PerspectiveMicrobiology

Pathogenic Bacteria Prefer Heme

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Science  10 Sep 2004:
Vol. 305, Issue 5690, pp. 1577-1578
DOI: 10.1126/science.1102975

Almost all cells and organisms require iron to facilitate basic cellular processes such as respiration and DNA biosynthesis [HN1]. Diverse and complex iron-uptake systems have evolved throughout the biological world to provide iron for numerous proteins, particularly those involved in energy capture and oxygen transport. Indeed, in parts of the great oceans, a complete lack of iron profoundly limits bacterial growth (1, 2). In most environments, however, iron uptake is limited not by its absence, but by the fact that it is insoluble and inaccessible. To facilitate iron uptake, free-living bacteria and fungi have adopted several strategies. Some secrete compounds known as siderophores [HN2] that solubilize and bind to an external source of iron with high affinity; the iron-siderophore complexes are then imported into the bacteria by specific transporter proteins. Others have uptake systems that import free iron salts directly (3, 4). To combat microbial infections, animals strictly limit the availability of free iron in their blood and tissues. They do this by ensuring that iron in blood and secretions is carried by the high-affinity iron-binding proteins transferrin and lactoferrin [HN3], which create a primary line of defense against infection termed the “iron blockade” (5). The two ferric iron-binding sites of transferrin are rarely fully saturated, and an excess of unsaturated transferrin (apo-transferrin) ensures that free iron is virtually eliminated from blood. Pathogenic microorganisms, therefore, must overcome major obstacles if they are to acquire iron and thrive in their animal hosts. On page 1626 of this issue, Skaar and colleagues (6) [HN4] explore how the pathogenic bacterium Staphylococcus aureus [HN5] acquires the iron that it needs for growth in two different animal hosts. By growing the bacteria in the presence of the two principal iron sources found in mammals—diferric transferrin and the iron-porphyrin heme [HN6]—the investigators show that most of the iron taken up by S. aureus during the initial phases of infection is obtained from heme.

Although most bacteria are unable to grow in media in which the only iron sources are transferrin-iron and heme, some bacteria have developed strategies that enable them to obtain iron from these two sources (7, 8). To determine whether S. aureus prefers transferrin-iron or heme as a source of iron, Skaar and colleagues labeled heme with 54Fe and transferrin with 57Fe, two rare and stable iron isotopes (9). After establishing that S. aureus grew well in medium supplemented with equimolar amounts of 54Fe heme and 57Fe transferrin, they analyzed the isotope content of cells using inductively coupled plasma- mass spectrometry (ICP-MS) [HN7] at various times during growth. They discovered that S. aureus markedly prefers a source of iron derived from heme. Analysis of the S. aureus genome [HN8] revealed that it encodes seven putative membrane transporter proteins that have some homology to known bacterial iron transporters. Using mutational inactivation and ICP-MS to monitor uptake of heme iron, the investigators identified a heme transport system in S. aureus composed of three genes (hts A, B, and C) that show homology with known heme transporter genes in other bacteria (Yersinia enterocolytica and Corynebacterium diphtheriae). Moreover, they identified a binding site for the bacterial ferric-uptake repressor protein, Fur, immediately upstream of the HtsA initiation codon, implying that the hts system is switched on in response to iron deficiency.

Although a heme-uptake system is of potential value, it would be of little use if S. aureus did not also possess mechanisms for liberating heme from the red blood cells where it is packaged in the form of hemoglobin (figure). S. aureus produces multiple hemolysins [HN9] that breach the red cell membrane, promoting osmotic lysis of the cells (10). Once hemoglobin is released, it is not clear whether S. aureus liberates heme from hemoglobin with specific IsdB and IsdA enzymes (11), by secreting proteases like Vibrio vulnificus (12), or by oxidizing hemoglobin to promote its spontaneous dissociation into globin and heme (13). The bacteria then import the free heme, which is catabolized by the bacterial enzymes IsdG and IsdI in the same way as mammalian heme oxygenases catabolize heme, resulting in the release of iron from the heme porphyrin ring (14). Thus, S. aureus is a versatile pathogen that liberates heme from a vast erythrocyte repository, imports heme across its bacterial membrane, and degrades it to yield free iron (figure).

Bloodletting explained.

S. aureus bacteria obtain most of the iron (Fe) that they need for growth in mammalian hosts from an iron-containing porphyrin ring called heme. S. aureus produces hemolysins that lyse red blood cells containing heme in the form of hemoglobin. It is unclear how the bacteria break down the released hemoglobin to heme, but the bacterial enzymes IsdA and IsdB may be involved. The bacteria then import heme via transporter proteins encoded by the hts ABC operon. The heme is then catabolized by the heme oxygenase-like enzymes, IsdG and IsdI, with the release of iron and biliverdin (a breakdown product of the porphyrin ring). The free iron released from heme is used to fuel further bacterial growth. The practice of bloodletting in the pre-antibiotic era may have been an attempt to starve pathogenic bacteria of the iron that they need for growth.

