Cancer cells should accommodate to oxygen- and energy-deficient microenvironment to keep growing. The transcription factors HIF-1a and HIF-2a sense the oxygen state, and express hypoxia-induced genes. The deacetylase SIRT1 senses the energy or redox state, and modulates activities of transcription factors that express metabolic genes. Previously, two studies demonstrated that SIRT1 deacetylates HIF-1a at K674 or HIF-2a at K741 and that SIRT1 inactivates HIF-1a but activates HIF-2a. Yet, how SIRT1 differentially regulates HIF- as is uncertain so far. We here tried to clarify the roles of SIRT1 in HIF-1a and HIF-2a regulations. In the present study, we established two assay systems to separately evaluate SIRT1’s actions on two HIF-a s; that is, immnoblotting with specific antibodies to acetylated HIF-1a_K674 or acetylated HIF-2a_K741 and reporter assay using Gal4(DBD)-HIF-1a(TAD) or Gal4(DBD)-HIF-2a(TAD), both of which contain the SIRT1 target sites. Consequently, PCAF acetylated the two lysine residues of HIF- as, which was reversed by SIRT1. Also, we confirmed the opposing actions of SIRT1 on HIF-1 and HIF-2 driven transcriptions, namely, it negatively and positively regulated HIF-1 and HIF-2, respectively. RT-PCR analyses showed that SIRT1 reduced mRNA levels of HIF-1 target genes but enhanced those of HIF-2 target genes. These results further support the differential roles of SIRT1 in HIF-1 and HIF-2 signaling under hypoxia. As one of two HIF- as tends to be chosen as a major adaptive factor during tumor evolution, the selective modulation of HIF-1 and HIF-2 signaling can be a promising strategy for cancer therapy. In this aspect, SIRT1 could be a candidate target to develop tailor-made cancer therapy for patients. Key words: SIRT1, HIF-1a, HIF-2a, hypoxia, cancer