# News this Week

Science  20 Dec 2002:
Vol. 298, Issue 5602, pp. 2296
1. BREAKTHROUGH OF THE YEAR

# Small RNAs Make Big Splash

1. Jennifer Couzin

## #1 The Winner

Just when scientists thought they had deciphered the roles played by the cell's leading actors, a familiar performer has turned up in a stunning variety of guises. RNA, long upstaged by its more glamorous sibling, DNA, is turning out to have star qualities of its own.

See Web links on RNA

For decades, RNA molecules were dismissed as little more than drones, taking orders from DNA and converting genetic information into proteins. But a string of recent discoveries indicates that a class of RNA molecules called small RNAs operate many of the cell's controls. They can turn the tables on DNA, shutting down genes or altering their levels of expression. Remarkably, in some species, truncated RNA molecules literally shape genomes, carving out chunks to keep and discarding others. There are even hints that certain small RNAs might help chart a cell's destiny by directing genes to turn on or off during development, which could have profound implications for coaxing cells to form one type of tissue or another. Science hails these electrifying discoveries, which are prompting biologists to overhaul their vision of the cell and its evolution, as 2002's Breakthrough of the Year.

These astonishing feats are performed by short stretches of RNA ranging in length from 21 to 28 nucleotides. Their role had gone unnoticed until recently, in part because researchers, focused on the familiar larger RNA molecules, tossed out the crucial small ones during experiments. As a result, RNA has long been viewed primarily as an essential but rather dull molecule that ferries the genetic code from the nucleus to the ribosomes, the cell's protein factories, and helps assemble amino acids in the correct order during protein synthesis.

Signs that RNA might be more versatile came in the early 1990s, when biologists determined that some small RNAs could quash the expression of various genes in plant and, later, animal cells. But they didn't appreciate the molecules' true powers until 1998. That's when Andrew Fire of the Carnegie Institution of Washington in Baltimore, Maryland, Craig Mello of the University of Massachusetts Medical School in Worcester, and their colleagues injected stretches of double-stranded RNA into worms. Double-stranded RNA forms when a familiar single strand kinks back in a hairpin bend, putting two complementary sequences alongside each other. To the researchers' surprise, double-stranded RNA dramatically inhibited genes that had helped generate the RNA in the first place. This inhibition, which was later seen in flies and other organisms, came to be known as RNA interference (RNAi). It helped prove that RNA molecules were behind some gene silencing.

Another crucial step came last year, when Gregory Hannon of Cold Spring Harbor Laboratory in New York and his colleagues identified an enzyme, appropriately dubbed Dicer, that generates the small RNA molecules by chopping double-stranded RNA into little pieces. These bits belong to one of two small RNA classes produced by different types of genes: microRNAs (miRNAs) and small interfering RNAs (siRNAs). SiRNAs are considered to be the main players in RNAi, although miRNAs, which inhibit translation of RNA into protein, were recently implicated in this machinery as well.

To bring about RNAi, small RNAs degrade the messenger RNA that transports a DNA sequence to the ribosome. Exactly how this degradation occurs isn't known, but scientists believe that Dicer delivers small RNAs to an enzyme complex called RISC, which uses the sequence in the small RNAs to identify and degrade messenger RNAs with a complementary sequence.

Such degradation ratchets down the expression of the gene into a protein. Although quashing expression might not sound particularly useful, biologists now believe that in plants, RNAi acts like a genome “immune system,” protecting against harmful DNA or viruses that could disrupt the genome. Similar hints were unearthed in animals this year. In labs studying gene function, RNAi is now commonly used in place of gene “knockouts”: Rather than delete a gene, a laborious process, double-stranded RNA is applied to ramp down its expression.

The year's most stunning revelations emerged in the fall, in four papers examining how RNA interference helps pilot a peculiar—and pervasive—genetic phenomenon known as epigenetics. Epigenetics refers to changes in gene expression that persist across at least one generation but are not caused by changes in the DNA code.

In recent years, researchers have found that one type of epigenetic regulation is caused by adjustments in the shape of complexes known as chromatin, the bundles of DNA and certain fundamental proteins that make up the chromosomes. By changing shape—becoming either more or less compact—chromatin can alter which genes are expressed. But what prompts this shape-shifting remained mysterious.

This year, scientists peering closely at RNAi in two different organisms were startled to find that small RNAs responsible for RNAi wield tremendous control over chromatin's form. In so doing, they can permanently shut down or delete sections of DNA by mechanisms not well understood, rather than just silencing them temporarily.

That news came from several independent groups. In one case, Shiv Grewal, Robert Martienssen, and their colleagues at Cold Spring Harbor Laboratory compared fission yeast cells lacking RNAi machinery with normal cells. When yeast cells divide, their chromosomes untangle and migrate to opposite sides of the cell. The researchers already knew, broadly, that this chapter of cell division is governed by a tightly wrapped bundle of chromatin, called heterochromatin, around the centromere—the DNA region at the chromosome's “waist.” The biologists found that their mutant cells, which were missing the usual small RNAs, couldn't properly form heterochromatin at their centromeres and at another DNA region in yeast that controls mating. This suggests that without small RNAs, cell division goes awry. The scientists theorized that in healthy yeast cells, small RNAs elbow their way into cell division, somehow nudging heterochromatin into position to do the job. That exposes DNA to different proteins and dampens gene expression.

Meanwhile, David Allis and his colleagues at the University of Virginia Health System in Charlottesville, along with Martin Gorovsky of the University of Rochester in New York and others, were focusing on a different organism, a single-celled ciliate called Tetrahymena. Biologists treasure Tetrahymena because it stores the DNA passed to offspring in a different nucleus from the one containing DNA expressed during its lifetime, making it easy to distinguish one gene set from the other. The researchers found that in Tetrahymena, small RNAs trigger deletion or reshuffling of some DNA sequences as a cell divides. RNAi appeared to be targeting structures analogous to heterochromatin, only this time strips of DNA were discarded or moved elsewhere. The mechanism remains unclear, however.

The two sets of experiments might help explain why small RNAs exist in the first place. In both the yeast and Tetrahymena, small RNAs' frenetic activity is focused on genome regions, such as centromeres, that contain repetitive DNA resulting from transposons. Transposons are bits of DNA that can jump around the genome and insert themselves at different locales; at times, they jam transcription machinery and cause disease. It appears possible—although still largely hypothetical—that small RNAs evolved very early in life's history to help protect the genome against instability.

This is just one of many areas that remain to be explored. Researchers are still trying to sort out how the well over 100 different miRNAs function and which species contain which ones. There are hints that they behave differently in plants and animals. And some recent work suggests that miRNAs exert more control over gene expression than previously believed. Also a focus of research are the proteins, such as Dicer, that are critical cogs in the RNAi machinery.

Researchers are also probing RNAi's possible role in development and disease. RNAi has been implicated in guiding meristems, the plant version of stem cells, so some biologists believe that it might help establish the path taken by human and other mammalian stem cells as they differentiate into certain tissues. If so, RNAi could prove an essential tool in manipulating stem cells. And if small RNAs influence cell division in humans as they do in yeast and Tetrahymena, minor disruptions in the machinery could lead to cancer.

The extraordinary, although still unfulfilled, promise of small RNAs and RNAi has split the field wide open and put RNA at center stage. Having exposed RNAs' hidden talents, scientists now hope to put them to work.

## Papers and Articles

Selected Research Papers

I. M. Hall et al., “Establishment and Maintenance of a Heterochromatin Domain,” Science 297, 2232 (2002)

K. Mochizuki et al., “Analysis of a piwi-Related Gene Implicates Small RNAs in Genome Rearrangement in Tetrahymena,” Cell 110, 689 (2002) [PubMed]

S. D. Taverna et al., “Methylation of Histone H3 at Lysine 9 Targets Programmed DNA Elimination in Tetrahymena,” Cell 110, 701 (2002) [PubMed]

B. J. Reinhart and D. P. Bartel, “Small RNAs Correspond to Centromere Heterochromatic Repeats,” Science 297, 1831 (2002)

T. A. Volpe et al., “Regulation of Heterochromatic Silencing and Histone H3 Lysine-9 Methylation by RNAi,” Science 297, 1833 (2002)

S. M. Elbashir et al., “Duplexes of 21-Nucleotide RNAs Mediate RNA Interference in Cultured Mammalian Cells,” Nature 411, 494 (2001) [PubMed]

M. Lagos-Quintana et al., “Identification of Novel Genes Coding for Small Expressed RNAs,” Science 294, 853 (2001)

N. C. Lau et al., “An Abundant Class of Tiny RNAs with Probable Regulatory Roles in Caenorhabditis elegans,” Science 294, 858 (2001)

Rosalind C. Lee and Victor Ambros, “An Extensive Class of Small RNAs in Caenorhabditis elegans,” Science 294, 862 (2001)

S. M. Hammond et al., “Argonaute2, a Link Between Genetic and Biochemical Analyses of RNAi,” Science 293, 1146 (2001)

E. Bernstein et al., “Role for a Bidentate Ribonuclease in the Initiation Step of RNA Interference,” Nature 409, 363 (2001) [PubMed]

A. Fire et al., “Potent and Specific Genetic Interference by Double-Stranded RNA in Canorhabditis elegans,” Nature 391, 806 (1998) [PubMed]

A. R. van der Krol et al., “Inhibition of Flower Pigmentation by Antisense CHS Genes: Promoter and Minimal Sequence Requirements for the Antisense Effect,” Plant Mol. Biol. 14, 457 (1990) [PubMed]

C. Napoli et al., “Introduction of a Chimeric Chalcone Synthetase Gene in Petunia Results in Reversible Cosuppression of Homologous Genes in trans,” Plant Cell 2, 279 (1990) [PubMed]

Reviews and Perspectives

T. Jenuwein, “An RNA-Guided Pathway to the Epigenome,” Science 297, 2215 (2002)

R. Allshire, “RNAi and Heterochromatin -- a Hushed-Up Affair,” Science 297, 1818 (2002)

R. H. A. Plasterk, “RNA Silencing: The Genome's Immune System,” Science 296, 1263 (2002)

P. D. Zamore, “Ancient Pathways Programmed by Small RNAs,” Science 296, 1265 (2002)

G. Ruvkun, “Glimpses of a Tiny RNA World,” Science 294, 797 (2001)

V. Ambros, “Dicing up RNAs,” Science 293, 811 (2001)

P. A. Sharp and P. D. Zamore, “RNA Interference,” Science 287, 2430 (2000)

Medical Applications

Here's a sample of recent articles highlighting the potential of RNAi-based therapeutics.

T. Tuschl and A. Borkhardt, “Small Interfering RNAs: A Revolutionary Tool for the Analysis of Gene Function and Gene Therapy,” Molecular Interventions 2, 158 (2002) [PDF file; posted at author's Web site]

M. Branca, “RNAi-Fever Heats Up Novel Drug Category Funding” (Bio-IT World, 9 October 2002)

S. Connor, “From Cancer to AIDS: The RNAi Revolution Is Gathering Pace” (The Independent, 6 September 2002)

M. Kitabwalla and R. M. Ruprecht, “RNA Interference-A New Weapon Against HIV and Beyond,” N. Engl. J. Med. 347, 1364 (2002) [PubMed]

K. Garber, “Prescription RNA,” Technology Review, December 2002/January 2003, p. 42

## Interesting World Website

Substantial collection of Web links to an array of RNA-related resources, from databases to tutorials; maintained by the Institute of Molecular Biotechnology, Jena.

Brief survey of historical development of RNAi science, from Ambion Inc.

A comprehensive list of papers, labs, and other resources focused on RNAi, provided by Orbigen, Inc.

Collection of Web sites of molecular biology groups interested in various aspects of RNA.

Interesting discussion of RNAi, on Web site developed to accompany textbook by S. F. Gilbert.

Catalog of results from RNAi phenotypic analysis of C. elegans genes.

Protocols for siRNA preparation for Drosophila knockout experiments, and links to commercial sources for siRNAs.

## Selected Labs

Robin Allshire (Wellcome Trust Center for Cell Biology)

Victor Ambros (Dartmouth Medical School)

David Bartel (MIT)

Martin A. Gorovsky (University of Rochester)

Ronald H. A. Plasterk (Hubrecht Laboratory)

Phil Sharp (MIT)

Thomas Tuschl (Max Planck Institute for Biophysical Chemistry)

Phillip D. Zamore (University of Massachusetts Medical School)

2. BREAKTHROUGH OF THE YEAR

# The Runners-Up

1. The News and Editorial Staffs

The News and Editorial Staffs

Science applauds discoveries ranging from the dawn of time to the dawn of our species.

#2 Neutrinos nailed. Neutrinos, mysterious and misunderstood, are finally getting the respect they deserve. For years, neutrinos were the terra incognita on the particle chart. Electrons, muons, taus, and quarks had all been analyzed for years, their properties measured and dissected. But neutrinos? Nobody knew even whether they had mass until a few years ago. They were essentially unknowns.

See Web links on neutrinos

No longer. In the last decade, physicists finally proved that neutrinos have mass, and since then, a flurry of experiments has begun to flesh out the elusive neutrinos' properties.

This year, the Sudbury Neutrino Observatory (SNO), a 1000-ton sphere of heavy water deep inside a nickel mine in Sudbury, Ontario, put the final nail in the coffin of the solar neutrino paradox. The nuclear reactions in the sun should produce a large number of electron neutrinos, but all observations had shown that only about one-third of the expected number were actually reaching Earth.

If neutrinos have mass, they can change flavors—from electron neutrinos into tau or mu neutrinos, for example—and that could explain the missing electron neutrinos. SNO showed, once and for all, that this is the case. In April, scientists at SNO announced that they had measured the abundances of all three types of neutrinos—electron, mu, and tau—by detecting when they split apart atoms of deuterium. When they added up the solar electron, mu, and tau neutrinos streaming through the detector, the total matched the number that should be created by nuclear reactions. Electron neutrinos change flavor during their journey to Earth.

As a bonus, the SNO measurements allowed scientists to drastically limit the “mixing angles” that define the neutrinos' flavor-changing abilities and, in December, the KamLAND experiment in Japan restricted the limits even further—with nuclear reactor- created antineutrinos instead of solar neutrinos. Although physicists still don't know how much neutrinos weigh, the evanescent beasties are no longer blank spots on the particle chart.

## Online Extras on Neutrinos

### Papers and Articles

G. A. McGregor [for the SNO Collaboration], “First Results from the Sudbury Neutrino Observatory,” [arXiv.org preprint server]

SNO Collaboration, “Direct Evidence for Neutrino Flavor Transformation from Neutral-Current Interactions in the Sudbury Neutrino Observatory,” Phys. Rev. Lett. 89, 011301 (2002) [arXiv.org preprint server]

SNO Collaboration, “Measurement of Day and Night Neutrino Energy Spectra at SNO and Constraints on Neutrino Mixing Parameters,” Phys. Rev. Lett. 89, 011302 (2002) [arXiv.org preprint server]

KamLAND Collaboration, “First Results from KamLAND: Evidence for Reactor Anti-Neutrino Disappearance” (2002) (PDF, 1.2 MB) [preprint; from KamLAND Web site]

C. Seife, “Neutrino Census Nails Chameleon Particles,” Science 296, 632 (2002)

C. Seife, “Particle Trap Confirms Antimatter Shuffle,” Science 298, 2107 (2002)

C. Seife, “Neutrino Traps and X-ray Eyes,” Science 298, 527 (2002)

Two of the three 2002 Physics Nobel laureates shared the prize for early work in neutrino detection.

### Interesting Web Sites

SNO site includes an excellent description of neutrinos and the solar neutrino problem, images of SNO events, and details about the SNO detector itself.

Official Japanese site of the KamLAND project (in English and Japanese); includes link to CERN article providing background on the project, as well as other resources.

Excellent resource on KamLAND project (and on neutrino issues generally), from Lawrence Berkeley Laboratory. Includes links to KamLAND sites of other collaborating institutions.

Particularly nice collection of online resources on neutrino science.

#3 Genomes head south. This year's DNA sequencing efforts should prove to be a boon for the developing world. Two international consortia took a stab at malaria—which kills 3 million people a year, primarily in tropical Africa—by deciphering the genome sequences of both the parasite and the mosquito responsible for transmitting most of the deadliest cases. With a highly organized draft of 278 million bases of the mosquito Anopheles gambiae genome and the genome sequence of the parasite Plasmodium falciparum now in hand, biomedical researchers hope to find better ways to fight this devastating tropical disease.

See Web links on genomes

Developing countries should also benefit from rapid progress toward sequencing the rice genome. In April, a private company and a Chinese group independently published draft sequences of the japonica and indica rice strains consumed in Japan and China, respectively. Just this week, the International Rice Genome Sequencing Project released a more highly polished draft sequence of japonica, two chromosomes of which are now published. Originally, this publicly funded effort planned to finish the sequence by 2008, but two companies have made their sequences available to the international group, allowing it to move up the target date for completing the work to 2005.

