A novel mode of action of YC-1 in HIF inhibition: stimulation of FIH-dependent p300 dissociation from HIF-1{alpha}.
Hypoxia-inducible factor (HIF)-1 plays a key role in tumor promotion by inducing approximately 60 genes required for tumor adaptation to hypoxia; thus, it is viewed as a target for cancer therapy. For this reason, YC-1, which down-regulates HIF-1alpha and HIF-2alpha at the post-translational level, is being developed as a novel anticancer drug. We here found that YC-1 acts in a novel manner to inhibit HIF-1. In the Gal4 reporter system, which is not degraded by YC-1, YC-1 was found to significantly inactivate the COOH-terminal transactivation domain (CAD) of HIF-1alpha, whereas it failed to inactivate CAD(N803A) mutant. In coimmunoprecipitation assays, YC-1 stimulated factor inhibiting HIF (FIH) binding to CAD even in hypoxia, whereas it failed to increase the cellular levels of hydroxylated Asn803 of CAD. It was also found that YC-1 prevented p300 recruitment by CAD in mammalian two-hybrid and coimmunoprecipitation assays. The involvement of FIH in YC-1-induced CAD inactivation was confirmed in EPO-enhancer and Gal4 reporter systems using FIH small interfering RNA and dimethyloxalylglycine FIH inhibitor. Indeed, FIH inhibition rescued HIF target gene expressions repressed by YC-1. In cancer cell lines other than Hep3B, YC-1 inhibits HIF-1alpha via the FIH-dependent CAD inactivation as well as via the protein down-regulation. Given these results, we suggest that the functional inactivation of HIF-alpha contributes to the YC-1-induced deregulation of hypoxia-induced genes.
Li
Shan Hua
SH
Department of Pharmacology, Seoul National University College of Medicine, Seoul, Korea.
Shin
Dong Hoon
DH
Chun
Yang-Sook
YS
Lee
Myung Kyu
MK
Kim
Myung-Suk
MS
Park
Jong-Wan
JW
D000600:Amino Acids, Dicarboxylic / Q000494:pharmacology; D045744:Cell Line, Tumor; D020536:Enzyme Activators / Q000494:pharmacology*; D015972:Gene Expression Regulation, Neoplastic; D017930:Genes, Reporter; D006801:Humans; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000037:antagonists & inhibitors* / Q000378:metabolism; D007191:Indazoles / Q000737:chemistry / Q000494:pharmacology*; D006899:Mixed Function Oxygenases; D008954:Models, Biological; D010957:Plasmids / Q000378:metabolism; D014176:Protein Biosynthesis; D034741:RNA, Small Interfering / Q000378:metabolism; D012097:Repressor Proteins / Q000378:metabolism*; D015533:Transcriptional Activation; D050880:p300-CBP Transcription Factors / Q000378:metabolism*
D000600:Amino Acids, Dicarboxylic; D020536:Enzyme Activators; C497442:HIF1A protein, human; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D007191:Indazoles; D034741:RNA, Small Interfering; D012097:Repressor Proteins; C090937:3-(5'-hydroxymethyl-2'-furyl)-1-benzylindazole; D006899:Mixed Function Oxygenases; C439790:HIF1AN protein, human; D050880:p300-CBP Transcription Factors; C040947:oxalylglycine