CREDIT: KATHARINE SUTLIFF/SCIENCE

If the ability to take up iron is a potent virulence factor and the hts system is a major determinant of iron uptake, then mutational inactivation of the hts genes should attenuate S. aureus virulence. The authors analyze the pathogenicity of mutant and wild-type S. aureus in two model systems: the worm Caenorhabditis elegans and the mouse. Mutations in the Hts B and C genes markedly decreased mortality in worms infected 48 hours previously, and abscess formation markedly decreased in the livers and kidneys of mice 96 hours after intravenous injection of mutant compared to wild-type S. aureus. These results strongly imply that heme is the major source of nutrient iron in the critical early stages of S. aureus infection.

In response to bacterial infection and inflammation, humans restrict iron uptake and sequester iron within macrophages throughout the body [HN10]. The peptide hormone hepcidin orchestrates these changes and causes a substantial decrease in serum iron levels (15). This hypoferremic response may be important for host defense by making iron even less available than usual to invading pathogens. The protective effects of hypoferremia may explain the mystery of why physicians embraced bloodletting [HN11] as a therapeutic procedure for more than 2500 years. As recently as 1942, Sir William Osler's highly regarded medical textbook advocated bloodletting as a treatment for acute pneumonia: “To bleed at the onset in robust healthy individuals in whom the disease sets in with great intensity and harsh fever is good practice” (16). The development and widespread use of antibiotics in the mid-20th century obviated the need to employ questionable treatments such as bloodletting. However, the discovery that S. aureus depends on heme iron for growth in its animal hosts suggests that bloodletting in the pre-antibiotic era may have been an effective mechanism for starving bacterial pathogens of iron and slowing bacterial growth.

Bacteria continue to discover new ways to combat antibiotics and the race is on to discover new therapeutic targets to combat bacterial infection. The heme-uptake proteins of S. aureus may represent a new target for molecular therapy. Efficient lysis of erythrocytes increases the concentration of iron available to S. aureus by 100-fold compared to the normal concentration of transferrin-iron in serum. This liberated heme-iron apparently fuels the rapid growth of virulent S. aureus infection. Thus, pathogenic S. aureus applies principles of logic similar to those of accomplished bank robbers: They go for the heme, because that's where the iron is.

HyperNotes Related Resources on the World Wide Web

General Hypernotes

Dictionaries and Glossaries

Microglossary is a glossary of microbiology terms and abbreviations made available by Hardy Diagnostics.

The On-line Medical Dictionary is provided by CancerWeb.

A searchable medical dictionary is provided by Medline Plus.

A life sciences dictionary is provided by BioTech, an online educational and resource tool provided by the University of Texas Institute for Cellular and Molecular Biology.

A microbial genetics glossary is provided by S. Maloy, Department of Biology, San Diego State University, for a microbial genetics course.

Web Collections, References, and Resource Lists

The WWW Virtual Library: Microbiology and Virology is maintained by Scott Sutton of The Microbiology Network.

Microbes.info is a comprehensive microbiology information portal.

The Wellcome Trust Sanger Institute provides sequence data for Staphylococcus aureus and other microbial pathogens.

The Yahoo! Web Directory provides links to Internet resources related to microbiology and microorganisms.

Online Texts and Lecture Notes

Medical Microbiology, 4th edition, edited by S. Baron, is an online textbook made available by the University of Texas Medical Branch.

The Microbiology and Immunology Online Textbook is made available by the University of South Carolina School of Medicine.

Todar's Online Textbook of Bacteriology is made available by Kenneth Todar, University of Wisconsin, and includes a chapter about Staphylococcus.

The U.S. Food and Drug Administration's Bad Bug Book provides basic facts regarding foodborne pathogenic microorganisms and natural toxins.

B. Grant, Department of Microbiology and Immunology, University of Leicester, offers lecture notes for an introductory microbiology course.

The Microbiology Textbook is made available by T. Paustian, Department of Bacteriology, University of Wisconsin-Madison.

The World Lecture Hall, a large collection of online courses, offers a section on microbiology.

General Reports and Articles

The Annual Review of Microbiology, vol. 54, 2000, had an article titled “Iron metabolism in pathogenic bacteria” by C. Ratledge and L. G. Dover.

The May-June 1999 issue of the CDC's Emerging and Infectious Diseases had an article by E. D. Weinberg titled “Iron loading and disease surveillance.”

An article titled “Structural biology of bacterial iron uptake systems” by T. E. Clarke et al. is made available by Bentham Science Publishers.

“Heme and Iron” from NetBiochem provides basic information about iron utilization and heme synthesis and breakdown.

The September-October 2000 issue of the CDC's Emerging and Infectious Diseases had an article by G. M. Weinstock titled “Genomics and bacterial pathogenesis.”