This past year, sequencers got detailed looks at several other large genome sequences as well. A U.S.-British team has completed and analyzed a high-quality draft sequence of the mouse genome; a U.S. public-private partnership has assembled a draft for the rat. Nailing these two genome sequences will cover two of the most important research animals. Researchers now know the order of the DNA bases in the genome of the Japanese puffer fish, which has the smallest known genome of all vertebrates. The DNA of two tunicates—sessile invertebrates with vertebrate-like larvae—has been deciphered as well. These sequences should provide clues about how vertebrates evolved.

In addition, the sequences of microbial genomes keep pouring in: another strain of anthrax and a bacterium called Shewanella, useful for bioremediation, to name just two. And the flood of genome sequences shows no sign of abating: Work has begun on chimp, corn, and poplar. The honey bee, dog, cow, chicken, and sea urchin are supposed to be up next.

## Online Extras on Genomes

### Papers and Articles

M. J. Gardner et al., “Genome Sequence of the Human Malaria Parasite Plasmodium falciparum Nature 419, 498 (2002).

Home page of Nature Plasmodium genome special issue (3 October 2002), with links to related papers, news, and commentary

R. A. Holt et al. “The Genome Sequence of the Malaria Mosquito Anopheles gambiae,” Science 298, 129 (2002)

Home page of Science Anopheles genome special issue (4 October 2002), with links to related papers, news, and commentary

J. Yu et al., “A Draft Sequence of the Rice Genome (Oryza sativa L. ssp. indica),” Science, 296, 79 (2002)

S. A. Goff et al., “A Draft Sequence of the Rice Genome (Oryza sativa L. ssp. japonica),” Science, 296, 92 (2002)

Home page of Science rice genome special issue (4 October 2002), with links to related papers, news, and commentary

T. Sasaki et al., “The Genome Sequence and Structure of Rice Chromosome 1,” Nature 420, 312 (2002) [PubMed]

Q. Feng et al., “Sequence and Analysis of Rice Chromosome 4,” Nature 420, 316 (2002) [PubMed]

Mouse Genome Sequencing Consortium, “Initial Sequencing and Comparative Analysis of the Mouse Genome,” Nature 420, 520 (2002)

S. Aparicio et al., “Whole-Genome Shotgun Assembly and Analysis of the Genome of Fugu rubripes,” Science 297, 1301 (2002)

P. Dehal et al., “The Draft Genome of Ciona intestinalis: Insights into Chordate and Vertebrate Origins,” Science 298, 2157 (2002)

### Interesting Web Sites

Collection of Web resources on Plasmodium and Anopheles, from Science's Functional Genomics Web site.

Includes information on rice functional genomics and bioinformatics, as well as access to IRIS (International Rice Information System).

Educational site promising “Everything you want to know about one of the world's longest running cereals.”

Rich collection of links on Mus musculus.

Regularly updated page from the Genome News Network of The Institute for Genomic Research (TIGR), providing brief descriptions and links for a comprehensive list of sequenced organisms.

Remarkably comprehensive and up-to-date compendium of information on complete and ongoing genome projects around the world.

#4 Cosmic twist. Even though it's less than 3° above absolute zero, the cosmic microwave background (CMB) is very, very hot. In 2002, astronomers and physicists watched the end of the tale of the beginning of the universe.

Discovered in 1965 by Arno Penzias and Robert Wilson of Bell Laboratories in New Jersey, the CMB is the remnant of a time, 400,000 years after the big bang, when free-streaming nuclei and electrons finally cooled and formed atoms. As the electrons settled down into their orbitals, high-energy light, liberated from its cage of matter, streamed forth. Stretched and attenuated by 14 billion years of travel, the CMB appears as a faint but ubiquitous microwave static coming from all regions of the sky.

See Web links on the CMB

In 2000 and 2001, airborne and ground-based microwave telescopes generated exquisitely detailed pictures of fluctuations in the CMB, fluctuations that reveal not only the universe's past but its future. Not only did these fluctuations give rise to the agglomerations of galaxies that we see today, but they also revealed the “curvature” of the universe, which shows that the universe will expand forever, rather than recollapsing in a big crunch. In May, the Cosmic Background Imager (CBI), a microwave telescope high atop the Andes mountains in Chile, put an exclamation point on those observations by detecting “peaks”—characteristic patterns in the fluctuations—that revealed structures far smaller than other telescopes had yet seen. In January 2003, other physicists are expected to reveal the first results from the Microwave Anisotropy Probe (MAP), an orbiting satellite that will be the ne plus ultra of fluctuation detection until the end of the decade.

But the real CMB triumph this year was the first detection of polarization by the Degree Angular-Scale Interferometer team at the University of Chicago. The discovery of this faint signal heralds the beginning of a new chapter in CMB research—one that might reveal the state of the universe when it was a minuscule fraction of a second old by revealing the subtle scarring caused by gravitational waves during the birth of the cosmos.

## Online Extras on the CMB

### Papers and Articles

B. S. Mason et al., “The Anisotropy of the Microwave Background to l = 3500: Deep Field Observations with the Cosmic Background Imager,” astro-ph/0205384 (2002) [arXiv.org preprint server]

T. J. Pearson et al., “The Anisotropy of the Microwave Background to l = 3500: Mosaic Observations with the Cosmic Background Imager,” astro-ph/0205388 (2002) [arXiv.org preprint server]

J. Kovac et al., “Detection of Polarization in the Cosmic Microwave Background Using DASI,” astro-ph/0209478 (2002) [arXiv.org preprint server]

E. Hivon and M. Kamionkowski, “A New Window to the Early Universe,” Science 298, 1349 (2002)

Perspective article on the implications of the DASI results.

C. Seife, “Subtle Signals in Ancient Light Promise New View of Cosmos,” Science 297, 2184 (2002)

C. Seife, “Best Big Bang Pictures Show New Wrinkles,” Science 296, 1588 (2002)

M. Tegmark, “Measuring Spacetime: From the Big Bang to Black Holes,” Science 296, 1427 (2002)

Review article, from Science's 24 May 2002 Spacetime Special Issue, surveys measurements of spacetime parameters, including the CMB, over 22 orders of magnitude.

### Interesting Web Sites

Includes information on the instrument, a collection of papers, and more.

DASI site features link to streaming video of the press conference on CMB polarization.

Web headquarters for NASA mission offers Cosmology 101, a rich and informative introduction.

Home site of the European Space Agency microwave background mission (slated to launch in 2007) includes a wealth of information on the CMB.

Impressive collection of online CMB resources and tutorials, from University of Chicago Center for Cosmological Physics.

#5 Fast moves. If action flicks seem to be getting faster these days, just wait. This year, laser physicists succeeded in making the first-ever movies in which individual frames were measured in attoseconds, or billionths of a billionth of a second. The new high-speed filmmaking techniques are expected to spawn a new genre of cinema devoted to tracking the motion of electrons around atoms.

See Web links on attosecond physics

Laser physicists have been refining their high-speed moviemaking approaches for years. But most rely on the same basic principle, using ultrashort pulses of laser light like bursts from a strobe light to freeze motion in flight. Researchers now routinely use the technique to capture the blur of molecules as they break and weld bonds in a chemical reaction, events that take place on the order of 1 to 100 femtoseconds, or 10−15 seconds.

Dutch and French researchers broke the attosecond barrier last year, when they trained ultrashort laser pulses on a gas of argon atoms, which in turn emitted a train of pulses, each lasting just 220 attoseconds. A team of Austrian, Canadian, and German researchers followed hard on their heels with a related technique that turned out individual 650-attosecond pulses, which are more easily used as moviemaking strobes.

This year, researchers turned their new attosecond strobes onto the action within atoms. In October, the Austrian and German members of the original team used their attosecond pulses to excite electrons in krypton atoms, each of which left behind an electron vacancy. With another laser pulse, they were then able to track the timing with which excited electrons gave up some of their energy and fell back into the more stable energy levels. It's not Hitchcock, but attosecond movies will give physicists a whole new view of life inside the atom.

## Online Extras on Attosecond Physics

### Papers and Articles

M. Drescher et al., “ Time-Resolved Atomic Inner-Shell Spectroscopy,” Nature 419, 803 (2002) [PubMed]

A. Cho, “X-ray Flashes Provide Peek Into Atom Core,” Science 298, 727 (2002)

R. Kienberger et al., “Steering Attosecond Electron Wave Packets with Light,” Science 297, 1144 (2002)

M. Lewenstein, “Resolving Physical Processes on the Attosecond Time Scale,” Science 297, 1131 (2002)

Perspective article on midyear developments in attophysics.

F. Krausz, “Tracking Light Oscillations: Attosecond Spectroscopy Comes of Age,” Opt. Photon. News 13 (5), 62 (2002) [PDF, posted on author's Web site]

Review article on potential for time-resolved spectroscopy in the sub-femtosecond domain.

Y. Bhattacharjee, “Photoelectrons Show How Quick a Flash Is,” Science 294, 1805 (2001)

R. F. Service, “Strobe Light Breaks the Attosecond Barrier,” Science 292, 1627 (2001)

### Interesting Web Sites

Photonics Institute, Vienna University of Technology

Research page of attophysics group at Bielefeld University.

News, resources, and features on photonics and optics.

#6 A taste for temperature. The heat of four-alarm chili and the coolness of spearmint chewing gum aren't just metaphorical: To some cells, taste and temperature are the same. This year, researchers tunneled into ion channels that respond to such sensations. They're tuned to warmth, minty coolness, or, in mice at least, another steamy stimulus: pheromones.

See Web links on TRP channels

So-called transient receptor potential (TRP) ion channels are proteins that snake in and out of the cell membrane. When they're tickled appropriately, they allow calcium or other ions to surge into a cell. In neurons, this can make the cell fire off a signal to its neighbors. Mammals harbor at least 21 flavors of TRP channels, but most of their functions are unknown.

The first report that certain TRP channels promiscuously respond to either a chemical or a thermal stimulus came in 1997, with the identification of a TRP channel that gets steamed up by either hot temperatures (above 43°C) or capsaicin, the active ingredient in chili peppers. This year, a similar multitasking channel was found in nerves of the mouth and skin. It reacts when exposed to either cool temperatures (15° to 25°C) or menthol, the chemical that makes mint minty.

A warmth-sensitive (∼34°C) TRP channel debuted this year as well. It is concentrated in skin cells, suggesting that the skin itself senses heat and passes the message to neurons.

TRP channels are also necessary for another type of body heat. Male mice lacking a certain TRP channel are particularly thickheaded when it comes to mating, seemingly unable to distinguish between females and fellow males. These TRP channels inhabit a part of the nose that sniffs out pheromones.

Mutations in other TRP channels are responsible for certain cancers and other diseases, including, as discovered this year, an inability to regulate magnesium levels. These and most other TRP functions are still poorly understood, but researchers are picking up more coherent signals from TRP channels that buzz to sensations of taste, heat, and pheromones.

## Online Extras on TRP Channels

### Papers and Articles

L. Stowers et al., “Loss of Sex Discrimination and Male-Male Aggression in Mice Deficient for TRP2,” Science 295, 1493 (2002)

M. Beckman, “When in Doubt, Mice Mate Rather Than Hate,” Science 295, 2228 (2002)

News report on the Stowers et al. Science paper.

A. M. Peier et al., “A TRP Channel That Senses Cold Stimuli and Menthol,” Cell 108, 705 (2002) [PubMed]

D. D. McKemy, W. M. Neuhausser, D. Julius, “Identification of a Cold Receptor Reveals a General Role for TRP Channels in Thermosensation,” Nature 416, 52 (2002) [PubMed]

H. Xu et al., “TRPV3 is a Calcium-Permeable Temperature-Sensitive Cation Channel,” Nature 418, 181 (2002) [PubMed]

A. M. Peier et al., “A Heat-Sensitive TRP Channel Expressed in Keratinocytes,” Science 296, 2046 (2002)

D. E. Clapham, “Hot and Cold TRP Ion Channels,” Science 295, 2228 (2002)

Perspective article on role of TRP proteins in sensations of heat and cold.

C. Montell, “Physiology, Phylogeny, and Functions of the TRP Superfamily of Cation Channels,” Science's STKE (10 July 2001)

Comprehensive review of TRP channels, on Science's Signal Transduction Knowledge Environment (subscription required for full-text access).

### Interesting Web Sites

Includes an extensive list of TRP families, with individual links to SWISS-PROT.

Collection of information sheets on individual human TRP genes.

Provides a list of resources containing relevant information about ion channels.

#7 Frozen images. Thirty years ago, researchers pitched the idea of reconstructing a 3D picture from electron micrographs. Today, cryoelectron tomography (cryo-ET) has overcome a series of technical obstacles to emerge as a breakthrough technique for viewing structures inside intact cells.

Biologists have long been able to capture the molecular structure of single proteins in cells, using techniques such as x-ray crystallography. But they haven't had a good way to get a 3D look at midsize organelles (∼5 nm), such as the protein-packaging Golgi apparatus or energy-producing mitochondria—especially without removing them from their native environment. Cryo-ET fills this resolution gap and gives scientists a way to link atomic-level detail to whole-cell organization.

See Web links on cryo-ET

Cryo-ET works something like a doctor's computerized tomography scan. Penetrating beams of electrons create two-dimensional image slices that a computer assembles into a 3D image. Cells are flash-frozen and do not need to be fixed or have their membranes disrupted. (For years, the problem with cryo-ET has been that too much radiation causes structures to degrade.)

Long, steady progress has solved many of the early snags. Autorotation of the specimen through a range of imaging angles and better calibration of the microscope stage have dramatically reduced exposure time. Improved clarity by reduced scattering of the electrons allows thicker specimens to be viewed. Advances in cryosectioning, slicing up the specimen in layers, have also enabled this technique to be used with thicker samples.

This year, cell imagers used cryo-ET to catch the first glimpse of actin filaments in the act, braced against the edge of the cell membrane. They also captured the first view of spatial arrangement of tubules and receptors in the sarcoplasmic reticulum, the components responsible for the chemical cascade that sets off a muscle contraction. And efforts are currently under way to create the first detailed 3D map of the spatial relationship of all the organelles in a eukaryotic cell.

## Online Extras on Cryo-ET

### Papers and Articles

O. Medalia et al., “Macromolecular Architecture in Eukaryotic Cells Visualized by Cryoelectron Tomography,” Science 298, 1209 (2002)

E. Goldman, “A New Window on the Cell's Inner Workings,” Science 298, 1155 (8 November 2002)

News article on the Medalia et al. result.

T. Wagenknecht et al., “Electron Tomography of Frozen-Hydrated Isolated Triad Junctions,” Biophys. J. 83, 2491 (2002) [PubMed]

B. F. McEwen et al., “Use of Frozen-Hydrated Axonemes to Assess Imaging Parameters and Resolution Limits in Cryoelectron Tomography,” J. Struct. Biol. 138, 47 (2002) [PubMed]

U. Ziese et al., “Automated High-Throughput Electron Tomography by Pre-Calibration of Image Shifts,” J. Microscopy 205, 187 (2002)

J. M. Plitzko, “In Vivo Veritas: Electron Cryotomography of Cells,” Trends. Biotechnol. 20 (Suppl.), S40 (2002) [PubMed]

M. L. Harlow et al., “The Architecture of Active Zone Material at the Frog's Neuromuscular Junction,” Nature 409, 479 (2001) [PubMed]

M. Auer, “Three-Dimensional Electron Cryo-Microscopy as a Powerful Structural Tool in Molecular Medicine,” J. Mol. Med. 78, 191 (2000) [PDF; posted on author's Web site]

R. G. Hart, “Electron Microscopy of Unstained Biological Material: The Polytropic Montage,” Science 159, 1464 (1968)

Early paper that introduced the technique; available to AAAS Members through the JSTOR portal of the AAASMember.org Web site.

### Interesting Web Sites

Describes the Institute's automated 3D electron tomography and energy-filtering capabilities.

National Center for Research Resources-funded lab develops new technologies for analyzing 3D structure of cells and tissues; site includes links to electron tomography movies from recent papers.

Comprises several state-of-the-art integrated microscope and computing components; site includes informative pages on visualization techniques.

Develops technology to improve and automate advanced imaging systems and for Web-based collaborative research in electron tomography.

#8 Clear skies ahead. This year, astronomers converted the promise of adaptive optics (AO) into crisp new views of the heavens. AO systems erase the blurring of Earth's atmosphere by flexing the surfaces of thin mirrors hundreds of times each second, precisely canceling the turbulence overhead. This optical wizardry is easier said than done—it took years of painstaking engineering to make AO work routinely at the world's biggest telescopes.

