FUNCTIONAL INTERFERENCE BETWEEN HYPOXIA AND DIOXIN SIGNAL-TRANSDUCTION PATHWAYS - COMPETITION FOR RECRUITMENT OF THE ARNT TRANSCRIPTION FACTOR

Hypoxia-inducible factor 1 alpha (HIF-1 alpha) and the intracellular d ioxin receptor mediate hypoxia and dioxin signalling, respectively. Bo th proteins are conditionally regulated basic helix-loop-helix (bHLH) transcription factors that, in addition to the bHLH motif, share a Per -Amt-Sim (PAS) region of homology and form heterodimeric complexes wit h the common bHLH/PAS partner factor Arnt. Here we demonstrate that HI F-1 alpha required Arnt for DNA binding in vitro and functional activi ty in vivo, Both the bHLH and PAS motifs of Amt were critical for dime rization with HIF-1 alpha. Strikingly, HIF-1 alpha exhibited very high affinity for Arnt in coimmunoprecipitation assays in vitro, resulting in competition with the ligand-activated dioxin receptor for recruitm ent of Amt. Consistent with these observations, activation of HIF-1 al pha function in vivo or overexpression of HIF-1 alpha inhibited ligand -dependent induction of DNA binding activity by the dioxin receptor an d dioxin receptor function on minimal reporter gene constructs, Howeve r, HIF-1 alpha- and dioxin receptor-mediated signalling pathways were not mutually exclusive, since activation of dioxin receptor function d id not impair HIF-1 alpha-dependent induction of target gene expressio n, Both HIF-1 alpha and Amt mRNAs were expressed constitutively in a l arge number of human tissues and cell lines, and these steady-state ex pression levels were not affected by exposure to hypoxia, Thus, HIF-1 alpha may be conditionally regulated by a mechanism that is distinct f rom induced expression levels, the prevalent model of activation of HI F-1 alpha function, Interestingly, we observed that HIF-1 alpha was as sociated with the molecular chaperone hsp90, Given the critical role o f hsp90 for ligand binding activity and activation of the dioxin recep tor, it is therefore possible that HIF-1 alpha is regulated by a simil ar mechanism, possibly by binding an as yet unknown class of ligands.