Special Perspectives

Life with Oxygen

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Science  05 Oct 2007:
Vol. 318, Issue 5847, pp. 62-64
DOI: 10.1126/science.1147949


The survival of all metazoan organisms is dependent on the regulation of O2 delivery and utilization to maintain a balance between the generation of energy and production of potentially toxic oxidants. Hypoxia-inducible factor 1 (HIF-1) is a transcription factor that functions as a master regulator of oxygen homeostasis and has essential roles in metazoan development, physiology, and disease pathogenesis. Remarkable progress has been made in delineating the molecular mechanisms whereby changes in cellular oxygenation are transduced to the nucleus as changes in gene transcription through the activity of HIF-1. Pharmacologic agents that activate or inhibit the hypoxia signal transduction pathway may be useful therapies for ischemic and neoplastic disorders, respectively, which are the major causes of mortality in industrialized societies.

Life with oxygen began some 2.5 billion years ago with the evolution of organisms capable of transducing solar energy into the chemical energy of carbon bonds. In this process of photosynthesis, carbon dioxide and water are converted into glucose, with O2 generated as a side product of the reaction. Photosynthetic organisms prospered and multiplied, leading to a progressive increase in atmospheric O2 concentrations. About 1.5 billion years ago eukaryotic organisms appeared containing mitochondria, subcellular organelles in which glucose is oxidized to carbon dioxide and water, thereby completing the energy cycle. Reducing equivalents are generated that pass through the mitochondrial respiratory complex, which results in the formation of a proton gradient that is used to drive the synthesis of adenosine 5′-triphosphate (ATP). A third landmark, the appearance of the first metazoan organisms, was attained ∼0.5 billion years ago. Just as the evolution of eukaryotes was dependent on the prior establishment of photosynthesis, metazoan evolution was dependent on the highly efficient recovery of energy contained within the chemical bonds of glucose through the process of oxidative phosphorylation, which, compared with glycolysis, produces 18 times as much ATP per mole of glucose and thus provides the energy necessary for developing and maintaining complex multicellular organisms.

The utilization of O2 as a substrate for energy production is not without risk. The electrons transferred through the mitochondrial respiratory chain ultimately react with O2 to form H2O, a process that is catalyzed by cytochrome c oxidase (complex IV). However, a fraction of electrons escape the respiratory chain and combine with O2 prematurely, resulting in the generation of superoxide anion, which is converted to hydrogen peroxide by the action of superoxide dismutase. The oxidation of lipids, nucleic acids, and proteins by these reactive oxygen species (ROS) can result in cellular dysfunction or death. Acute increases or decreases in the cellular O2 concentration (hyperoxia and hypoxia, respectively) result in generation of excess ROS (1). This finding implies that efficient respiratory chain function occurs within a narrow range of O2 concentrations (2). Hypoxia may also result in deficient ATP production due to substrate limitation. All eukaryotic organisms must maintain oxygen homeostasis, and this requirement is a critical organizing principle of metazoan evolution and biology, as described in detail below.

The transcription factor HIF-1 (hypoxia-inducible factor 1) has an essential role in the maintenance of oxygen homeostasis in metazoan organisms. Within any given cell type, HIF-1 controls the expression of hundreds of genes (2), and because the battery of target genes varies considerably from one cell type to another, the complete HIF-1 transcriptome is likely to include thousands of genes. HIF-1 is a heterodimer composed of an O2-regulated HIF-1α subunit and a constitutively expressed HIF-1β subunit (2). HIF-1α is continuously synthesized and degraded under normoxic conditions, whereas under hypoxic conditions, HIF-1α degradation is inhibited, the protein accumulates, dimerizes with HIF-1β, binds to cis-acting hypoxia-response elements in target genes, and recruits coactivator proteins, all of which leads to increased transcription (Fig. 1A). An important element of complexity derives from the HIF-1α paralogue HIF-2α, which is also O2-regulated, dimerizes with HIF-1β, and activates transcription of an overlapping but distinct set of target genes (3). Another paralogue, HIF-3α, appears to function as an inhibitor of HIF-1α (4). Establishing the specific roles of HIF-1α, HIF-2α, and HIF-3α in oxygen homeostasis is a major challenge of current research.

Fig. 1.

Molecular mechanisms of oxygen homeostasis. (A) O2-dependent posttranslational modifications of the HIF-1α subunit by PHD2 and FIH-1 (not shown) serve as molecular switches that regulate the interaction of HIF-1α with VHL and p300, which, in turn, determine the half-life and transcriptional activity of HIF-1, respectively. Two additional proteins form multivalent complexes that increase the efficiency of these reactions: OS-9 (a protein that was originally identified in osteosarcomas) interacts with HIF-1α and PHD2 and promotes hydroxylation, whereas SSAT2 (a paralog of spermidine/spermidine acetyltransferase-1) interacts with HIF-1α, VHL, and Elongin C and promotes ubiquitination. Finally, pharmacological inhibitors of HSP90, such as 17-allylaminogeldanamycin (17-AAG), promote the binding of HIF-1α to RACK1, which recruits the Elongin C ubiquitin-ligase complex in an O2-independent manner. Blue, red, and green arrows denote processes that occur under hypoxic, normoxic, or O2-independent conditions, respectively. Additional factors regulating HIF-1 are described in the STKE Connections Map (17). (B) Under hypoxic conditions, HIF-1 induces the expression of genes encoding the following proteins: (1) glucose transporters GLUT1 and GLUT3, which increase intracellular glucose uptake; (2) glycolytic enzymes, which convert glucose into pyruvate; (3) lactate dehydrogenase A, which converts pyruvate into lactate; (4) pyruvate dehydrogenase (PDH) kinase 1, which phosphorylates and inactivates PDH, the enzyme that converts pyruvate into acetyl CoA for entry into the mitochondrial tricarboxylic acid (TCA) cycle, which generates reducing equivalents that are passed on to the electron transport chain (ETC); and (5) cytochrome c oxidase subunit COX4-2, which replaces COX4-1 and, thereby, increases the efficiency of mitochondrial respiration under hypoxic conditions.