The wait was worth it. Both the W. M. Keck Observatory in Hawaii, with its twin 10-meter telescopes, and the European Southern Observatory's Very Large Telescope array of four 8.2-meter telescopes in Chile used AO this year to peer at the heart of our Milky Way. Sharp images of the central stars dashing around a hidden body gave the best evidence yet of a supermassive black hole. Other striking AO images included a huge volcanic blast on Jupiter's moon Io and new details about the shapes of distant galaxies. Even a daytime observatory joined the club. A Swedish solar telescope on the Canary Islands snapped the clearest pictures of the sun's surface with a new AO system, revealing dark ribbons of seething magnetic fields around sunspots.

These studies require a bright star or planet, providing enough light for the AO sensors to gauge the air's ripplings. However, lasers mounted on the sides of telescopes can create “artificial” stars high in the atmosphere, making it possible to clarify vision anywhere in the sky. Astronomers showed this year that laser AO works well on smaller telescopes; the big eyes on the sky should have laser systems within a year.

## Online Extras on Adaptive Optics

### Papers and Articles

S. Gezari et al., “Adaptive Optics Near-Infrared Spectroscopy of the Sagittarius A* Cluster,” astro-ph/0205186 (2002) [arXiv.org preprint server]

F. Marchis et al., “High-Resolution Keck Adaptive Optics Imaging of Violent Volcanic Activity on Io,” (2002) [preprint of in-press Icarus article; on author's Web site]

S. Gezari et al., “A Keck Adaptive Optics Search for Young Extrasolar Planets,” astro-ph/0210169 (2002) [arXiv.org preprint server]

G. Schilling, “Latest Observations Bring the Unseen Into View,” Science 295, 616 (2002)

A. Lawler, “Glimpsing the Post-Hubble Universe,” Science 295, 1448 (2002)

R. Angel and B. Fugate, “Adaptive Optics,” Science 288, 455 (2000)

Primer on the principles of adaptive optics, in Science's Tech.Sight section.

### Interesting Web Sites

Web site of NSF-funded center at the University of California, Santa Cruz, includes a thorough introduction to the principles of AO, images comparing observations with and without AO, a substantial collection of AO links, and more.

Organization in charge of the Very Large Telescope offers information about adaptive optics.

#9 Retina receptors. Researchers hit the jackpot this year in understanding how light resets the circadian clock, our internal timepiece that regulates daily patterns of behavior and physiology. After years of searching hard for the so-called photoreceptor cells that relay that light signal to the clock in mammals, circadian biologists had tantalizing clues but no answer. They knew the photoreceptors must be in the eye. But the eyes' only known photosensitive cells, the rods and cones, weren't doing the job. Then last winter, five independent research teams discovered a brand-new class of light-responsive cells in the mammalian retina that connect directly to the brain's clock.

See Web links on melanopsin

First, researchers found a pigment called melanopsin in a small subset of retinal ganglion cells (RGCs) in the eyes of rats. Most RGCs don't respond to light, but it turned out that the melanopsin-containing ones do, making them a brand-new class of previously unknown light-responsive retinal cells. What's more, researchers traced their connections and found that they hook up directly to the suprachiasmatic nucleus, the brain area that houses the clock.

That's not all. More recent neuroanatomy studies have shown that the melanopsin-containing RGCs also link up to brain areas that control a variety of responses to light that don't require the image-forming visual system, such as constriction of the pupils and the direct effect of light on sleep-wake state—what makes us drowsy in dark seminar rooms or wakeful if the lights are kept on all night.

Most of the pieces have fallen neatly into place, but technical difficulties prevented researchers from proving that melanopsin responds chemically to light. Without that evidence, some were reluctant to accept it as the RGCs' light-capturing pigment. Now that issue has been put to rest: In last week's issue of Science (13 December, pp. 2211 and 2213), researchers showed that mice that lack melanopsin do not normally reset their circadian clocks in response to light, suggesting that melanopsin is capturing and relaying the light signal.

## Online Extras on Melanopsin

### Papers and Articles

J. J. Gooley et al., “Melanopsin in Cells of Origin of the Retinohypothalamic Tract,” Nature Neurosci. 4, 1165 (2001) [PubMed]

J. Hannibal et al., “The Photopigment Melanopsin Is Exclusively Present in Pituitary Adenylate Cyclase-Activating Polypeptide-Containing Retinal Ganglion Cells of the Retinohypothalamic Tract,” J. Neurosci. 22, RC191 (2002) [PubMed]

I. Provencio et al., “Photoreceptive Net in the Mammalian Retina,” Nature 415, 493 (2002) [PubMed]

D. M. Berson et al., “Phototransduction by Retinal Ganglion Cells that Set the Circadian Clock,” Science 295, 1070 (2002)

S. Hattar et al., “Melanopsin-Containing Retinal Ganglion Cells: Architecture, Projections, and Intrinsic Photosensitivity,” Science 295, 1065 (2002)

N. F. Ruby et al., “Role of Melanopsin in Circadian Responses to Light,” Science 298, 2211 (2002)

S. Panda et al., “Melanopsin (Opn4) Requirement for Normal Light-Induced Circadian Phase Shifting,” Science 298, 2213 (2002)

M. Barinaga, “How the Brain's Clock Gets Daily Enlightenment,” Science 295, 955 (2002)

M. P. Pando and P. Sassone-Corsi, “Signaling to the Mammalian Circadian Clocks: In Pursuit of the Primary Mammalian Circadian Photoreceptor,” Science's STKE (6 November 2001; subscription required for full-text access)

### Interesting Web Sites

“Virtual museum exhibit” on HHMI's superb BioInteractive educational site provides an engaging introduction to the study of biological rhythms.

Site of HHMI Holiday Lectures for 2000 features four Webcasts on aspects of biological clocks.

General discussion of how the biological clock is set and maintained in various organisms (including mammals).

Introductions, tutorials, and Web links from the University of Warwick Department of Biology.

#10 Evolutionary headlines. Only a decade ago, the earliest known human ancestor was a species whose most famous member, Lucy, lived in east Africa about 3.2 million years ago. But in July, the nearly complete cranium of a primate that lived twice as long ago—between 6 million and 7 million years ago—was introduced as the oldest known hominid, the lineage that includes humans but not other apes. This fossil, found by a team of French and Chadian researchers, fills a crucial gap at the dawn of human evolution when almost nothing is known; the next oldest published hominid skull is almost 3 million years younger.

See Web links on Toumaï

It also is important because it was found in an unexpected place: along the shores of the ancient Lake Chad in western Africa. Until now, the earliest ancestors of the human family were found in east Africa, which has been called the cradle of humanity.

The fossil, nicknamed Toumaï for “hope of life” in the Goran language, shows that the earliest hominids were more widely distributed across Africa than previously thought, and it challenges old views about where the first hominid arose.

At this early age, Toumaï looks most like an ancient ape, with a brain the size of a chimpanzee's, large incisors, and widely spaced eyes like those of a gorilla. But the shape and size of its canines and lower face resemble those of human ancestors that came later; it has small, unsharpened canines and a flat lower face, unlike the protruding snout of living apes. The mix of features convinced the fossils' discoverers that they had found a new genus and species of hominid, which they named Sahelanthropus tchadensis.

Controversy is another prominent feature of these fossils. A competing team of researchers (who have discovered another, slightly younger fossil that they say is the earliest hominid) argues that Toumaï is the ancestor of an extinct ape or gorilla, partly because there are no skeletal bones to show whether it walked upright—the hallmark of being a hominid. Others who have seen the skull, however, disagree. Although detailed analysis has just begun, they say that on the face of it, Toumaï looks like a hominid.

## Online Extras on Toumaï

### Papers and Articles

M. Brunet et al., “A New Hominid from the Upper Miocene of Chad, Central Africa,” Nature 418, 145 (2002) [PubMed]

P. Vignaud et al., “Geology and Palaeontology of the Upper Miocene Toros-Menalla Hominid Locality, Chad,” Nature 418, 152 (2002) [PubMed]

Comment and Reply: “Sahelanthropus or 'Sahelpithecus'?,” Nature 419, 581 (2002)

Comment: M. H. Wolpoff et al.

Response: M. Brunet

A. Gibbons, “First Member of Human Family Uncovered,” Science 297, 171 (2002)

A. Gibbons, “One Scientist's Quest for the Origin of Our Species,” Science 298, 1708 (2002)

A profile of Michel Brunet, discoverer of the Chad fossil.

A. Gibbons, “In Search of the First Hominids,” Science 295, 1214 (2002)

### Interesting Web Sites

Web site on the Chad discovery from French National Center for Scientific Research.

In-depth, up-to-date discussions of hominid species, fossils, type specimens, and more (including Tomaï), from the Talk.Origins Archive.

Superb site from the Arizona State University Institute of Human Origins includes paleoanthropology news, a Flash-driven “documentary,” and copious links and educational resources.

Searchable portal of anthropology resources.

3. BREAKTHROUGH OF THE YEAR

# Areas to Watch in 2003

Science's editors use their powers of prognostication to come up with next year's hot research topics.

Whither the ice? Glaciologists are scrambling to sort out which of the world's ice houses may be about to empty themselves under the onslaught of greenhouse warming. Mountain glaciers are clearly receding, and high-mountain tropical glaciers could soon disappear. But the behavior of the great ice stores of Greenland, Antarctica, and West Antarctica is proving more subtle. Satellite-borne radar and other new geophysical tools will be monitoring the comings and goings of ice in these constantly shifting sheets, providing a better understanding of what our warmer future holds.

A sun-climate connection. As more and more wiggles matching the waxing and waning of the sun show up in records of past climate, researchers are grudgingly taking the sun seriously as a factor in climate change. They have included solar variability in their simulations of the past century's warming. And the sun seems to have played a pivotal role in triggering droughts and cold snaps. To gain complete respectability, sun-climate researchers are working to identify the physical link between relatively feeble solar fluctuations and climate. A leading candidate: solar-modulated cosmic rays and their effects on clouds.

See Web links on areas to watch

Budget bust. Will 2002 be remembered as the year the good times ended? That's likely to be a little too dire, but it's a growing worry among scientists in developed nations, as a slumping world economy could dramatically slow the growth of government and private spending on basic science. Italy, Germany, and France are already facing cuts or freezes in government spending. In the United States, cratering stock prices have shrunk university and foundation endowments by one-third or more. The White House has already signaled that it won't support continued double-digit increases for biomedical research spending. And war with Iraq could quash growing hopes of doubling taxpayer outlays on the physical sciences. But there is a bright side: Low interest rates are allowing stretched institutions to keep many lab construction projects on track.

R-evolutionary genomics. With genome sequences for most of the major microbial groups in hand and ever more DNA of complex organisms being deciphered, researchers expect to be able to make better sense of life's many evolutionary relationships. Meanwhile, studies of human genetic variation will continue to shed light on our deep past, and the chimp genome project may begin to reveal what makes us human.

A different light. Several satellites tuned to wavelengths outside the glamorous optical band should shine in 2003. The European Space Agency's Integral mission, launched in October, will soon observe gamma rays from black holes, supernovas, and other scenes of violence, and NASA's Swift explorer will start tracking gamma ray bursts by December. The Space Infrared Telescope Facility, slated for launch in April, will examine the heat from distant galaxies and dusty clouds where stars and planets form. And astronomers will get their best map of microwave ripples in the sky—a chilly imprint of the big bang—when results from the Microwave Anisotropy Probe are released early in the year.

Important matter. In 2002, two rival teams at the CERN laboratory near Geneva produced cold, slow-moving antihydrogen atoms—antielectrons orbiting antiprotons—for the first time. Antihydrogen will be a powerful tool for studying the difference between matter and antimatter, but scientists have to trap significant amounts of it before they can zap it with a laser and measure its properties. It might not happen in the coming year or even in the next, but there's no question that the game is afoot. Antihydrogen futures are brighter than ever.

## Online Extras on Areas to Watch

### Disappearing Glaciers

Science Special Issue: Polar Science, 30 August 2002

Special issue content includes news and review articles on impacts of recent climate change in polar regions.

“Antarctic Ice Shelf Collapses,”, Science295, 2359 (2002)

Site includes informative “topic sheets” about climate change and the Antarctic ice sheet.

Center's State of the Cryosphere site provides an overview of snow and ice as indicators of climate change.

### Sun-Climate Connection

D. Rind, “The Sun's Role in Climate Variations,” Science 296, 673 (2002)

K. S. Carslaw et al., “Cosmic Rays, Clouds, and Climate,” Science 298, 1732 (2002)

S. K. Solanki, “Solar Variability and Climate Change: Is There a Link?” (2002) [PDF]

Review of debate regarding the sun's influence on Earth climate, posted at the Royal Astronomical Society Web site.

Well-known NASA site includes extensive links on climate and sun-earth interactions.

Electronic workspace for scientists from the Collaboratory for Atmospheric Science and Technology of the British Atmospheric Data Centre. Includes a public-access area.

### U.S. Science Budget

D. Malakoff, “GOP Takes Senate, Budget Uncertain,” Science 298, 1313 (2002)

D. Malakoff, “News Focus: 2003 Budget -- War Effort Shapes U.S. Budget, With Some Program Casualties,” Science 295, 952 (2002)

### Genomes and Evolution

E. Pennisi, “Comparative Biology Joins the Molecular Age,” Science 296, 1792 (2002)

J. Couzin, “NSF's Ark Draws Alligators, Algae, and Wasps,” Science 297, 1638 (2002)

S. Olson, “Seeking the Signs of Selection,” Science 298, 1324 (2002)

Ambitious Web project providing information about evolutionary relationships (including molecular ones) among Earth's diverse organisms.

Comprehensive, up-to-date compendium of information on complete and ongoing genome projects around the world.

Chimpanzee Genomics Page at Baylor College of Medicine

### Gamma-Ray, Infrared, and Microwave Astronomy

G. F. Bignami, “Gamma-Ray Astronomy with INTEGRAL,” Science 298, 1560 (2002)

R. Irion, “Sensing the Hidden Heat of the Universe,” Science 298, 1870 (2002)

Web site accompanying Science's Star Formation special issue of 4 January 2002 includes links to many ground and spaceborne telescope projects.

### Antihydrogen

M. Amoretti et al., “Production and Detection of Cold Antihydrogen Atoms,” Nature 419, 456 (2002) [PubMed]

C. Seife, “CERN Team Produces Antimatter in Bulk,” Science 297, 1979 (2002)

C. Seife, “Antihydrogen Rivals Enter the Stretch,” Science 298, 1327 (2002)

Web site of the CERN project that first captured antihydrogen in bulk.

Outstanding CERN educational site including pages on the science of antimatter and Webcasts on the theoretical and technical challenges of making antimatter.

Another CERN site, featuring QuickTime movies and information on the ATRAP antihydrogen-trapping program.

4. BREAKTHROUGH OF THE YEAR

# Scorecard 2002

In which we take our lumps for predictions made last year

## Online Extras on Last Year's Picks

### Stem Cells

News Focus: Stem Cell Lines, Science 297, 923 (2002)

C. Holden and G. Vogel, “'Show Us the Cells,' U.S. Researchers Say”

C. Holden and G. Vogel, “Plasticity: Time for a Reappraisal?,”, Science 296, 2126 (2002)

B. Vogelstein et al., “Please Don't Call It Cloning!,” Science 295, 1237 (2002)

J. G. Reich, “Embryonic Stem Cells: The Debate in Germany,” Science 296, 265 (2002)

Science 2002 Policy Forum articles on aspects of the international controversy regarding embryonic stem cells.

Overview on stem cell issues from AAAS Center for Science, Technology, and Congress.

### Proteomics

J. Kaiser, “Proteomics: Public-Private Group Maps Out Initiatives,” Science 296, 827 (2002)

Science Special Issue: Systems Biology (1 March 2002)

Issue contained a review articles on biological networks, including protein-protein interactions.

Interesting Weblog/portal to proteomics resources.

### Astronomical Viewing

Includes an “Image of the Week,” as well as a wealth of other information on the project.

Web site accompanying Science's Star Formation special issue of 4 January 2002 includes links on sky-survey programs, brown dwarfs, and globular clusters, among many other things.

Well-curated collection of links on virtual observatory projects.

### Genes and Disease

J. Couzin, “New Mapping Project Splits the Community,” Science 296, 1391 (2002)

J. Couzin, “HapMap Launched With Pledges of $100 Million,” Science 298, 941 (2002) Science Special Issue: The Puzzle of Complex Diseases (26 April 2002) News, Viewpoints, and Review on diseases that resist attribution to a single genetic or environmental “smoking gun.” ### Optical Clocks Web site of this standard-setting and research group offers a variety of interesting links and a time-and-frequency glossary. ### Visualization M. N. Teruel and T. Meyer, “Parallel Single-Cell Monitoring of Receptor-Triggered Membrane Translocation of a Calcium-Sensing Protein Module,” Science 295, 1910 (2002) V. Marx, “Beautiful Bioimages for the Eyes of Many Beholders,” Science 297, 39 (2002) D. Clery, “Brainstorming Their Way to an Imaging Revolution,” Science 297, 761 (2002) S. Paddock, “Optical Sectioning--Slices of Life,” Science 295, 1319 (2002) M. Chicurel, “Cell Migration Research Is on the Move,” Science 295, 606 (2002) U. H. von Andrian, “T Cell Activation in Six Dimensions,” Science 296, 1815 (2002) 5. BREAKTHROUGH OF THE YEAR # Bioterrorism: The Calm After the Storm 1. Martin Enserink The 11 September terrorist attacks and the mysterious anthrax letters, mailed a few weeks later, are beginning to put their stamp on the research enterprise, especially in the United States. But although 2002 has been marked by much talk about bioterror, it has also become a year of waiting for action, with major decisions on research funding, regulation, and smallpox vaccination stalled by politics and technical debate. Meanwhile, despite one of the most expansive investigations in FBI history and a$2 million reward, the anthrax killer is still on the loose.

