The 2019 Nobel Prize in Physiology or Medicine

has today decided to award

the 2019 Nobel Prize in Physiology or Medicine

jointly to

William G. Kaelin Jr., Sir Peter J. Ratcliffe and Gregg L. Semenza

HIF enters the scene  

 In addition to the carotid body-controlled rapid adaptation to low oxygen levels (hypoxia), there are other fundamental physiological adaptations. A key physiological response to hypoxia is the rise in levels of the hormone erythropoietin (EPO), which leads to increased production of red blood cells (erythropoiesis). The importance of hormonal control of erythropoiesis was already known at the beginning of the 20th century, but how this process was itself controlled by O2 remained a mystery. 

 Gregg Semenza studied the EPO gene and how it is regulated by varying oxygen levels. By using gene-modified mice, specific DNA segments located next to the EPO gene were shown to mediate the response to hypoxia. Sir Peter Ratcliffe also studied O2-dependent regulation of the EPO gene, and both research groups found that the oxygen sensing mechanism was present in virtually all tissues, not only in the kidney cells where EPO is normally produced. These were important findings showing that the mechanism was general and functional in many different cell types.   

 Semenza wished to identify the cellular components mediating this response. In cultured liver cells he discovered a protein complex that binds to the identified DNA segment in an oxygendependent manner. He called this complex the hypoxia-inducible factor (HIF) . Extensive efforts to purify the HIF complex began, and in 1995, Semenza was able to publish some of his key findings, including identification of the genes encoding HIF. HIF was found to consist of two different DNA-binding proteins, so called transcription factors, now named HIF-1a and ARNT. Now the researchers could begin solving the puzzle, allowing them to understand which additional components were involved and how the machinery works. 

VHL: an unexpected partner  

 When oxygen levels are high, cells contain very little HIF-1a. However, when oxygen levels are low, the amount of HIF-1a increases so that it can bind to and thus regulate the EPO gene as well as other genes with HIF-binding DNA segments (Figure 1). Several research groups showed that HIF-1a, which is normally rapidly degraded, is protected from degradation in hypoxia. At normal oxygen levels, a cellular machine called the proteasome, recognized by the 2004 Nobel Prize in Chemistry to Aaron Ciechanover, Avram Hershko and Irwin Rose, degrades HIF-1a. Under such conditions a small peptide, ubiquitin, is added to the HIF-1a protein. Ubiquitin functions as a tag for proteins destined for degradation in the proteasome. How ubiquitin binds to HIF-1a in an oxygen-dependent manner remained a central question. 

 The answer came from an unexpected direction. At about the same time as Semenza and Ratcliffe were exploring the regulation of the EPO gene, cancer researcher William Kaelin, Jr. was researching an inherited syndrome, von Hippel-Lindau's disease (VHL disease). This genetic disease leads to dramatically increased risk of certain cancers in families with inherited VHL mutations. Kaelin showed that the VHL gene encodes a protein that prevents the onset of cancer. Kaelin also showed that cancer cells lacking a functional VHL gene express abnormally high levels of hypoxia-regulated genes; but that when the VHL gene was reintroduced into cancer cells, normal levels were restored. This was an important clue showing that VHL was somehow involved in controlling responses to hypoxia. Additional clues came from several research groups showing that VHL is part of a complex that labels proteins with ubiquitin, marking them for degradation in the proteasome. Ratcliffe and his research group then made a key discovery: demonstrating that VHL can physically interact with HIF-1a and is required for its degradation at normal oxygen levels. This conclusively linked VHL to HIF-1a. 

  Oxygen sHIFts the balance 

 Many pieces had fallen into place, but what was still lacking was an understanding of how O2 levels regulate the interaction between VHL and HIF-1a. The search focused on a specific portion of the HIF-1a protein known to be important for VHL-dependent degradation, and both Kaelin and Ratcliffe suspected that the key to O2-sensing resided somewhere in this protein domain. In 2001, in two simultaneously published articles they showed that under normal oxygen levels, hydroxyl groups are added at two specific positions in HIF-1a (Figure 1). This protein modification, called prolyl hydroxylation, allows VHL to recognize and bind to HIF-1a and thus explained how normal oxygen levels control rapid HIF-1a degradation with the help of oxygen-sensitive enzymes (so-called prolyl hydroxylases). Further research by Ratcliffe and others identified the responsible prolyl hydroxylases. It was also shown that the gene activating function of HIF-1a was regulated by oxygen-dependent hydroxylation. The Nobel Laureates had now elucidated the oxygen sensing mechanism and had shown how it works.