O2-dependent degradation of HIF-1α is triggered by binding of the von Hippel–Lindau tumor-suppressor protein (VHL), which interacts with the protein Elongin C, thereby recruiting an E3 ubiquitin–protein ligase complex that ubiquitinates HIF-1α and targets it for degradation by the 26S proteasome. VHL binding is dependent on the hydroxylation of proline residue 402 or 564, or of both residues, by the prolyl hydroxylase PHD2, which is a dioxygenase that uses O2 and α-ketoglutarate as substrates and generates CO2 and succinate as by-products (58). PHD1 and PHD3 also hydroxylate HIF-1α when overexpressed, but their physiological functions have not been established. PHD2 activity is reduced under hypoxic conditions either as a result of substrate limitation (7) or as a result of inhibition of the catalytic center [which contains Fe (II)] by ROS generated at complex III of the mitochondrial respiratory chain (1). ROS levels increase in response to hyperoxia, but HIF-1α levels do not, which suggests that the site of ROS generation may be different in hyperoxic cells or that ROS generation by complex III is necessary, but not sufficient, to induce HIF-1α under hypoxic conditions. FIH-1 (factor inhibiting HIF-1) is another dioxygenase that hydroxylates asparagine residue 803 of HIF-1α and, thereby, blocks its interaction with the coactivator p300 (9). The half-life of HIF-1α is also regulated in an O2-independent manner by the competitive binding of either heat shock protein HSP90, which stabilizes the protein, or RACK1, which interacts with Elongin C and, thereby, promotes HIF-1α ubiquitination and degradation that is independent of PHD2 and VHL (10). A second major O2-independent regulatory mechanism is the stimulation of HIF-1α protein synthesis by signal transduction via phosphatidylinositol 3-kinase, protein kinase B (AKT), and mammalian target of rapamycin (11).

HIF-1 is present in the roundworm Caenorhabditis elegans (7), which consists of fewer than one thousand cells and is so small and simple that all cells obtain O2 by direct diffusion from the atmosphere. However, O2 becomes limiting when the worms burrow into the soil in search of nutrients. The resulting hypoxia induces the HIF-1–mediated expression of genes encoding glucose transporters and glycolytic enzymes, which catalyze the anaerobic production of ATP through the fermentation of glucose to lactic acid (Fig. 1B). In mammalian cells, HIF-1 also regulates synthesis of mitochondrial acetyl coenzyme A (acetyl CoA), subunit composition of cytochrome c oxidase, and mitochondrial biogenesis (2, 12). The evolution of larger and more complex metazoans required the concomitant evolution of anatomic structures and physiological mechanisms designed to ensure the delivery of optimal concentrations of O2 to every cell. In the fruit fly Drosophila melanogaster, this is accomplished by tracheal tubules through which O2 is transported to the interior of the organism. In mammals, such as the laboratory mouse Mus musculus, complex respiratory and circulatory (heart, blood, and vasculature) systems evolved to deliver O2 to each of the trillions of cells that compose the adult organism. The development of these systems before birth and their use after birth are controlled by HIF-1. Thus, HIF-1 mediates developmental and physiological pathways that either promote O2 delivery to cells or allow cells to survive O2 deprivation (Fig. 1).

In the case of Homo sapiens, the ability of the species to prosper and multiply over the last 0.0001 billion years has been associated with an increase in life span. Humans in industrialized societies are less likely to die of infection, predation, or starvation and more likely to die of cardiovascular and neoplastic disorders in which aging, dietary excess, and physical inactivity play important etiological roles. Atherosclerotic stenosis of large arteries in the coronary and femoral circulations results in myocardial and limb ischemia, respectively, conditions in which cells are deprived of O2 and glucose and accumulate toxic metabolites. The deprivation of O2 in ischemic cells induces adaptive homeostatic responses in young healthy experimental animals, such as the increased production of vascular endothelial growth factor and other angiogenic cytokines, which promote tissue perfusion. These processes appear to be blunted in humans as a result of aging, atherosclerosis, cigarette smoking, diabetes, and hypertension (13). A major challenge of current research in this field is to understand the mechanisms underlying the impairment of O2 homeostasis and to devise therapeutic strategies to counteract them. In contrast, cancer cells co-opt adaptive mechanisms mediated by HIF-1 to promote their survival, proliferation, perfusion, and invasion of body tissues (14, 15), and clinical trials of therapeutic strategies designed to block these pathways are ongoing.

The last landmark on our timeline, a mere 0.0000002 billion years ago, is the discovery of oxygen, which has been called the most important discovery in the history of science (16). Life, after all, depends on it.

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