See Web links on bioterror

Infectious-disease researchers are confident that the attacks will eventually produce a funding windfall. In his 2003 budget, President George W. Bush requested a $1.5 billion increase for the National Institute of Allergy and Infectious Diseases, which in turn has asked researchers for proposals on everything from new drugs and vaccines to new research centers and specialized labs. Congress was supposed to approve the spending by 1 October, but election-year politics has stalled any decision until at least January. Still, the vulnerability of the United States to the ultimate bioterror nightmare—a smallpox attack—has diminished considerably. Old smallpox vaccine supplies were dusted off and proven to still work. Together with new vaccine produced by Acambis, a government contractor, there's now enough to cover the entire U.S. population. But government officials were locked in debate for months about how many people should get preemptive shots, primarily because the vaccine is known to cause severe infections and death in a small number of recipients. Last week, the Administration finally announced plans to start vaccinating half a million health care workers and first responders, and another half million in the military. But eventually, the vaccine will be made available to anyone who wants it—a decision that's drawing outspoken criticism from public health experts (see News Focus story). Another topic of intense but unfinished debate is how best to balance security needs against scientific freedom. Researchers are anxious, for example, about upcoming regulations on work with potential bioweapons. Poorly written rules could lead to misguided law enforcement, they say, pointing to the case of Tomas Foral, a 26-year-old graduate student at the University of Connecticut, Storrs, who became the first researcher to be criminally charged with mishandling dangerous agents after he allegedly stored anthrax samples in a lab freezer. Foral avoided indictment by agreeing to perform community service, but research leaders worry that the incident heralds a new, chilly era in their labs. And the National Academy of Sciences' decision to censor a “sensitive” chapter from a recent report about agricultural bioterrorism has helped spark debate about what kinds of unclassified information scientists should withhold from the public in the name of security. The trail of the real anthrax killer, meanwhile, appears to have grown cold, despite extensive help from anthrax scientists. In August, a break in the case appeared imminent after FBI sleuths twice searched the home of Steven Hatfill, a former Army microbiologist with an interest in bioterrorism. Hatfill lost his job as a bioterrorism preparedness instructor at Louisiana State University, Baton Rouge, after Attorney General John Ashcroft called him a “person of interest,” but he was never charged with any offense. ## Online Extras on Bioterror ### Papers and Articles D. Malakoff, “Congress Adopts Tough Rules for Labs,”, Science 296, 1585 (2002) M. Enserink, “How Devastating Would a Smallpox Attack Really Be?,” Science 296, 1592 (2002) D. Malakoff, “One Year After: Tighter Security Reshapes Research,” Science 297, 1630 (2002) M. Enserink, “One Year After: Hunt for NIH Funds Fosters Collaboration,” Science 297, 1630 (2002) J. Couzin, “A Call for Restraint on Biological Data,” Science 297, 749 (2002) D. Malakoff, “Student Charged With Possessing Anthrax,” Science 297, 751 (2002) J. Mervis and E. Stokstad, “NAS Censors Report on Agriculture Threats,” Science 297, 1973 (2002) M. E. Halloran et al., “Containing Bioterrorist Smallpox,”, Science 298, 1428 (2002) J. Koopman, “Controlling Smallpox,” Science 298, 1342 (2002) Perspective article on the Halloran et al. Science paper. R. M. Atlas, “National Security and the Biological Research Community,” Science 298, 753 (2002) Policy Forum calls for an “Asilomar-type” conference to reckon with balancing scientific openness with national security. R. Bayer and J. Colgrove, “Public Health vs. Civil Liberties,” Science 297, 1811 (2002) Policy Forum, with links to additional Web resources. ### Interesting Web Sites Biodefense and Bioterrorism Internet links from MEDLINEplus Biological Warfare from the NLM's Specialized Information Services Bioterrorism-related articles in the CDC's Emerging Infectious Diseases NIAID's Biodefense Web site FDA's Bioterrorism Web site Federation of American Scientists' Chemical & Biological Arms Control Program Johns Hopkins' Center for Civilian Biodefense Strategies St. Louis University's Center for the Study of Bioterrorism Epidemiologic Information on Bioterrorism from the UCLA School of Public Health 6. BREAKTHROUGH OF THE YEAR # Breakdown of the Year: Physics Fraud 1. Robert F. Service The past year witnessed more than just high points. The physics community suffered two stunning setbacks when separate investigations concluded that a physicist at Bell Laboratories in Murray Hill, New Jersey, and another at Lawrence Berkeley National Laboratory (LBNL) in California committed fraud. At Bell Labs, device physicist Jan Hendrik Schön was fired on 24 September, shortly after officials there received word from an independent committee that Schön fabricated data and falsified reports from 1998 through 2001 (Science, 4 October, p. 30). Bell Labs officials organized the committee in May to look into allegations that portions of figures in separate experiments appeared to have been duplicated (Science, 24 May, p. 1376). Ultimately, the committee members concluded that Schön either falsified or fabricated data in 16 of the 24 cases they reviewed, and they raised questions about the other eight cases. The committee found no evidence of misconduct by any of Schön's 20 co-authors on the suspect papers. Schön, who has denied the charges, and co-authors have moved to retract the 16 suspect papers, and Bell's parent company, Lucent Technologies, has pulled six patent applications based on Schön's work. See Web links on physics fraud Schön, unfortunately, doesn't have a monopoly on this year's alleged misconduct in physics. One of LBNL's claims to fame is a rich history of creating new elements, unstable heavy species that decay in a fraction of a second. But that history has been tarnished. In July, a team of physicists from the lab withdrew a paper from Physical Review Letters that contained evidence for the creation of element 118—evidence that disappeared when investigators looked closely at the original tapes. After the laboratory concluded that the data had been fabricated, Victor Ninov, who was in charge of the analysis, was fired. Ninov has filed a grievance contesting the charges. Ninov's influence might have extended beyond LBNL. According to Sigurd Hofmann of the Institute for Heavy Ion Research (GSI) in Darmstadt, Germany, there was evidence of fabricated data in two other experiments that Ninov worked on when he was part of Hofmann's element-hunting team. Luckily, only element 118 has vanished in a puff of smoke. ## Online Extras on Physics Fraud ## The Schön Affair ### The Story in Science R. F. Service, “Molecules Get Wired,” Science 294, 2442 (2001) Science's Breakthrough of the Year article for 2001, on molecular electronics, included brief mention of the Bell Labs work. R. F. Service, “Pioneering Physics Papers Under Suspicion for Data Manipulation,” Science 296, 1376 (2002) R. F. Service, “Physicists Question Safeguards, Ponder Their Next Moves,” Science 296, 1584 (2002) R. F. Service, “Winning Streak Brought Awe, and Then Doubt,” Science 297, 34 (2002) R. F. Service, “Bell Labs Fires Star Physicist Found Guilty of Forging Data,” Science 298, 30 (2002) D. Kennedy, “Next Steps in the Schön Affair,” Science 298, 495 (2002) An editorial, by Science's editor-in-chief, in the aftermath of the Bell Labs report. Retraction, Science298, 961 (2002) Official retraction, by the nine coauthors, of the eight Science papers included in the Bell Labs investigation. ### Other Links Press release and Web page posted by Lucent Technologies to report investigation findings. Includes links to PDF versions of full report and executive summary. D. Goodstein, “In the Matter of J. Hendrik Schön,” Physics World (November 2002) Viewpoint article on physics misconduct, by Caltech vice provost. Editorial statement, Nature419, 425 (2002) Statement by editors on the five Nature papers included in the Bell Labs investigation [full text; subscription required]. ## Element 118, “Found” and Lost ### The Story in Science R. F. Service, “Berkeley Crew Bags Element 118,” Science 284, 1751 (1999) C. Seife, “Berkeley Crew Unbags Element 118,” Science 293, 777 (2001) C. Seife, “Heavy-Element Fizzle Laid to Falsified Data,” Science 297, 313 (2002) ### Other Links 7 June 1999 news release from Lawrence Berkeley Lab, announcing the initial discovery. 27 July 2001 LBL news release on the retraction of the element 118 results. “Misconduct Case Gets News Coverage,”Currents (LBL newsletter), 26 July 2002 “Editorial Note: Observation of Superheavy Nuclei Produced in the Reaction of 86Kr with 208Pb [Phys. Rev. Lett. 83, 1104 (1999)],”Phys. Rev. Lett.89, 039901(E) (2002) PRL item withdrawing Ninov et al. element 118 results [page contains link to full-text PDF of retraction; subscription required for full text]. 7. BIOLOGICAL AGENTS # New U.S. Rules Set the Stage for Tighter Security, Oversight 1. David Malakoff One of science's hottest fields is now becoming one of its most heavily regulated, too. The U.S. government last week unveiled sweeping new bioterror research regulations that will require 20,000 scientists at nearly 1000 laboratories to beef up security—or face hefty fines and jail sentences. The interim rules, due to go into effect early next year, could also force scientists to get prior approval for a growing list of sensitive experiments. Academic and industry scientists peppered government officials with questions at a public meeting to review the new rules earlier this week in downtown Washington, D.C. Some worried that looming deadlines and steep start-up compliance costs—up to$700,000 per lab—will disrupt important research. Others praised the government for striking the right balance between science and security. And all sides predicted that it will take time to work out the kinks in the new system.

“It's a major change, and there are a lot of questions about how it's going to work in practice,” says Andy Garcia-Rivera, who heads biosafety programs at Cornell University in Ithaca, New York. “We're doing the best we can under very tight deadlines,” said Larry Sparks, a senior adviser at the Centers for Disease Control and Prevention (CDC) in Atlanta, which issued the new rules in conjunction with the U.S. Department of Agriculture (USDA).

The 13 December announcements in the Federal Register, which ran nearly 50 pages, are a response to the 2001 anthrax letter attacks. Alarmed by reports of weak security in labs where researchers study deadly viruses, bacteria, and other potential bioweapons, Congress this summer passed a bioterror bill that called for stricter controls on dozens of “select agents” that could imperil people, farm animals, and crops (Science, 31 May, p. 1585).

The new rules mete out responsibilities among universities, private companies, and government laboratories, all of whom must agree to unannounced inspections. Labs that handle any of nearly 100 select agents, for instance, must register with the government, submit detailed physical security and training plans, and provide the names—and probably fingerprints—of all workers for background checks. Researchers will also need permission to send or receive these agents.

In what may be one of the plan's most controversial provisions, prior approval from the Department of Health and Human Services will be needed for genetic engineering experiments that might make a select agent more toxic or more resistant to known drugs. Government-funded scientists are already subject to that restriction under National Institutes of Health (NIH) guidelines, which require the agency's Recombinant DNA Advisory Committee (RAC) to approve such experiments. And government officials say the list of restricted experiments could grow.

One researcher thinks that expansion is a good idea. Biochemist Richard Ebright, a Howard Hughes Medical Institute investigator at Rutgers University in Piscataway, New Jersey, says the list should include experiments that could lead to less effective vaccines or better methods for making or spreading bioweapons. “It's common sense that such work get stricter scrutiny,” he says.

But Ron Atlas, a bioterrorism expert at the University of Louisville in Kentucky and president of the American Society for Microbiology, is skeptical. “I'm not sure the government should start proscribing experiments … and locking rules into regulations” which can be difficult to adjust, he says. Instead, he recommends that the government instruct researchers to follow the NIH guidelines, which he says are more flexible and can change with the times.

Another issue, Atlas and other researchers say, is exactly who would review sensitive experiments. RAC, they note, has narrowed its focus in recent years, as genetic engineering has become commonplace, and it conducts most of its business in public. But secrecy might be a better fit with bioweapons research. “There is going to be an issue around transparency,” says one scientist, who asked to remain anonymous.

Other debates are likely to erupt before the two agencies finalize their rules this winter. One involves the best way to regulate protein and gene fragments that might be useful to weapons makers. Another centers on how the Department of Justice will ensure timely and accurate background checks and evaluate security plans. Researchers also want CDC and USDA to make sure that their rules agree. Ebright, for instance, wonders what will happen if the two agencies disagree on whether a scientist should be allowed to use a nonlethal variety of a select agent. “There should be a consensus,” he says.

Researchers have until 11 February to send in comments. And scientists hope that the final answers arrive by next fall, when all labs currently handling select agents must be in compliance.

8. CALIFORNIA BUDGET

# Latest Cuts Send Universities Reeling

1. Andrew Lawler

A rapidly ballooning budget deficit is forcing California to make substantial cuts in state-funded science programs. The reductions affect facilities used by a global community of researchers, from astronomical observatories to oceanographic collections. And the bad news is expected to get worse: Next month, Governor Gray Davis will propose chopping at least $29 million, roughly 10%, from the state's next research budget in response to a$21 billion deficit.

California's woes are a result of a nationwide economic downturn, declining tax revenues, and an increased need for social services across the country. State officials who only a few years ago were debating how to spend surpluses are now putting the squeeze on universities and research institutes. “There's not much joy in finding misery elsewhere, but this problem is no longer unique to California,” says Joseph Miller, director of the Lick Observatory at the University of California (UC), Santa Cruz. “It's a precarious situation,” adds Nils Hasselmo, president of the Washington, D.C.-based Association of American Universities.

Earlier this year, the California legislature imposed a $32 million cut on state-supported science at the nine-school UC system as part of a 10% spending rollback for the fiscal year that began 1 July. Last week, the governor proposed taking another$18 million bite out of research as part of an extraordinary set of midyear cuts to higher education. And in January, Davis will propose another round of cuts of at least 10%, according to UC spokesperson Brad Hayward. “These [new] budget cuts will be painful because we have already absorbed major cuts,” says UC president Richard Atkinson. The details of the 2002-03 cuts will be fleshed out this week at a meeting of the university system's Board of Regents, which is also reviewing a hike in student fees.

For the Scripps Institution of Oceanography at UC San Diego, the belt tightening means reducing by half the $300,000 budget for its oceanographic collections. Among the most extensive in the world, they include marine vertebrates and invertebrates as well as sediment cores and dredged rocks. If the state subsidy is eliminated in 2004–05, as expected, the collections would need to find alternative funding to remain open, says Scripps Director Charles Kennel. He says commitments to staff salaries and collaborative ventures come first, leaving him with little choice. Kennel hopes that by sounding the alarm, curators will be able to generate enough nonstate funding to keep the collections intact and operating. “Our folks aren't going to go quietly into the night; they will fight like hell,” says Kennel. But Scripps's Mark Ohman, curator of the pelagic invertebrate collection, says it will be difficult to find alternate sources. “Scientists who run the collections think that it is unrealistic to expect to raise a permanent endowment of$5 million to $10 million in 9 months,” he says. The next round of cuts may also leave Scripps with nothing for its portion of the California Cooperative Oceanic Fisheries Investigation, a 50-year effort with the National Oceanic and Atmospheric Administration (NOAA) and California's Fish and Game Department to monitor the ocean ecosystem. Kennel says that NOAA is setting up a committee to examine the problem. At Lick Observatory, Miller is coping with a$500,000 cut to his $5 million budget by laying off a handful of technicians. “This is hitting us at our heart,” he says. The cuts are also expected to delay several projects, including work on advanced coatings for telescopes and planning for the$600 million, 30-meter California Extremely Large Telescope project. “Research at UC has been a driver for the economy,” Miller adds. “Such cuts are so shortsighted.”

Agricultural research is taking it on the chin, too. The renowned agricultural and environmental sciences department at UC Davis will be losing 25 research positions through attrition, says its dean, Neal Van Alfen, who notes that the department has not yet recovered from the last recession in the early 1990s. But until the U.S. economy bounces back, he and other research managers can expect more hard times.

9. SCIENCE AND SECURITY

# Academy Asks to Ease Visas for Scholars

1. David Malakoff

University of Utah physicist Xiaomei Jiang rushed home to China this fall after her parents died in a car crash. But new security reviews adopted after the 11 September terrorist attacks have so far blocked the fifth-year doctoral student from rejoining her lab in Salt Lake City. Jiang, unfortunately, has plenty of company as she waits for a new visa. Security reviews are causing delays that threaten the health of U.S. science, say the leaders of the National Academies, which last week called on the government to fast-track foreign researchers seeking to enter the country.

