Hypoxia-Associated Factor, a Novel E3-Ubiquitin Ligase, Binds and Ubiquitinates Hypoxia-Inducible Factor 1α, Leading to Its Oxygen-Independent Degradation

The hypoxia-inducible factor 1 alpha (HIF-1 alpha) is the master regulator of the cellular response to hypoxia. A key regulator of HIF-1 alpha is von Hippel-Lindau protein (pVHL), which mediates the oxygen-dependent, proteasomal degradation of HIF-1 alpha in normoxia. Here, we describe a new regulator of HIF-1 alpha, the hypoxia-associated factor (HAF), a novel E3-ubiquitin ligase that binds HIF-1 alpha leading to its proteasome-dependent degradation irrespective of cellular oxygen tension. HAF, a protein expressed in proliferating cells, binds and ubiquitinates HIF-1 alpha in vitro, and both binding and E3 ligase activity are mediated by HAF amino acids 654 to 800. Furthermore, HAF overexpression decreases HIF-1 alpha levels in normoxia and hypoxia in both pVHL-competent and -deficient cells, whereas HAF knockdown increases HIF-1 alpha levels in normoxia, hypoxia, and under epidermal growth factor stimulation. In contrast, HIF-2 alpha is not regulated by HAF. In vivo, tumor xenografts from cells overexpressing HAF show decreased levels of HIF-1 alpha accompanied by decreased tumor growth and angiogenesis. Therefore, HAF is the key mediator of a new HIF-1 alpha-specific degradation pathway that degrades HIF-1 alpha through a new, oxygen-independent mechanism.