In a previous study we compared and contrasted the rate and mode of HIF-1alpha molecular evolution between teleost fishes and mammals. Although the overall evolutionary rate in teleost HIF-1alpha was at least twice as fast as in mammalian HIF-1alpha, the crucial interaction domains were observed to be under stringent negative selection in all vertebrates. In teleosts the positions with partial support for positive selection were not found in the vicinity of the oxygen-dependent degradation domain of HIF-1alpha, but in the bHLH-PAS domain responsible for DNA binding and dimerization, suggesting the possibility that these areas may be evolutionarily interesting. To broaden the analysis of hypoxia-inducible factors, we now analyze and compare the rates and modes of evolution of all HIF isoforms: (HIF1-3alpha). High expression of especially HIF-2alpha is observed in some hypoxia-tolerant tumor cells. Thus, it is useful to investigate what kind of selective forces have acted on isoform-specific amino acid residues and test if this analysis can predict potentially important differential protein-protein interactions in the crucial interaction domains.