The academies issued a 3-page warning after hearing “numerous” reports of immigration problems from academic researchers and seeing the impact on several of its own meetings, says Bruce Alberts, president of the National Academy of Sciences. The restrictions, enacted “in the name of national security, are having serious, unintended consequences for American science, engineering, and medicine,” says the 13 December statement, which was also signed by William Wulf, president of the National Academy of Engineering, and Harvey Fineberg, president of the Institute of Medicine. To prevent future disruptions, they ask the Department of State to reinstate a “precleared” status for foreign scientists who travel frequently to the United States, create a special visa for researchers with solid credentials and invitations from U.S. scientists, and consult with U.S. scientists on which fields should raise red flags.

Consular officials have a strong incentive to err on the side of extreme caution, the statement notes, as they face criminal penalties for granting visas to terrorists. The academy presidents urge the State Department to create some type of counterweight that would also encourage the officials to smooth the way for “scholars who benefit our nation.” A State Department official familiar with the problem says the suggestions are “helpful and on target; we're already working to make them happen.” But he and White House officials warn that progress could be slow. Congressional action might be needed to address the consular liability issue, which is enshrined in law, or to carve out special visas for visiting scientists.

Recent enrollment statistics suggest that the delays are so far having a limited impact on U.S. academic life. The number of foreign students at 20 major research universities rose by 4% this fall, to 36,656, according to a survey released last month by the Association of American Universities. But there has been a 10% drop in the number of foreign faculty members and researchers on campus, the survey found, and more students and scholars reported visa delays or denials than in the previous year. Those numbers could climb in the short run, as the government struggles to beef up security reviews and begin monitoring foreign students studying in “sensitive” fields. On 12 December, the State Department unveiled one monitoring program, and more plans are expected shortly.

In the meantime, Jiang's colleagues say they miss her and her talent in running key experiments. They also worry that her visa troubles could sour the promising physicist, who co-authored a paper in Science on plastic lasers (4 February 2000, p. 839), on her long-term career prospects in a country that once welcomed her for training.

10. STEM CELL MEDICINE

# Stanford Gets Gift for New Institute

1. Constance Holden

Stanford University last week announced the formation of a new, privately funded institute to marry research on stem cells and cancer in a search for new therapies. The announcement precipitated a brief media flurry over the issue of cloning, leaving university officials scrambling to beat down press accounts that suggested the school might become a baby factory.

A $12 million gift from an anonymous donor has kicked off the Institute for Cancer/Stem Cell Biology and Medicine, to be headed by hematopoietic stem cell researcher Irving Weissman. The university will build on existing faculty research but also hopes to recruit more scientists. Stanford medicine Nobelist Paul Berg says the goal is to raise$100 million to support research on genetically based treatments for cancer, Parkinson's disease, heart disease, and other illnesses.

Stanford got in hot water after the institute stated an intention to develop new human embryonic stem (ES) cell lines to study particular diseases. During an interview with the Associated Press (AP), Weissman acknowledged that scientists might someday try to create human stem cell lines for this type of research through nuclear transfer—otherwise known as therapeutic cloning. The resulting AP story, declaring that “Stanford University has announced its intention to clone human embryos,” forced Stanford officials to hold a press conference immediately to deflate the brouhaha. They followed it with a statement emphasizing that “creating human stem cell lines is not equivalent to reproductive cloning.” Berg, who appeared at the press conference, calls the episode “bizarre. … You have an audience [that] seems not to have been on this planet for the last 2 years.”

Stanford's plans are consistent with a state law passed in September that endorses both stem cell and therapeutic cloning research (Science, 27 September, p. 2185), although Berg says that the new center “was in the works long before that.” The University of California, San Francisco (UCSF), sponsored such work before researcher Roger Pedersen moved from there to the U.K.'s Cambridge University last year, and UCSF may revive it as part of another privately funded stem cell initiative launched earlier this year (Science, 16 August, p. 1107). Another boost may come from Massachusetts: Legislators there introduced a bill this month that would create a state fund for ES cell research.

Researchers applaud the Stanford initiative, which they see as necessary in light of the current prohibition on federal funding for research with human ES cells derived after 9 August 2001. MIT stem cell researcher George Daley says he hopes it is “just one of the first of what should be many privately funded institutes to take up the slack.” The biggest one to date is the Institute for Cell Engineering at Johns Hopkins University, formed last year with an anonymous donation of $58.5 million. UCSF is currently focused on distributing its two lines of presidentially approved stem cells to the 40 groups that have requested them. 11. RESEARCH FUNDING # Italian Researchers Facing Lean Times 1. Alexander Hellemans* 1. Alexander Hellemans is a writer in Naples, Italy. NAPLES—Italian researchers are distraught this week after legislators approved a 2003 budget that could shutter some national facilities and threaten Italy's contributions to major international research centers. “It is the worst situation in research in Italy since [World War II],” says legal historian Luigi Capogrossi Colognesi, a member of the governing council of the National Research Council (CNR), the country's largest research organization. The parliamentary vote came after days of heated debate in the Senate, punctuated by outraged researchers demonstrating noisily outside and a mass resignation by university rectors, who say that a written commitment to adequate funding is the only thing that will bring them back. The chamber of deputies was preparing this week to rubber-stamp the budget decision by the upper house. The Senate budget contained$1.6 billion for public research organizations, a cut of 1.6%, and level funding of $6.3 billion for universities. Fixed costs such as salaries and operating expenses consume most of the budget, leaving ongoing research projects most vulnerable to cuts, says Rino Falcone, an artificial-intelligence researcher at CNR. It is not just the level of funding that has angered researchers. They are also incensed that the Ministry of Finance has proposed raising additional funds for universities by increasing the state tax on cigarettes. “I don't agree with people smoking to finance my research,” says oncologist Alfredo Budillon of the University of Naples. Scientists are also galled by a government proposal to create a$98 million special science fund distributed at the personal discretion of Prime Minister Silvio Berlusconi. “This is a new attack on the autonomy of the scientific community and its institutions,” says Falcone. Parliament was expected to vote this week on both proposals.

CNR, with about 100 research centers throughout Italy, will receive $477 million, about 2.5% less than in 2002, says Capogrossi Colognesi. But the actual impact on science will be greater because its shrunken budget must cover raises written into existing labor agreements. CNR recently announced that it might have to rescind its membership in the European Science Foundation, and other international collaborations might also be sacrificed. The government gave no explanation for why some organizations were hit much harder than others. “It is very hard to understand what the original motivations for these moves are, apart from trying to cut budgets across the board,” says physicist Alfonso Franciosi, chair of the National Committee for Synchrotron Radiation Research at the National Institute for the Physics of Matter (INFM). Facing a 30% cut, INFM will be forced to slash its contribution to the Elettra x-ray synchrotron in Trieste, which each year hosts 800 researchers from across Europe. Elettra could be shut down temporarily as early as next month. INFM officials are also wondering how to meet their$11-million-a-year obligation to other European x-ray and neutron projects, among them the European Synchrotron Radiation Facility, the Institute Laue-Langevin neutron source, both in Grenoble, and the future European Spallation Source.

The National Institute for Nuclear Physics says it will be able to maintain its subscription to major facilities such as the CERN particle physics lab near Geneva, despite a 10% cut. But it will have to reduce its contribution to the building of several detectors for the Large Hadron Collider there, reports physicist Carlo Bernardini of the University of Rome.

Researchers say that the new cuts only underscore the country's status as the stepchild of European research. Italy spends less than 1% of its gross national product on research, about half the European average. Nobelist Carlo Rubbia, whose National Agency for New Technologies, Energy, and the Environment faces a 15% cut, says that Italy is “marginal both in Europe and in the world in the field of science.”

12. 2004 BUDGET

# No Holiday Cheer for NIH, NSF

1. Jeffrey Mervis,
2. Jocelyn Kaiser

When is a budget increase not really a budget increase? When President George W. Bush prepares a 2004 request to Congress before legislators have completed work on this year's budget.

Although the president's request for the next fiscal year won't become public until early February, Science has learned that the White House has settled on a 9% increase for the National Science Foundation (NSF), to roughly $5.4 billion. That sounds like a hefty increase for a domestic research agency when the economy is in a slump, a war against Iraq looms, and the budget deficit is growing. But it might be no more than Congress gives NSF this year. The$23.3 billion National Institutes of Health (NIH) has received similarly Scrooge-like news for the holidays: The White House has offered less than a 1% hike, and Department of Health and Human Services (HHS) officials are appealing.

The NSF request is less generous than it seems for two reasons. First, it's based on the president's 2003 request for a 5% hike, a number that Congress is almost certain to surpass when it finishes work next month on the budget for the fiscal year that began 1 October. (The House has already approved a 13% increase, and a Senate panel has endorsed a 12% hike.) Second, before calculating the 9% increase, budget officials subtracted $76 million from a dead-on-arrival proposed transfer of funds to NSF from three other agencies. The result is a presidential request of roughly$600 million over 2002 levels, which exactly splits the difference between the $633 million hike for 2003 approved by the House and the$564 million added by the Senate panel.

There's no sugar coating on the NIH request, which sources say is a mere $50 million over the expected 2003 total of$27.3 billion. HHS Secretary Tommy Thompson is pushing for his original 5% request. But NIH watchers are dubious of anything more than the 2% that the White House has projected for future years. And unlike previous years, nobody is counting on Congress to come to NIH's rescue.

13. HIGH-ENERGY PHYSICS

# CERN Council Chooses ITER's Head as Chief

1. Charles Seife*
1. With reporting by Pallava Bagla.

Europe's premier accelerator laboratory has elected a director general without training in particle physics but skilled in managing large projects. That's no accident: CERN's governing council made it clear last week that building the Large Hadron Collider (LHC) on time and within budget is the lab's top priority, with everything else—including a streamlined research portfolio—taking a back seat.

The new leader is Robert Aymar, a 66-year-old French plasma physicist who currently directs the International Thermonuclear Experimental Reactor (ITER). A multibillion-dollar international tokamak project, ITER has survived a downsizing of its original design and the withdrawal of the United States before regaining its momentum; the partners are now in the final stages of selecting a site (Science, 20 September, p. 1977). That performance under fire was not lost on the CERN council. “For the time being, CERN's activities are centered about building the LHC and not exploiting the science, and [Aymar] has long-standing experience,” says Jean-Pierre Ruder, the Swiss delegate to the CERN council. The outgoing director-general of CERN, Luciano Maiani, agrees: “I find that Aymar is very well qualified, even though he's not a particle physicist.”

Aymar, who will begin his 5-year term in January 2004, is best known for directing the Tore Supra project, which used a large magnetic bottle called a tokamak to study very hot plasmas. But he also has had a lot of exposure to particle physicists. During the 1990s, he oversaw particle-physics experiments as head of the Sciences of Matter directorate of France's atomic energy lab, CEA. He also helped design the LHC and chaired the LHC external review committee when the project ran into budgetary problems (Science, 5 October 2001, p. 29). “I was involved in the decisions about the LHC at all levels,” says Aymar. “The big challenge [now] is to make sure that the LHC is achieved correctly. The timing should be controlled by technology concerns, not financial ones.”

Financial concerns have dominated CERN for more than a year. A 30% cost overrun in the LHC's $1.6 billion budget has forced the council to shut down several key experiments in 2005 and pare research and development projects to a bare minimum. Although the council's acceptance of the cuts last week was no surprise—the details had been announced earlier this year (Science, 29 March, p. 2341)—physicists remain concerned about the impact of the cuts. “It leaves very little scope for preparing for the long-term future,” says Phil Allport, a physicist at Liverpool University in the U.K. and adviser to CERN. “There's pain in the lost physics programs, but more pain in the strategic R&D that underpins CERN's future post-LHC.” According to Aymar, the damage can be mitigated by a closer collaboration between CERN and other European laboratories, but “CERN has to devote all its financial capacity to make LHC possible.” Current estimates put the cost to complete the collider in 2007 at approximately$210 million, with a $96 million contingency budget. In other actions, the council added a third category to CERN's two formal levels of participation. Traditionally, member states make a yearly budgetary contribution and in return receive full access to the facilities and a vote in the CERN management. Non-European states could contribute to experiments on an approval-only basis or, like the United States, could become observers, with limited access to the laboratory and no voting rights. India, which has been participating in CERN experiments, including the LHC, is now the newest CERN observer state and may be in line for a new, more active status as an associate member. “India was very happy at being given the observer status, for it was a well-earned recognition of the scientific and technical contribution Indians have been making at CERN,” says Ravi Bhushan Grover, the director of India's Strategic Planning Group of the Department of Atomic Energy in Mumbai. Associates can have full-time employees at CERN and can bid on contracts, although they would still lack a vote. In return, the associate state would pledge a yearly contribution to CERN's budget, pegged to the associate's gross domestic product. “India welcomes the new tier that has been opened up,” says Grover. However, he adds, “whether we will be able to afford this … will have to be assessed only when the details are made available.” 14. ASTRONOMY # Europe's Telescope Builders Aim High 1. Govert Schilling* 1. Govert Schilling is an astronomy writer in Utrecht, the Netherlands. LONDON—To reach for the heavens, you need to think big. European astronomers are taking that motto to heart. Last week, teams from across the continent met here to start work on a joint design for what they hope, 12 years hence, will be the largest telescope in history. The European Large Telescope (ELT) would take an order-of-magnitude leap from the scale of today's telescopes, which have mirrors about 10 meters across, to one up to 100 meters in diameter, capable of revealing the workings of the universe and examining nearby stars and planets in unimaginable detail. “This is astronomy beyond the wildest dreams of anybody in the world,” says Tim Hawarden of the U.K.'s Astronomy Technology Centre in Edinburgh. It's no dream, ELT boosters say. Over the past 15 years, astronomers building today's top-rank telescopes have refined key new technologies needed to make larger scopes—techniques such as building big mirrors from smaller hexagonal segments and tweaking a telescope's optical system in real time to compensate for atmospheric turbulence. Astronomers in the United States have set their sights on a 30-meter Giant Segmented Mirror Telescope, which a 2001 decadal review by the National Research Council identified as the top priority for U.S. ground-based astronomy. But researchers from the European Southern Observatory (ESO) and a number of national institutes across Europe have decided that a much bigger telescope is technologically feasible. “No one has as yet managed to find anything [in these plans] that we can't do,” says Gerry Gilmore of the University of Cambridge. Last Friday, at a meeting hosted by the Royal Astronomical Society in London and attended by most of the European and American teams working on extremely large telescopes, Gilmore announced that the European groups will work together on a single project. And although the telescope's size, optical design, and location are still to be decided, the ELT steering committee met for the first time this week in Garching, Germany, to plan future work. “This is a big step ahead for Europe,” says Torben Andersen of Lund Observatory in Sweden. The ELT will build mainly on the results of two large design studies carried out over the past few years. Euro50 is a proposed 50-meter telescope under study by scientists in Finland, Ireland, Spain, Sweden, and the United Kingdom. Like all of the planned giant telescopes, Euro50 would have a segmented mirror consisting of hundreds of hexagonal tiles combined to make a reflecting surface longer and wider than a Boeing 747. Euro50 would cost about$600 million, says Andersen, its technical project leader. Much more ambitious is the Overwhelmingly Large Telescope (OWL), designed by ESO. With a 100-meter mirror, OWL would stand almost as tall as the Great Pyramid but still cost a “mere” $900 million, says project engineer Philippe Dierickx, thanks to standardization and mass production in its design. Even a 50-meter ELT would have more light-gathering power than all telescopes in the history of astronomy put together and would revolutionize the study of just about every possible class of objects, scientists at the London meeting said. Members of the design team acknowledge that building ELT won't be easy. “These are very exciting prospects, but many details need to be worked out,” says Isobel Hook of the University of Oxford. In particular, astronomers still have to design workable adaptive optics for very large telescope apertures. But Andersen is confident: “Many bright people are working on adaptive optics, and they have already made wonderful progress.” Funding will pose another huge challenge. Gilmore estimates that operating costs alone will add up to$1 billion over 10 years. To garner such sums, the European partnership may have to go global. Andersen and Dierickx say they would welcome a collaboration with colleagues in the United States. But American astronomers might need to be convinced that jumping straight to a 50- or 100-meter mirror will work. “Thirty meters is the next logical step,” says Richard Ellis of the California Institute of Technology in Pasadena, one of the leaders of the 30-meter California Extremely Large Telescope project, which got a boost this fall when Caltech made it the centerpiece of an ambitious fundraising campaign (Science, 8 November, p. 1151). Building a much larger telescope is just too risky, Ellis told those attending the London meeting.

