roles of coactivators in hypoxic induction of the erythropoietin gene辅活化因子的角色在缺氧诱导的红细胞生成素基因.pdfVIP

Roles of Coactivators in Hypoxic Induction of the Erythropoietin Gene 1 2 1 1 Feng Wang , Ruixue Zhang , Xiaomeng Wu , Oliver Hankinson * 1 Department of Pathology and Laboratory Medicine, and Jonsson Comprehensive Cancer Center, University of California Los Angeles, Los Angeles, California, United States of America, 2 Department of Microbiology, Immunology and Molecular Genetics, University of California Los Angeles, Los Angeles, California, United States of America Abstract Background: Hypoxia-inducible expression of the erythropoietin (EPO) gene is mediated principally by hypoxia-inducible factor 2a (HIF-2a) in Hep3B cells under physiologic conditions. How/whether p300/CBP and the members of p160 coactivator family potentiate hypoxic induction of endogenous EPO and other HIF-2a and hypoxia-inducible factor 1a (HIF- 1a) target genes remains unclear. Methodology/Principal Findings: We demonstrate, using chromatin immunoprecipitation (ChIP) analysis, that the histone acetyl transferase (HAT) coactivators p300, SRC-1 and SRC-3 are recruited to the 39 enhancer of the EPO gene upon hypoxic stimulation, and that each associates with the enhancer in a periodic fashion. Hypoxia induced acetylation of the EPO gene 59 promoter at histone 4 and lysine 23 of histone 3. Knocking down SRC-3, but not SRC-1 or SRC-2, using short interfering RNAs (siRNAs), reduced EPO transcriptional activity. Knocking down p300 resulted in dramatic down-regulation of hypoxic stimulation of EPO gene transcription, negated recruitment of RNA polymerase II to the gene’s promoter, and eliminated hypoxia-stimulated acetylation at the promoter and recruitments of SRC-1 and SRC-3 to the enhancer. The inhibitory effects of knocking down p300 and the chromati