Abstract 3941: HAF promotes the switch from HIF-1α to HIF-2α signaling during chronic hypoxia and promotes stem cell characteristics

Abstract

Abstract Most solid tumors and their metastases experience periods of hypoxia, which promotes tumor progression and resistance to therapy. Hypoxia is also an important micro-environmental factor supporting the maintenance of pluripotent cancer stem/progenitor cells (CSCs) within the stem cell niche. The clinical relevance of CSCs is underscored by their resistance to cytotoxic therapies, invasive potential and role in tumor recurrence. Critical mediators of the hypoxia response are the hypoxia-inducible factors HIF-1α and HIF-2α. Although they have some overlapping functions, the HIFs are generally non-redundant whereby HIF-1α regulates the acute metabolic changes required for hypoxic survival, whereas HIF-2α promotes the adaptation to chronic hypoxia, including the maintenance of CSCs. However, the mechanism determining selectivity for HIF-1α versus HIF-2α activation is unclear. Here we describe a new mechanism for the switch between HIF-1α and HIF-2α dependent transcription during chronic hypoxia by the hypoxia associated factor (HAF). HAF is overexpressed in a number of tumor types and is an E3 ubiquitin ligase that ubiquitinates HIF-1α independently of oxygen and pVHL, thus targeting HIF-1α for proteasomal degradation. HAF also binds to HIF-2α but at a different site than it binds to HIF-1α, increasing HIF-2α transactivation without causing its degradation. Hence HAF switches cellular dependence from HIF-1α towards HIF-2α, resulting in the activation of HIF-2α specific target genes (MMP9, OCT3/4)and the simultaneous inhibition of HIF-1α specific target genes (CA-IX, DDIT4). HAF itself is hypoxia regulated - decreasing during acute hypoxia, but increasing with hypoxic duration. Hence HAF is a key determinant of HIF-2 versus HIF-1 activation in response to hypoxia. Using cell-free recombinant proteins and in vitro binding assays, we show that HAF binding to HIF-2α requires a novel eukaryotic post-translational modification of HIF-2α within a unique region of HIF-2 that is absent from HIF-1. HAF binding to HIF-2α is increased in cells exposed to prolonged hypoxia and promotes the transcription of HIF-2α target genes important in CSC maintenance such as OCT3/4 and SOX2. HAF levels are elevated in CD133+ U87 glioblastoma stem/progenitor cells and HAF knockdown decreases neurosphere formation suggesting a key role for HAF in the maintenance of the stem cell phenotype. The HAF-induced switch to HIF-2α dependent transcription promotes invasion and enriches the proportion of stem/progenitor CD133+ U87 cells. This increases the rate of tumor take and increases tumor morbidity in vivo. Funded by NIH CA095060, CA17094, CA098920 Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3941. doi:1538-7445.AM2012-3941