But ESO's Dierickx says that the American approach, which scales up the design of the 10-meter Keck telescopes, is too conservative. ESO plans to come up with an entirely new design that makes maximum use of standardization and mass production, he says. According to Gilmore, the ELT design study will likely take 4 years to complete. After that, another 8 years will be needed to build the monster telescope, probably at La Palma in the Canary Islands or in Chile's Atacama Desert. “If Europe could build it without the U.S., we would,” says Gilmore. “Competition is a good thing. It makes you try harder.”

15. PUBLIC HEALTH

# Rough-and-Tumble Behind Bush's Smallpox Policy

1. Jon Cohen,
2. Martin Enserink

After months of ideological tugs-of-war over whether to vaccinate the public to protect against a bioterrorist attack, the Administration settled on a compromise that most scientists can live with

On the afternoon of Monday, 9 December, top public health officials met at Vice President Richard Cheney's residence for a crucial meeting to help hammer out the details of the Bush Administration's policy for smallpox vaccination. For some 7 months, a debate had raged within the Administration over how widely to distribute the vaccine. Now the scientists came armed with data they thought would, at long last, bring closure.

Without question, the decision to vaccinate against an eradicated virus required careful deliberations. The central issue: how to balance the known risks of the vaccine, which can injure and kill, against the unknown risk that rogue states or terrorists would use the smallpox virus as a weapon. Almost uniformly, the public health experts were opposed to immunizing the general public to protect against an uncertain attack. But there was strong support among some in the White House—and especially in the vice president's office—for widespread vaccination “pre-attack,” in part because they questioned whether the Department of Health and Human Services (HHS) could halt an outbreak should a bioterrorist attack occur against a largely unvaccinated population.

Around the dining room table, HHS officials—including the heads of the Centers for Disease Control and Prevention (CDC) and the Office of Public Health Preparedness (OPHP)—unveiled a detailed plan to Cheney, his staff, and Homeland Security chief Tom Ridge on how an outbreak could be contained. If the unthinkable should happen, within 12 days highly coordinated public health teams would distribute millions of doses of vaccine, and disease detectives would fan out to find and isolate patients and vaccinate their contacts, derailing the virus before it could spread far beyond the initial victims.

By the end of the meeting, according to one participant, the scientists had convinced Administration officials that HHS's containment plan would work, leading them, at last, to a consensus that had eluded them to date. “Had we not reassured [Vice President] Cheney on that day that we really did know what we were talking about, I think the vaccine would have been offered more widely to the general public,” says one scientist who attended the meeting.

Four days later, on 13 December, President George W. Bush, with HHS officials at his side, announced the policy: immediate, mandatory vaccination of 500,000 military personnel and a voluntary campaign, to be completed by summer 2003, among a similar number of health care workers or “first responders”—those at highest risk of coming in contact with the virus. After that, the government would offer the vaccine to up to 10 million additional health care workers, police, firefighters, and other personnel deemed essential. Bush explicitly did not “recommend” that anyone beyond these groups receive the vaccine—indeed, he discouraged it by saying neither his family nor his staff would get it. If people “insisted” on being vaccinated, he said, the government would “work to accommodate” them.

Reaction in the public health community has largely been relief that mass vaccination was staved off. Ronald Atlas, for example, who heads the American Society for Microbiology and co-runs the Center for the Deterrence of Biowarfare and Bioterrorism at Kentucky's University of Louisville, calls the president's decision “prudent.” Others, such as Tara O'Toole of the Center for Civilian Biodefense Strategies at Johns Hopkins University in Baltimore, Maryland, remain concerned that millions might still receive a live-virus vaccine that could pose a potential threat not only to themselves but also to others who come in contact with them. Extending vaccination to police and firefighters is unnecessarily risky, says O'Toole, because they're unlikely to encounter the virus.

To understand how the Bush Administration crafted its smallpox vaccination policy, Science spoke with several players in the tortuous negotiations. Experts entered with strongly held scientific concerns, but they quickly found that the issues went far beyond science. Public health officials found themselves wrestling with the traditional openness of scientific exchange versus the necessary secretiveness of intelligence data and a famously tight-lipped White House that, as one scientist close to the process said, “has a great concern for managing news.” As making the difficult decision dragged on, one insider dubbed it “a soap opera.”

Given the national security issues at stake and their close working relationships with the president and the vice president, many would speak candidly only if they remained unidentified. Most give the Administration credit for its willingness to dive into complex scientific issues, and several advisers came away impressed with Cheney's grasp of the current data. “There was a complete lack of trying to bully public health officials,” insists Anthony Fauci, head of the National Institute of Allergy and Infectious Diseases (NIAID). And in the end, the White House arrived at a compromise that most of the scientists felt was, in Fauci's words, “reasonable.”

## Supply and demand

Fears of a smallpox attack erupted after 11 September and the subsequent anthrax attacks, but a public vaccination campaign was then not even on the agenda. The government had only about 15 million doses of an old vaccine called Dryvax on hand, so discussions focused on how to increase the supply—fast—to be prepared should an outbreak occur.

The tenor of the discussions changed last winter. Studies showed that some 85 million doses of another old vaccine donated to the government by Aventis Pasteur, the Franco-German pharmaceutical giant, remained potent. And government-sponsored scientists reported that even when diluted five or 10 times, Dryvax remained effective. From then on, there was enough vaccine to cover the entire country before a single case of smallpox even occurred, says HHS's Philip Russell, a special adviser on vaccines at OPHP.

The first indication of an ideological rift between the White House and HHS surfaced soon thereafter. In late winter, General Bruce Lawlor of the Office of Homeland Security, at policy-coordinating committee meetings that included top brass from different agencies, began to raise questions about how quickly HHS could vaccinate the public, sources say. Lawlor suggested that it made sense to immunize the public widely before an attack occurred. That would make an attack easier to contain, resulting in less panic and far less economic damage; vaccination might even preempt an attack altogether because a largely immune population would make an unattractive terrorist target. Donald A. Henderson, then head of OPHP (and now a key adviser to that office), was said to be “quite offended” by Lawlor's arguments.

When The New England Journal of Medicine published the dilution studies on 25 April, the issue went public. In an accompanying editorial, William Bicknell, a former commissioner of the Massachusetts Department of Public Health, made the case for reducing the threat by allowing the public to receive the vaccine; in another article, Fauci called for “an open and public dialogue on the advantages and disadvantages of universal voluntary vaccination.”

By then, a vociferous debate was already raging behind the scenes. Henderson, Russell, Fauci, and others urged the White House to move slowly in vaccinating the public, stressing that, short of convincing evidence that a smallpox attack was imminent, the benefits simply did not outweigh the risks.

The existing smallpox vaccine—a living, replicating virus called vaccinia—causes side effects ranging from minor to life threatening (see sidebar, p. 2313). Even with rigorous screening to rule out high-risk people, for each 1 million people vaccinated, between 49 and 935 are expected to suffer severe adverse reactions, and one or two will die. In addition to individual reactions, the vaccine virus can spread inadvertently to unimmunized persons, such as household members or colleagues, simply by rubbing the vaccination site. Because these people did not choose to be immunized and may be at higher risk of side effects, “it's not just about individual rights,” as some libertarians argue, says O'Toole. “It's a societal issue.”

Some also worried that the severe side effects could undermine public confidence in both the government's biodefense effort and vaccination in general. “Once the first two kids with progressive vaccinia are on TV, the public could decide that the government has no idea what it's talking about,” says O'Toole.

## Behind closed doors

In June, CDC's Advisory Committee on Immunization Practices (ACIP) held a special meeting about the issue. After 2 days of intense debate, the panel recommended a cautious approach: vaccination of “smallpox response teams” that would investigate suspect cases, as well as a limited group of workers in a small number of hospitals specially designated to treat smallpox patients. That would result in, at most, 20,000 vaccinations, ACIP chair John Modlin told reporters after the meeting. The committee recommended against vaccinating the general population.

Within the Administration, this “minimalist” view initially seemed to prevail. On 18 July, Cheney and his staff invited the main players, including OPHP's new director, Jerome Hauer, Henderson, and their vaccination coordinator Russell, to a “surprise” visit at the CDC in Atlanta “to thank the frontline troops” for protecting the nation from bioterrorism. Aboard Air Force II, vaccination policy dominated the discussions. “I think we all walked away and said, ‘very good meeting,’” recalls Henderson, who headed the World Health Organization's program that by 1977 had eradicated smallpox. “We finally got all the facts out. There was general unanimity that we have to move deliberately.” To Henderson, that meant vaccinating the frontline responders at highest risk and leaving the public out of the equation. “From the beginning, I've had the feeling we were best off if we took one step at a time,” Henderson says.

But over the summer, the mass-vaccination forces gained momentum. In hearings, op-ed pieces, and back-channel phone calls, Republican Senators Bill Frist, Arlen Specter, and Judd Gregg made the case that anyone who wanted the vaccine should have it. Cheney shared this opinion. According to several sources close to the process, Cheney's chief of staff, Lewis Libby, argued forcefully for widespread immunization. “There's little doubt [that] Libby is the driver,” said one scientific adviser. Cheney staffer Carole Kuntz was also “very animated” about the issue.

Neither Libby nor Kuntz would speak to Science. Several sources ascribe their motives to the libertarian argument, described in an April report by the Cato Institute think tank, that the government has no business telling informed citizens that they cannot have a vaccine bought with tax dollars.

Another source cited the “Dark Winter” scenario, an exercise staged a year before, in which a simulated epidemic spiraled out of control after a smallpox release at shopping malls. Both Libby and Kuntz also have strong ties to the Department of Defense (which Cheney used to head), where pressure was mounting to “take that card from Saddam's deck” before an attack on Iraq. But the vice president also had on his staff voices of moderation, including Noreen Haynes, a physician at Johns Hopkins University. She and her husband Seth Carus, a bioterrorism expert at the National Defense University, were said to play an important role.

The debate ratcheted up in July, when the Monterey Institute of International Studies published a report suggesting that a smallpox outbreak in the former Soviet Union in 1971 had been caused by secret bioweapons tests on Vozrozhdeniye Island in the Aral Sea. The study offered the first, chilling evidence that smallpox could be successfully “aerosolized” and transported by the wind over many kilometers. Some saw this as another argument for widespread vaccination, especially because Russian authorities were unwilling to help study the incident or the virus strain.

Later that month, ACIP met again to advise CDC on a number of specific questions, such as what types of workers to vaccinate at each hospital and how to bandage each person's vaccination site to prevent inadvertent spread. As part of those deliberations, ACIP significantly upped the number of frontline responders to be vaccinated. In a press conference, ACIP's Modlin said half a million might be the right number. (The lone dissenter was Paul Offit of the University of Pennsylvania School of Medicine in Philadelphia, who wanted to stick with the original recommendations.)

Some observers contend that the Administration pressured ACIP into toeing the party line. Others say the panel changed its mind to avoid a huge rift with the government policy that seemed to be shaping up. The new recommendations were “fairly careful waffling,” says Michael Lane, a former director of CDC's smallpox eradication unit, who followed the discussions closely. “They wanted to accommodate the government, create a little wiggle room.”

Not so, Modlin told Science. The numbers changed because since its June meeting, HHS had convinced the committee that the strategy of treating patients in a small number of “designated hospitals” would never work. Hospitals were reluctant to assume this responsibility, for fear that even one suspected case could lead all other patients to avoid them; they also thought that smallpox patients would simply show up at the nearest emergency room. So instead of just a couple of hospitals, HHS had argued, every acute-care hospital in the country had to prepare. “We accepted that,” says Modlin.

Media attention was now riveted on the debate. News stories began appearing nearly every week, all indicating that the policy would extend far beyond the 500,000 that Modlin had mentioned. But despite predictions that a policy statement was imminent, Bush remained mum. Some top officials became frustrated by the delays. “This is complicated, but it's not that complicated,” said one official. The University of Louisville's Atlas decided that the Administration was floating one trial balloon after another to see how they fared. “They're trying to do what they do during political campaigns,” complained Atlas.

## 20/20?

To almost everyone's surprise, Bush finally revealed a few details about the policy in an interview with Barbara Walters on ABC's 20/20 program, snippets of which the network aired on Wednesday, 11 December. The president made clear that the general public could get the vaccine. First Lady Laura Bush added that she would be comfortable having her children vaccinated, a remark that seemingly signaled government support for general vaccination.

But in the official announcement 2 days later, Bush's tone was much different: This time, the message had been very carefully crafted. The government does not recommend widespread vaccination, he said, but for those who “insist” on receiving it, the government would develop ways to give them access. As Administration officials made clear later, this will be anything but easy. To get the vaccine right away, members of the public will have to enter clinical trials of the preparation, assuming they are eligible, and agree to potentially cumbersome follow-up studies. If more than 10,000 people—the number that can be accommodated in the trials—request the vaccine, HHS plans to organize special access to the Aventis Pasteur vaccine, which is not yet licensed, as an Investigational New Drug.

Still, shortly after the White House announcement ceremony, many at HHS rubbed the sweat off their brows. Fauci, who flanked the president during the announcement, said he was glad the Administration had chosen the option it did, but he still felt unsettled. “I'm not happy that we're dealing with a disease that so much effort was put into eradicating and that was such a public health triumph,” said Fauci. “But the fact is, we do have to deal with it.” And, he concluded, “if we do face the horrors of an attack, then we'll be very glad we went through this.” As another high-ranking HHS official said, “Given the world situation, the uncertainty of the threat, the number of voices and noises out there, and the combination of public health and political considerations, this isn't a bad place to be.”

16. PUBLIC HEALTH

# Treating Vaccine Reactions: Two Lifelines, But No Guarantees

1. Martin Enserink

The pictures are almost too gruesome to look at. Dozens of them, displayed in neat rows on an educational Web page* by the U.S. Centers for Disease Control and Prevention (CDC), remind viewers of the horrific side effects of the smallpox vaccine in a small minority of cases. Unfortunately, doctors will have only limited means to help such patients when the United States resumes vaccination. Although two treatments are recommended for vaccine complications, little hard evidence supports the efficacy of either one.

Like smallpox itself, complications from the vaccine—a live, replicating virus called vaccinia—were fading from memory until the terrorism threat brought them back. Most of the researchers who have witnessed these rare cases have retired or died, and most papers describing their work are more than 35 years old. Back then, expertise with the vaccine's side effects was concentrated at the University of Colorado Medical Center in Denver. There, a renowned pediatrician named Henry Kempe led a national reference center where patients—most of them children—were brought to be treated.

Kempe died in 1984, but his younger co-worker Vincent Fulginiti, now retired and living in Arizona, vividly remembers some of the children—especially those with progressive vaccinia, an uncontrollable and usually fatal infection, triggered by a malfunctioning immune system, that spread from the vaccination site. “The virus literally ate up their entire bodies,” Fulginiti recalls.

In the 1950s, Kempe pioneered what is still the first line of defense for vaccine-induced disease: vaccinia immune globulin (VIG), a product made from the blood plasma of recently vaccinated people, which is high in antibodies against vaccinia. The United States has about 600 to 700 doses left from the 1960s, enough to treat the side effects expected when 4 million to 6 million people are vaccinated. But that cache is rapidly being supplemented with new VIG, produced by Cangene, a company inWinnipeg, Canada, and Dynport, a military contractor in Frederick, Maryland.

That's less reassuring than it sounds, because VIG is no wonder drug. Kempe became convinced early on of VIG's efficacy in treating some side effects, says Michael Lane, a former director of CDC's smallpox eradication unit, so he never carried out a controlled clinical trial. “He was a great clinician and a wonderful humanitarian but not a man of science,” says Lane. In fairness to Kempe, Lane adds, there might not have been enough patients available for a rigorous trial. Instead, Kempe compared the fate of VIG-treated patients with historic data.

Those and other studies suggested that VIG worked well and reduced the death rate from eczema vaccinatum, an occasionally fatal complication in eczema patients, by as much as 70%. It also seemed beneficial in severe cases of generalized vaccinia, a pocklike rash that covers the body. But it rarely helped in progressive vaccinia, says Fulginiti, and it was useless against encephalitis. Anecdotal as they may be, such early experiences form the basis for today's treatment guidelines (see table).

View this table:

Today's proposed dosing regimen is based on similarly soft data. In the 1960s, Fulginiti recalls, “we more or less went by the seat of our pants,” starting out with 0.6 milliliters of VIG per kilogram of body weight and—absent improvement—ratcheting up to as much as 10 milliliters per kilogram.

In the past, VIG also has been used to prevent, rather than treat, side effects. In the Netherlands, for instance, adults routinely received a shot of VIG when they were vaccinated, after a 1962 trial suggested that this could cut the encephalitis rate by as much as 77%. (To mass-produce VIG, recalls former director Hans Cohen of the Dutch National Institute for Public Health, vaccinated military recruits were asked to donate plasma in return for “an afternoon off and a pancake.”) But the Dutch had an unusually high encephalitis rate to begin with—perhaps because they used a more virulent vaccine strain—and little is known about VIG's ability to prevent other side effects. The United States has no plans to try a similar strategy today.