Numbered Hypernotes

1. Iron in biology. The WebElements Periodic Table contains extensive information on the history, uses, and properties of iron. The Linus Pauling Institute's Micronutrient Information Center provides information about biological role of iron in human health. Alvin Crumbliss, Duke University, provides a tutorial on iron in biology. The Ferritin Molecular-Graphics Tutorial, made available by the Department of Chemistry at the University of Washington in St. Louis, considers iron in biology, focusing on the iron content in ferritin, the iron-storage protein. A tutorial provided by the Information Center for Sickle Cell and Thalassemic Disorders offers articles on a variety of iron-related topics including iron absorption, tranferrin and iron kinetics, and iron and infection.

2. Siderophores. D. Stack, Department of Chemistry, Stanford University, provided lecture notes on siderophores. A review article by J. B. Neilands in the 10 November 1995 Journal of Biological Chemistry discussed the structure and function of microbial iron transport compounds. The January 2004 issue of Infection and Immunity had a research article by Dale et al. titled “Role of siderophore biosynthesis in virulence of Staphylococcus aureus: Identification and characterization of genes involved in production of a siderophore.”

3. Transferrin and lactoferrin. Transferrins, a comprehensive web site maintained by Lisa Lambert at Chatham College, includes links to sequence information and other transferrin resources. A summary of lactoferrins is provided by Tej Singh from the All India Institute of Medical Sciences. A GeneCard for human transferrin is provided by the Weizmann Institute of Science. Transferrin is defined in the Medterms online medical dictionary. Transferrin and lactoferrin are defined in the Wilkipedia Encyclopedia. An overview of transferrin and iron kinetics is provided by the Information Center for Sickle Cell and Thalassemic Disorders.

4. Eric Skaar, Olaf Schneewind, Bae Taeok, and Kristin DeBord are in the Department of Molecular Genetics and Cell Biology, University of Chicago. Munir Humayun is at the National High Magnetic Field Laboratory, Florida State University.

5. Staphylococcus aureus. Todar's Online Textbook of Bacteriology includes a chapter on Staphylococcus. A fact sheet on Staphylococcus aureus can be found in the U.S. Food and Drug Administration's Bad Bug Book. An article about S. aureus infection is made available on the eMedicine Web site. S. Baron's Medical Microbiology textbook includes a chapter on Staphylococcus by T. Foster. Medline Plus offers a fact sheet on Staphylococcal infections.

6. Heme and other porphyrins. The Wilkipedia free encyclopedia contains a page on heme. The Porphyrin Page was created by S. Leung at Washburn University and provides basic information about porphyrins and related compounds. Porphynet provides links to a variety of porphyrin-related resources. Heme and Iron is a comprehensive online tutorial made available by NetBiochem. M. King, at the University of Indiana School of Medicine, offers lecture notes on heme and porphyrin metabolism from a medical biochemistry course. J. Diwan, Rensselaer Polytechnic Institute offers course notes on heme synthesis. A tutorial on heme and hemoglobin is made available by the Department of Chemistry at Washington University in St. Louis.

7. Inductively coupled plasma-mass spectrometry (ICP-MS). An introduction to ICP-MS is provided by the University of Missouri-Columbia Research Reactor Center. The FreeDictionary.com provides a definition of ICP-MS. “A Guide to Inductively Coupled Plasma Mass Spectroscopy” is a comprehensive feature provided by Spectroscopy magazine.

8. S. aureus genome. Published sequences of two clinical strains of S. aureus are made available by the Sanger Institute.

9. Hemolysins. Hemolysins are defined in the Medical.WebEnds.com medical terminology dictionary. G. Songer, Department of Veterinary Science and Microbiology, University of Arizona, offers lecture notes on hemolysins of S. aureus.

10. Iron and infection. J. Brock and V. Mulero provide symposium notes on iron metabolism in macrophages from a presentation at the 6th Internet World Congress for Biomedical Sciences. The May-June 1999 issue of the CDC's Emerging and Infectious Diseases had an article by E. D. Weinberg titled “Iron loading and disease surveillance.” An article by N.C. Andrews in the 1 May 2004 issue of The Journal of Clinical Investigation discussed the role of hepcidin in the anemia of inflammation. The MedLine Plus Medical Encylopedia has an entry for anemia.

11. Bloodletting. Bloodletting is defined in the Medicine Encyclopedia at AllRefer.com. The UCLA Biomedical Library hosts an online exhibit on bloodletting. “Bloodletting Antiques,” an article by Dr. Douglas Arbittier, includes images of various bloodletting devices. The PBS feature “Red Gold: The epic story of blood” includes a brief history of bloodletting.

12. Tracey A. Rouault is in the Section on Human Iron Metabolism, Cell Biology and Metabolism Branch, at the National Institute of Child Health and Human Development, NIH.

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