The 2019 Nobel Prize in Physiology or Medicine has been awarded jointly to William G. Kaelin Jr., Sir Peter J. Ratcliffe and Gregg L. Semenza “for their discoveries of how cells sense and adapt to oxygen availability.”
Gregg Semenza of Johns Hopkins University School of Medicine, William Kaelin of Dana Farber Cancer Institute in Boston, and Peter Ratcliffe of the Francis Crick Institute in London made groundbreaking discoveries relating to the HIF system, proteins that fine-tune cells’ response to oxygen.
In the early 1990s, Semenza and Ratcliffe were trying to understand how oxygen deprivation—hypoxia—triggers the erythropoietin gene to generate the hormone erythropoietin and boost red blood-cell production. They identified a DNA sequence that is necessary for hypoxia-dependent activation and placement of this DNA stretch next to other genes renders those genes inducible by low-oxygen conditions too. Semenza showed that protein from the nucleus sticks to this DNA-control region, but only when oxygen is scarce and furthermore, sequence alterations that eliminate protein binding to DNA also obliterate hypoxia-spurred gene stimulation. He proposed that the nuclear protein, which he called Hypoxia-Inducible Factor 1 (HIF-1), sits down on DNA when cells lack oxygen and rouse adjacent genes. Semenza and his postdoctoral fellow, Guang Wang, achieved a major breakthrough in 1995 when they purified HIF-1 and found that it contains two protein partners, HIF-1α and HIF-1β. The HIF-1α component was novel, and they isolated its gene from human cells. HIF-1α vanished quickly when Semenza shifted cells from low- to high-oxygen conditions. Semenza and Ratcliffe subsequently showed that multiple kinds of cells use HIF-1 to goad numerous genes in response to low-oxygen conditions and the list of hypoxia-induced genes expanded rapidly. In 1996, for instance, Semenza demonstrated that HIF-1 activates the gene for a key participant in blood-vessel formation, vascular endothelial growth factor (VEGF). This result extended HIF-1 into another key system by which the body can augment oxygen delivery. These and other findings established that HIF-1 lies at the core of an elaborate physiological network that ensures advantageous responses to oxygen.
The observation that amount of HIF-1 plummets when cells are shifted to high-oxygen conditions squared with the factor’s hallmark ability to activate target genes only when oxygen is limited, also raised a crucial question of what exactly drove HIF-1 destruction?
The answer came from an unexpected direction. A familial cancer syndrome called von Hippel-Lindau (VHL) disease owes its pathology to faulty versions of a particular protein. William Kaelin was trying to figure out how defects in this VHL protein cause the illness. The classic VHL tumor is composed of inappropriate, newly formed blood vessels, and surplus VEGF characterizes these growths; excessive erythropoietin production also can occur. Kaelin wondered whether VHL influences activity of the genes for these and other hypoxia-induced substances. In 1996, he and his colleagues grew human cells with and without operational VHL. Then they measured the abundance of several messenger RNAs, including VEGF’s, that normally disappear in response to oxygen. Even when oxygen was plentiful, VHL-defective cells contained large amounts of these mRNAs. Addition of intact VHL restored normal hypoxia-dependent quantities. Kaelin then showed that VHL’s capacity to quash the accumulation of particular mRNAs in rich oxygen conditions relies on its ability to physically assemble with several other proteins, including one that was later shown to mark certain proteins for destruction by attaching the chemical tag ubiquitin, the signal that sends certain proteins to the cell’s incinerator, the proteasome. Soon it was established that HIF-1α is degraded through the ubiquitin pathway when oxygen is profuse and in 1999, Ratcliffe and colleagues demonstrated that the oxygen-dependent elimination of HIF-1α depends on VHL. Ratcliffe further proposed that VHL interacts with HIF-1α under high-oxygen conditions and targets it for destruction, and Kaelin subsequently found that VHL binds directly to HIF-1α—and that optimal binding requires a region in HIF-1α that was known to be needed for its oxygen-triggered destruction. Kaelin and Ratcliffe showed that the same region is ubiquitinated and that defects in VHL prevent addition of the chemical tag and together, these observations established that the VHL assembly directs ubiquitination of HIF-1α when oxygen is present.
Ratcliffe and Kaelin further wondered what allows VHL to bind HIF-1α under high- but not low-oxygen conditions. Independently, they used various tricks to block the ubiquitin pathway, thereby allowing HIF-1α to accumulate even when oxygen abounds. Their observations suggested the existence of an enzyme that modifies HIF-1α such that it can snag VHL. To identify the presumptive HIF-1α modification, both teams homed in on the region of HIF-1α that grabs VHL. Elimination of a single amino acid—a proline—in this region abrogates VHL’s ability to attach ubiquitins and thus safeguards HIF-1. Further analysis revealed that this proline had picked up an oxygen atom next to one of its hydrogens. HIF-1α had thus acquired a chemical modification called a hydroxyl group. Together, these and other findings indicated that a prolyl hydroxylase, named for its deeds, adds a hydroxyl to a critical proline in HIF-1α, thereby rendering HIF-1α recognizable by VHL and fostering its consequent ruin. Because prolyl hydroxylases require molecular oxygen to do their jobs, the observations explained why HIF-1α is not degraded under hypoxic conditions and thus, how the enzyme translates oxygen levels into HIF-1α stability. The experiments had not, however, uncovered the specific prolyl hydroxylase that acts on HIF-1α, and the amino acid sequence around the hydroxylated HIF-1α proline did not match target sites for known prolyl hydroxylases. Then Ratcliffe proposed that the enzyme belongs to a larger molecular family and identified three related prolyl hydroxylases that govern the cellular response to oxygen in mammals and subsequently demonstrated that hydroxylation of another proline in HIF-1α also promotes its oxygen-dependent, VHL-mediated demolition.
Today’s prize honours three physician-scientists who illuminated the core molecular events that explain how almost all multicellular animals tune their physiology to cope with varying quantities of the life-sustaining element, thus exposing a unique signalling scheme underlying a cornerstone of life on Earth. Their seminal discoveries have revealed the mechanism for one of life’s most essential adaptive processes and established the basis for our understanding of how oxygen levels affect cellular metabolism and physiological function. Kaelin, Ratcliffe, and Semenza have unveiled the fundamental workings of HIF, which presides over a complex and exquisitely controlled response to oxygen fluctuations. The researchers’ findings touch a tremendous array of biological processes, and the details they have unearthed present possible strategies for revving up or reining in these activities. Such ventures might lead toward new therapeutics for a wide range of disorders and lead to the development of novel treatments for anaemia, strokes, cancer and a host of other conditions.