If VIG doesn't work, CDC recommends an experimental antiviral drug called cidofovir. Cidofovir is already approved to treat cytomegalovirus infections of the retina in AIDS patients, so many hospitals have the drug on hand and doctors have some experience with it. In lab studies, cidofovir halts vaccinia's replication in the test tube and can save mice from otherwise lethal vaccinia infections.

But whether the mouse data are applicable to humans remains to be seen, because vaccinia gives mice a very severe pneumonia instead of the side effects seen in humans, says John Huggins of the U.S. Army Medical Research Institute of Infectious Diseases in Fort Detrick, Maryland. Good animal models for side effects such as progressive vaccinia and eczema vaccinatum don't exist and would be hard to develop, says Huggins. Cidofovir can also have severe side effects.

The bottom line, says Lane, is that sick vaccinees have two potential lifelines, but neither one offers any guarantees. That makes it all the more important to ensure that those with known risk factors, such as eczema or compromised immune systems, don't get the vaccine. Even with those precautions, as vaccination rates rise, so does the likelihood that Kempe's work will have a 21st century follow-up—and nobody's looking forward to it.

17. PUBLIC HEALTH

# Looking for Vaccines That Pack a Wallop Without the Side Effects

1. Jon Cohen

Like many pox researchers, Bert Jacobs of Arizona State University in Tempe believes it is “premature” to distribute the smallpox vaccine widely. The threat of an outbreak is uncertain, he points out, and dangerous side effects from the existing vaccine, which contains vaccinia virus, are all too real. Why not wait, he wonders, given that “we have the potential for alternatives, at least in advanced research, in the next couple of years.”

Since the 11 September terrorist attacks, the quest for a safer smallpox vaccine has taken on a new urgency, drawing in a number of leading labs. (These efforts are in addition to the vaccinia already in production at Acambis, a U.S.-U.K. biotech company in both Cambridge, Massachusetts, and Cambridge, U.K. Scheduled to reach the market by 2004, the Acambis vaccine is not expected to be much safer than the existing one is.) Some researchers have dusted off experimental vaccines from the 1970s; others have turned to cutting-edge genetic approaches. “There are really exciting new ways to possibly immunize against smallpox,” says Alfred Prince, head of virology at New York City's New York Blood Center, who collaborates with Jacobs. Among the new approaches:

Modified vaccinia Ankara. MVA is a weakened strain of vaccinia developed 40 years ago by Anton Mayr and colleagues at Munich University in Germany. The German researchers reported that by 1978, 120,000 people had received it without any serious side effects. Unlike garden-variety vaccinia, MVA cannot replicate in human cells, but this key safety feature means it may not generate a protective immune response. And MVA never received a real-world test in an actual smallpox outbreak. Researchers have a myriad of tests planned on monkeys and humans over the next several months. In one study, researchers will vaccinate monkeys and then “challenge” them with monkeypox, a simian virus akin to smallpox. A human study will compare vaccinia and MVA head to head, challenging with vaccinia and carefully assessing immune responses.

If the vaccine looks promising, the U.S. National Institute of Allergy and Infectious Diseases may contract for 30 million doses. Even if MVA does not produce an impressive immune response by itself, the earlier German studies suggest it might work as a prevaccine to blunt vaccinia's side effects. Bavarian Nordic, a German-Danish company headquartered in Copenhagen, has begun manufacturing the vaccine; it recently announced that it sold 1 million doses to the German army for that purpose.

LC16m8. This is another weakened vaccinia strain but, unlike MVA, it replicates in human cells. Developed at the Chiba Research Institute in Japan, LC16m8 caused few side effects when tested in 50,000 children in the 1970s. But some leading poxvirus researchers have serious reservations about LC16m8 because it doesn't go through the stage in the viral life cycle called extracellular enveloped virus, which produces the main protective antibody response in vaccinia.

“We're pushing a rapid research agenda into LC16 very hard,” says Donald A. Henderson, a key biodefense adviser to the U.S. Department of Health and Human Services. With his help, VaxGen of Brisbane, California, recently struck a deal with Japan's Kaketsuken to start clinical trials of the vaccine.

Vaccinia-tetracycline combo. Paula Traktman, a poxvirus researcher at the Medical College of Wisconsin in Milwaukee, and her co-workers have genetically engineered vaccinia, stitching in a repressor that shuts down specific viral genes unless the antibiotic tetracycline is present. In theory, a person could take tetracycline and the vaccine simultaneously. If an adverse reaction occurred, stopping the drug immediately would shut down the vaccine virus. Traktman, in collaboration with Prince, is considering testing the idea in humans.

Engineered vaccinia mutants. Like Traktman, Jacobs genetically engineers various versions of vaccinia to study it. By mutating specific genes that control vaccinia's virulence, Jacobs and co-workers have created mutants that replicate well but, when injected into the brains of mice, seem much less likely to cause neurological damage. Clinical trials are a long way off, but a similar vaccine in humans might avoid the brain swelling caused by vaccinia, one of its most devastating side effects.

18. TRANSGENIC CROPS

# China Takes a Bumpy Road From the Lab to the Field

1. Ding Yimin,
2. Jeffrey Mervis*
1. Ding Yimin writes for China Features in Beijing.

China's rejection of applications to sell GM crops and a ban on foreign investment coexist with an expanding research portfolio

BEIJING—The Chinese government has long maintained that transgenic crops are the key to feeding and clothing the country's huge population in the 21st century. And it is backing that conviction with rapid increases in research budgets. But although it once enthusiastically embraced efforts to get genetically modified (GM) seeds into the hands of farmers and welcomed foreign investment in developing GM crops, the government is now proceeding with extreme caution. Officials point to growing concerns about biosafety as the reason for the shift, whereas others see trade policy as the driving force.

The latest example of this dichotomy between supporting research and postponing the harvest of its commercial fruits came last month, when the government's biosafety committee turned down an application for the commercial distribution of a domestically developed, herbicide-resistant variety of rice. It wasn't a big surprise: Although China has approved commercial cultivation of five species of GM crops, most notably Bt cotton in 1997, no staple food crop has ever passed muster. Earlier this year, the government drew the line around GM crops a little tighter by announcing a ban on new foreign investment in agbiotechnology ventures. The rules, issued in March, are part of a series of edicts governing the importation and labeling of GM crops.

Although the two actions are independent, together they emphasize the prevailing attitude toward GM crops. Chinese officials say their cautious approach is needed to make sure all GM crops are safe. The restriction on foreign investment, they argue, simply adds another protective layer by ensuring that overseas companies don't unleash varieties that could threaten local strains. “Our government is not trying to ban foreign investment from the domestic market,” insists Duan Wude, deputy director of the Department of Science, Technology, and Education at the Ministry of Agriculture. “It is only being careful with the introduction of foreign technology.”

That explanation sounds to many observers like a smokescreen for a tougher trade policy, however. “By trying to protect its own research-based industries, China is creating a situation in which the real losers are the farmers,” contends Scott Rozelle, a professor of agricultural and resource economics at the University of California, Davis. “I favor a go-slow system for approving new releases, but I am against the ban on foreign investment.”

The ban on foreign investment “is more of a trade issue than a scientific issue,” says Xing-Wang Deng, director of the new Center for Plant Molecular Genetics and Agribiotechnology, which involves scientists at Yale and Beijing universities. Deng and others say that China is worried that other countries might impose restrictions on its agricultural exports if farmers begin harvesting large amounts of transgenic crops, along with traditional varieties, and they fail to segregate the two types. That has happened in the past with Chinese-made soy sauce, derived in large part from GM soybeans grown in the United States. The ban, he says, is seen as a way to limit the amount of land planted with GM crops and thus make oversight easier.

U.S.-based Monsanto is by all accounts the company most affected by the restrictions on foreign investment. It's the biggest foreign source of GM crops in China; its Bollgard Bt cotton seeds, for example, are growing in a majority of the 350,000 hectares of GM cotton cultivated this year. John Killmer, president of Monsanto China, says the new rules exacerbate an already difficult situation for the company, which has repeatedly failed to win approval for Bt corn and has abandoned an earlier collaboration with Huang Danian of the National Rice Research Institute in Hangzhou on a herbicide-resistant rice variety that is also awaiting government approval.

“The current process is so onerous that it can take 7 to 9 years to win approval,” says Killmer, noting that Monsanto has tried unsuccessfully for 6 years to introduce the same Bt corn that is now growing in the United States. “And some corn hybrids have a life of only 3 to 4 years.” Killmer says companies such as Monsanto currently have little incentive to push ahead with GM crops in China, in view of the almost-certain rejection of any application for a commercial license. He also dismisses the official explanation for the ban on new foreign investment: “The idea that it's for safety considerations doesn't wash,” he says, because those considerations should apply equally to domestic activities.

China's Bt cotton crop is the fourth largest in the world and the most successful of five species under cultivation (see table). Two species of GM rice—one herbicide tolerant and the other blast resistant—have been approved for field trials and environmental release, but Duan says the review for a commercial license could take quite a long time because of concerns about whether genes from the transgenic plants could spread to wild species.

View this table:

While these debates play out, plant scientists are benefiting from the government's growing investment in research. The current Five-Year Plan projects a fivefold rise in spending on agbiotech research, to perhaps $500 million by the end of 2005, and some three dozen Chinese institutes are working on transgenic plant research. The Beijing-Yale center, which officially opened 18 months ago, is riding that wave of support, says Deng: “We try to understand basic biology. And we focus on Arabidopsis and rice because it's a good model system as well as an important economic crop.” The recent ban on foreign investment doesn't affect Yale's participation in the center, Deng says, nor did it stop Monsanto this summer from pledging approximately$750,000 over 5 years to support the center's programs. “It's not considered an investment by a foreign company,” notes Deng about the contribution, which he says will “strengthen an exchange program and lessen our dependence on university funds.”

Despite the current impasse, both Deng and Killmer believe that properly licensed GM crops will one day be commonplace among China's 350 million peasant farmers. But they offer different reasons for their optimism. “They are facing a use-it-or-lose-it situation,” says Killmer, referring to work by Chinese scientists, including discoveries based on the sequencing of the indica rice genome, that could wind up in the hands of foreign competitors. “And that may be the thing that breaks the logjam.”

For Deng, it's a simple matter of waiting until the government decides the time is right: “In the long term, these GM crops will be approved [for commercial sale]. All of the scientists I know are optimistic. But I don't know how long it will take.”

19. ARCHAEOLOGY

# Armenia Uncovers a Bronze Age Treasure Trove

1. Richard Stone

With the help of private money, Armenian researchers are unraveling a site full of impressive stonework and ritual artifacts

AGARAK, ARMENIA—From a distance, the cliff face and its curling overhang look like a giant wave about to break over the central Armenian plain. Up close, something more spellbinding comes into view: The lip of the 15-meter bluff sports what seems to be a meter-long, v-shaped ram's head, complete with coiling horns, chiseled from the volcanic rock. Farther along the edge, in eroded bas relief, are what looks like the nostrils and horns of a bull. A third, more deeply eroded feature might have been a dragon's head.

“Just look at it, it's beautiful,” says Boris Gasparyan of the Institute of Archaeology and Ethnography (IAE) in Yerevan. In just two field seasons since excavations began here in Agarak, an Armenian team has uncovered a prodigious number of artifacts and mysterious carvings that promise to shed new light on the peoples of the Caucasus from the Early Bronze Age some 5000 years ago right up to the Middle Ages. “It's an incredibly important site,” says Adam T. Smith, an archaeologist at the University of Chicago and one of the few Western experts to have visited Agarak. “A comparatively small portion of the site has been excavated, and yet the results are quite fantastic.”

A few years ago, the site was on the verge of being lost forever. Just before the excavations started, a nearby quarry was planning to expand onto the site and extract the rock, known as tuff, a popular building material in the Caucasus. Fortuitously, Gasparyan had befriended Michael Gfoeller, then an official at the U.S. Embassy in Yerevan who also happens to be an archaeology buff. Gasparyan showed him around a number of Armenia's rich historical sites. “I realized that Armenia today is like Egypt in the 1820s: an undiscovered country of immense cultural wealth,” Gfoeller recalls. Gasparyan similarly wowed Gfoeller's brother Joachim, a wealthy businessperson, during a visit. Joachim Gfoeller was so enthralled that he set up a foundation to support archaeological research in the impoverished nation.

Agarak was an obvious first project, says Gasparyan, now the foundation's local director. The cliff is hard to miss: Part of it was blasted away during construction of a road that passes within feet of the site, and fragments of Bronze Age pottery had also been found there. But once excavations started, even Gasparyan and his colleagues were astonished by the extent of the remains. In 2001, the dig team swelled to 100 at the height of the summer season. That many workers “is unheard of in this day and age,” says anthropologist Philip Kohl, an expert on the Caucasus at Wellesley College in Massachusetts, who says the operation's scale is reminiscent of archaeology's golden age in the colonial era before World War II.

By the end of last summer, the huge team, led by IAE's Pavel Avetisyan, had peeled away the surface soil from nearly 5000 square meters of the site—still only a fraction of its claimed 200-hectare extent. “Agarak is notable for its sheer size,” says Smith, who is co-leader of an excavation of Late Bronze Age fortresses in Armenia. If it is truly that large, says Kohl, who has not yet seen it, then “understanding and evaluating the significance of the site may take decades.”

The Armenian team has found traces of intensive stone working: steps leading to niches carved into the cliffs, horseshoe-shaped cavities connected by channels cut into the tuff, and trapezoidal blocks. “The total expanse appears to have been carved, shaped, and molded to human life,” says Smith, who adds that such features “suggest some sort of ritual installations, such as altars or other monuments.”

The Armenian team has also begun unearthing the remains of stone houses clustered along a street, along with a bounty of terra cotta statuettes and ceramic artifacts linking the site to the Kuro-Araxes, a culture widespread in the Caucasus in the 29th to 27th centuries B.C. The researchers suggest that the bluff was an open-air temple, complete with housing for the priests. According to Smith, that's “an entirely reasonable starting hypothesis that will undoubtedly be subject to intense questioning as the research proceeds.”

Although it might take decades to fix Agarak's position in the Kuro-Araxes culture, the site has already begun to spark vigorous debate. Some experts doubt, for instance, whether the bas relief heads along the cliff edge were carved by human hand. “I would have some reservations about the anthropic character of many of these ‘carvings,’” says archaeologist Pierre Lombard of Lumière University in Lyon, France. Although impressed by the site and the Armenian team's work, Lombard, an expert on the ancient Middle East, suggests that the “animal heads” could possibly be an effect of erosion. Others, however, argue that weathering alone is unlikely to have produced such features.

Also mysterious are thousands of tiny round pits, ranging from the size of silver dollars up to teacup saucers, which are scattered across much of the exposed top of the bluff. “We can't explain these,” says Gasparyan, who speculates that they might have been used for anchoring posts. “They are far too regular” to have been simply from erosion, says Smith, but when they were made might be an “irresolvable issue,” he says.

If the Early Bronze Age finds aren't tantalizing enough, the Armenian team has also uncovered evidence of habitation from more recent periods: Urartian amphoras from the 8th to 6th century B.C.; coins bearing the profiles of Alexander the Great and Octavian Augustus, and sarcophagi with skeletons buried with either pagan or Christian ritual that span a period from the 4th century B.C. to the 4th century A.D.; glazed and cooking pottery from the 12th to 14th centuries; and ceramics and coins of the Khanate of Yerevan from the 17th and 18th centuries. “I cannot think of a site anywhere in the Caucasus with such a millennia-long length of occupation,” says Kohl, who is eager to see Agarak for himself. “That fact alone makes the site important and noteworthy.”

After the threat of destruction just a few years ago, Agarak's future now seems secure. The Armenian government has designated much of the site a national park, and the Gfoeller Foundation has put up $60,000 to fund the first few years of a long-term excavation. It's also funding preliminary excavations elsewhere in Armenia and claims to have found a cavern with paintings that could date from the Stone Age, as well as a site in northern Armenia with evidence of early hominids—an important find if true, but not a big surprise considering its proximity to the famed Dmanisi site in neighboring Georgia. The foundation's support, predicts Smith, “will be a magnet for researchers and students from around the world.” And that's exactly what Armenia's suddenly resurgent community of archaeologists and anthropologists is hoping for. 20. ANIMAL MODELS # Can a Mouse Be Standardized? 1. John Bohannon* 1. John Bohannon writes from Lyon, France. With new generations of mutant mice on the horizon, some researchers question the meaningfulness of standard behavioral tests and the wisdom of minimizing the mouse environment OXFORD, UNITED KINGDOM—Georgia Mason moves quietly from cage to cage, peeking in on mice dimly visible by the glow of a red lamp. “This one spends her time ‘route tracing,’” says Naomi Latham, one of Mason's graduate students at the University of Oxford, as she slides a cage from the rack. Within the shoebox-sized cage, a mouse appears to perform a slapstick comedy in fast-forward mode, chasing herself around and around in a tight circle. “She does that for hours at a time,” Latham says. Mason points out a mouse with all its whiskers missing. “Sometimes they pluck out all the fur from the face.” By filming their mice in near darkness with a sensitive camera, Latham and Mason are cataloging a variety of such repetitive behaviors collectively known as stereotypy. Studying bizarre mouse habits might seem obscure, but the need to work out which aspects of a mouse's environment cause such behavioral quirks is becoming urgent. New techniques are speeding the creation of mutant mice used to study the links between genes and behavior. But there's accumulating evidence that the typical living conditions of lab mice might induce odd behaviors, from the subtle to the profound, that might obscure genetically based differences. The result: The same experiment can have different outcomes in different labs. That point was driven home 3 years ago in a project led by John Crabbe, director of the Veterans Affairs' Portland Alcohol Research Center in Oregon (Science, 4 June 1999, p. 1599). This was the experiment no one in the field wanted to do: Mice of the same strains, born on the same day, were tested with a standard array of behavioral tests in three independent laboratories. To the horror of behavioral geneticists, the results varied wildly between labs, despite efforts to rear the animals in the same way and test them at the same time under seemingly identical conditions. Following up on this study, Crabbe and his colleagues report in the January 2003 issue of the Journal of Neuroscience that certain strains of mice are much more active in some laboratories than others, and this confounds certain behavioral tests. But the critical differences between laboratory environments remain a mystery. One ultimate aim of research such as Mason's is to figure out those differences and use that knowledge to improve standards for the mouse environment. But that will involve “a balancing act,” says Pat Nolan, a geneticist at the U.K. Medical Research Council's Mammalian Genetics Unit near Oxford. The advantage of simplifying the environment in which mice are housed and tested, says Nolan, is that results are more easily replicated. But too narrow an environment limits the range of behavioral phenotypes that mice can exhibit. What's more, Mason points out, keeping mice in cramped and barren quarters elicits behaviors such as stereotypy that interfere with gathering meaningful data. Mason argues that “noisier” environments that mimic a mouse's natural habitat—offering, for example, places to hide—could reduce noise in mouse behavior. The pharmaceutical industry is taking a keen interest in these efforts to understand the behavior of Mus musculus. Big pharma devotes most of its$30 billion research budget to diseases of the central nervous system (CNS) and uses the mouse almost exclusively as its model organism. As in other mammals, between 30% and 40% of mouse genes are responsible for the development and maintenance of the CNS. Aided by the publication of the mouse genome—which is 80% similar to our own—researchers are racing to produce mutant mice that mimic human CNS diseases.

To determine whether a particular gene is important for schizophrenia, for example, researchers compare the behavior of mice with normal or mutated forms of the gene. But this step, explains Mason, can be a stumbling block, because behavior is variable and highly sensitive to interactions between genes and environment.

Crabbe has firsthand experience with these vexing problems. He had created a strain lacking the gene for a receptor of serotonin, a neurotransmitter thought to be important in addiction, and in 1996 demonstrated that these mice drink more alcohol than other mice—a result he replicated four times. But when mice from this strain were raised elsewhere as part of the comparative study Crabbe conducted 3 years later, they had no particular fondness for alcohol. Crabbe concluded that subtle differences between laboratory environments were behind these divergent results.

Mason believes that stereotypy could be part of the explanation. Joseph Garner, a former student of Mason's who is now at the University of California, Davis, has recently shown that stereotyping C57 mice—the strain most commonly used for genetic experiments—differ profoundly from their normal-seeming cage mates, in that they get “stuck in ruts.” Garner found that stereotyping mice have difficulty switching to a new task, even when the old task goes unrewarded, Mason reported in September at the Festival of Science in Leicester, U.K. Although recognized years ago, says Mason, stereotypy is “often overlooked because it happens at night.” According to Garner and Mason, it is caused by early separation from mothers, a lack of sensory stimulation, and cages 1/20th the size of a mouse's natural territory.

Adding to the problem, even lab-bred mice are social animals. “After 4 weeks in isolation, mice become very strange. It's better to study their behaviors in a social context,” says Hans-Peter Lipp, a biologist at the University of Zürich, Switzerland. To this end, he has invented a computerized cage that keeps track of the location and behaviors of individual mice within groups (see sidebar).

Lipp admits that his approach is not amenable to the high-throughput testing of mutant mice that many researchers are calling for. According to the National Association for Biomedical Research in Washington, D.C., well over 20 million laboratory mice are produced annually in the United States alone. The numbers are expected to balloon with new breeds of mutant mice created using ethylnitrosourea (ENU), which mutates the DNA in sperm. Whereas most breeds of mutant mice have been produced by “knocking out” a specific gene, researchers are using ENU to build huge libraries of mice containing single, random mutations.

To run each mouse produced by ENU through time-consuming tests in search of often-subtle behavioral differences would be impossible, says Robert Gerlai, a biotech consultant formerly of Eli Lilly Research Laboratories in Indianapolis, Indiana, so the system needs to be automated—but just how is unclear. Large-scale behavioral testing of mutants can produce misleading results, claims Hanno Würbel of the University of Giessen, Germany, the first to observe nocturnal mouse stereotypy. Such testing is likely to use simple environments, explains Würbel, “but this is naïve. The behavioral effect of a gene should be tested in many contexts, not just a single, standard laboratory environment.”

Others are adopting these ideas. Nolan's research group is developing ENU mouse models of human disease as part of a European Union consortium formed last month called Eumorphia. One of its goals is to develop new environmental and phenotype-testing standards. The research showing that mice are exquisitely sensitive to their environments will guide the group's efforts, Nolan says, and he hopes new techniques will improve the data mining of ENU mice.

21. ANIMAL MODELS

# To Build a Better Mouse Cage

1. John Bohannon*
1. John Bohannon writes from Lyon, France.

When it comes to mouse behavior, a perennial source of experimental “noise” is the scientist herself. Mice are sensitive to differences between people, such as their handling techniques or scents. And not only do humans complicate experiments by their presence, they limit the pace of research by observing no more than one mouse performing one task at a time. “Behavioral analysis is space and time intensive,” says Robert Gerlai, a biotech consultant formerly with Eli Lilly Research Laboratories in Indianapolis, Indiana. That's why many mouse researchers would like to remove humans from the picture altogether.

One attempt to create a standard human-free mouse environment is the SmartCube, a high-tech testing environment for a single mouse being developed by PsychoGenics in Hawthorne, New York. According to its creators, by keeping track of many parameters over time—the order and duration of certain behaviors, along with physiological variables such as heart rate—the SmartCube will detect subtle effects that would be invisible to an old-fashioned human researcher.

Whereas the SmartCube observes a single mouse with better-than-human watchfulness, the IntelliCage keeps track of up to 16 mice living in the same cage. A computer follows the location of mice by means of microchips implanted in the loose skin at the back of their necks and records their interactions with various behavioral testing modules at each corner of the cage. Hans-Peter Lipp of the University of Zürich, Switzerland, created the IntelliCage to observe mouse social behavior while reducing the total number of mice needed for his research. Mice behave strangely unless given the illusion of privacy from people, says Lipp, “because, after all, humans are seen as dangerous predators.”

22. NANOTECHNOLOGY

# Biology Offers Nanotechs a Helping Hand

1. Robert F. Service

Rather than building tiny devices atom by atom, nanoscientists are raiding biology's molecular toolbox in hopes of revolutionizing sensors, medical diagnostics, and electronics

BOSTON, MASSACHUSETTS—When it comes to nanotechnology, physicists, chemists, and materials scientists can't hold a candle to the simplest bacteria. Billions of years of evolution have outfitted organisms of all stripes with a wealth of nanomachines—from the information-storage medium of DNA to the proteins that capture sunlight and copy DNA during cell division. Early nanotech visionaries dreamed of crafting their own versions of nanomachinery and even went so far as to draw up molecular specs for tiny gears and motors. But at the Materials Research Society meeting here earlier this month,* it was clear that as nanotechnology begins to leave its infancy and find its feet, most nanobuilders are looking to biology not just for inspiration but also a little practical help.

In labs around the globe, researchers are working to marry biology and nanotechnology, fusing useful biomolecules to chemically synthesized nanoclusters in arrangements that do everything from emitting light to storing tiny bits of magnetic data. The result is a merger that attempts to blend biology's ability to assemble complex structures with nanoscientists' capacity to build useful devices.

That merger hasn't created any Fortune 500 companies yet. But for basic researchers, blending biology and nanotech is a white-hot field. “It's gaining huge momentum right now,” says Chad Mirkin, a chemist and nanoscience expert at Northwestern University in Evanston, Illinois. And that momentum, say many researchers, could lead to the development of a wide range of applications, from novel medical diagnostics and sensors to data-storage materials and nanomanufacturing tools. “There are a lot of ideas emerging right now,” says Günter Schmid, a chemist at the University of Essen, Germany.

One of the biggest drivers behind nanotechnology's enthusiasm for biological systems revolves around an organism's impressive ability to manufacture complex molecules such as DNA and proteins with atomic precision. Chemists create molecules up to hundreds of atoms in size without too much trouble, controlling the position of every atom. But beyond that, traditional synthetic schemes become unwieldy and too inefficient to be practical. Computer chip engineers—the most advanced materials makers—do much better. They can craft chips with 200 million transistors, each with features on the order of 100 nanometers, or 100 billionths of a meter.

“There's a big gap in between those two,” says Joseph Jacobson, a physicist at the Massachusetts Institute of Technology (MIT) in Cambridge—but not for biology. “The champions of that [size range] are the pieces of machinery of biology, DNA and proteins.” So as a stopgap, nanotechnologists are looking to see what they can appropriate. “There is an incredible toolbox [of biomolecules] that we can incorporate for our own ends,” says Paul Alivisatos, a chemist at the University of California, Berkeley.

Complex biological machines also show an uncanny knack for homing in on and binding to molecular targets amid a sea of other molecules. “Biomachinery is a powerful way of bringing organization into a system,” notes Keith Williams, a physicist and nanotech expert at the Delft Institute of Technology in the Netherlands. By contrast, engineered nanosized objects such as carbon nanotubes and tiny spherical metal and semiconducting particles lack any guidance mechanism. That makes it extremely difficult to put those tubes and particles where you want them to go. “As materials become so small, they become difficult to handle with traditional methods such as lithography,” the technique used to pattern computer chips, says Williams.

As a result, until researchers learn to construct complex nanostructures from the ground up, they have little choice but to become small-time thieves. “Instead of trying to build [nanostructures] from scratch, let's just steal them from biology,” says Jacob Schmidt, a bioengineer at the University of California, Los Angeles. A handful of nanotech research groups has been perpetrating such theft in recent years, Mirkin and others say, but now the nano field is in the midst of a kleptomania epidemic.

## Bioelectronic assembly

At the meeting, for example, Williams described an emerging effort to harness the selective binding capabilities of a chemical relative of DNA called peptide nucleic acid (PNA) to assemble carbon nanotubes into molecular-scale electronic devices. Williams is a postdoc in the lab of Cees Dekker, whose team reported making the first nanotube-based transistor in 1998. To construct that device, Dekker's team scattered nanotubes across a surface patterned with tiny gold electrodes and then used an atomic-resolution microscope to find a lone nanotube draped across two electrodes. The team then measured how much current flowed through the nanotube when a voltage applied to the material beneath it made the nanotube more conductive—the essence of a transistor.

But randomly scattered nanotubes can't be the scaffold for more-complex molecular-scale circuitry. So Dekker and Williams have turned to PNA, a molecule that, like DNA, is made up of a series of nucleotide bases (A's, T's, G's, and C's) that bind selectively to one another. The difference is that PNAs replace DNA's backbone of sugar and phosphate groups with more stable links based on peptides. That substitution enables PNAs to withstand higher temperatures and solvents often used to process nanotubes.

Williams started with bundles of nanotubes that he placed in a bath of nitric acid. The acid slowly ate away at the nanotubes, digesting them from the ends in toward the center. After a few hours, Williams removed the acid and was left with short tubes, each studded with carboxylic acid groups at both ends. These groups readily react with amide groups on the ends of the PNA molecules, making it possible for Williams to link PNAs to the tips of each nanotube. Williams then spiked the nanotube-PNA mixture with DNA strands harboring nucleotide sequences complementary to those in the PNAs and confirmed that the partners found their mates, a result published in this week's issue of Nature. Next, Williams says he plans to attach single-stranded DNA to electrodes. The DNA should bind to the PNAs and direct the nanotubes to bridge electrodes, the first step in assembling nanoscale circuitry.

The Delft group has plenty of company in turning to DNA for its assembly skills. Mirkin reported that his team at Northwestern is making progress on using DNA to make sensors and a new nanomanufacturing platform. In 1997, Mirkin's group showed that it could detect free-floating target DNA strands with complementary DNA linked to tiny gold nanoparticles. When the target strands bind to the gold-bound complementary strands, they pack the gold particles close together. That changes the color the particles reflect, creating a simple color-based detector for specific DNA sequences.

Mirkin's DNA detection scheme is already finding applications. At the meeting, James Storhoff, a chemist at Nanosphere—a Northbrook, Illinois-based start-up founded by Mirkin and his Northwestern colleague Robert Letsinger—reported that the company has turned it into a bench-top tool for rapid diagnoses of infectious diseases and detection of tiny genetic mutations called single-nucleotide polymorphisms. Storhoff says the diagnostic scheme can readily distinguish antibiotic-resistant bacteria, such as Staphylococcus aureus, from nonresistant strains, a development that could cut in half the time needed to alert doctors to the status of a patient's infection. Storhoff noted that the detection scheme is so sensitive that it can pick up DNA from a target bacterium without first amplifying the DNA by the now-standard polymerase chain reaction, a shortcut that could speed up detection rates even more. Eventually, the advances might enable doctors to diagnose infections in hours rather than waiting for days as samples are processed in a lab. It could also speed the detection of smallpox and other potential bioweapons and give emergency workers a leg up in containing an outbreak before it becomes widespread.

## Biomechanics

Mirkin's group is also marrying DNA with tiny mechanical devices in hopes of revolutionizing nanomanufacturing. In 1999, the researchers reported developing a technique called dip pen nanolithography, in which they use the tip of an atomic-force microscope to write nanosized letters or other features on a surface (Science, 15 October 1999, p. 389). Later, they showed that by using a DNA-based “ink,” they could write an initial pattern in DNA and then use it to bind complementary DNA sequences toting nanoparticles. The technique carried the promise of patterning materials at fantastic resolution. But it was painfully slow, because features had to be written out one by one.

No longer. At the Materials Research Society meeting, Mirkin reported that his team had constructed an array of 10,000 microscope tips, each capable of acting independently from the others. Team members reported that by using 10 tips in concert they can draw essentially any desired shape. “The goal is to use dip pen nanolithography to generate [DNA-based] templates on surfaces that guide the assembly of nanoscale building blocks,” Mirkin says. “It opens the door to placing electronic particles right where you want them. We think it's ultimately going to be a production tool.” What's more, Mirkin says there's still room to grow: “This is not going to stop at 10,000. We can make arrays of arrays.”

Like Mirkin, MIT's Jacobson has linked gold nanoparticles to DNA and other biomolecules. His aim, though, is to disrupt the capabilities of biomolecules to recognize targets. Earlier this year, Jacobson's team reported that by guiding the nanoparticles with a simple radio-frequency (RF) transmitter, they can control whether DNA fragments exist as two separate single strands or as bound pairs. With the RF field turned on, the gold nanoparticles spin, heating the associated DNA enough to melt the bonds between complementary strands. Turn the RF field off, and the strands knit themselves back together. Although still in its early stages, the technique promises to provide researchers with an electronic switch that they can use to turn genes on and off.

At the Boston meeting, Jacobson announced that his team had extended its electrical control over biomolecules to proteins as well. The researchers started with an RNA-chopping protein called ribonuclease (RNAase), which they cut into two pieces: a large protein segment made up of 104 amino acids, and a small 18-amino-acid strand called the S-peptide. The RNA-chewing machine couldn't do its job unless the small strand sat in the mouth of the protein. Jacobson's team linked gold nanoparticles to the end of S-peptide strands and used the particles as a switch to turn the enzyme on and off. In the absence of an RF field, the nanoparticle-toting S-peptides adopt their usual conformation, allowing RNAase to do its job. But when the team switched on an external RF field, the field set the nanoparticles twirling, which prevented them from assembling with the larger protein.

Down the road, Jacobson says he hopes that electronically controlled proteins and DNAs will enable molecular biologists to cut and splice genetic information electronically and eventually program computers to engineer new organisms. “It's [the] early days,” says Jacobson. “But we're trying to feel our way to see how complex systems we can build and to see if we can get biology to build these systems for us.” Many other groups are starting to feel their way along as well. Where they succeed, expect the marriage of nanotech and biology to provide big payoffs.

• *2002 Fall Meeting, 2 to 6 December.