Braliou Georgia G, Venieris Emmanouil, Kalousi Alkmini, Simos George
Laboratory of Biochemistry, School of Medicine, University of Thessaly, 41222 Larissa, Greece.
Biochem Biophys Res Commun. 2006 Aug 11;346(4):1289-96. doi: 10.1016/j.bbrc.2006.06.043. Epub 2006 Jun 16.
Hypoxia inducible factor 1 (HIF-1), the master regulator of hypoxia-activated genes, is involved in many diseases and is a valid drug target. In order to develop a simple and genetically tractable in vivo system for HIF-1 analysis, we tested the inducible expression of both human HIF-1 subunits (HIF-1alpha and ARNT) in the yeast Saccharomyces cerevisiae and showed the formation of transcriptionally active HIF-1. The use of this system for the identification and characterization of HIF-1 effectors was first validated by showing that two chemical Hsp90 inhibitors, geldanamycin and radicicol, impaired the activity of HIF-1 in yeast. By applying this system in mutant yeast strains, we then identified Hsp90 co-chaperones, which were required for HIF-1 activity. Furthermore, using yeast strains co-expressing truncated forms of HIF-1alpha with ARNT or both HIF-1alpha and ARNT, we characterized fragments of HIF-1alpha that acted as dominant negative mutants and suppressed HIF-1 activity.
缺氧诱导因子1（HIF-1）是缺氧激活基因的主要调节因子，参与多种疾病，是一个有效的药物靶点。为了开发一种用于HIF-1分析的简单且易于进行基因操作的体内系统，我们在酿酒酵母中测试了人类HIF-1两个亚基（HIF-1α和ARNT）的诱导表达，并证明了转录活性HIF-1的形成。通过证明两种化学Hsp90抑制剂格尔德霉素和放线菌酮损害酵母中HIF-1的活性，首次验证了该系统用于鉴定和表征HIF-1效应物的用途。通过将该系统应用于突变酵母菌株，我们随后鉴定了HIF-1活性所需的Hsp90共伴侣蛋白。此外，使用共表达HIF-1α截短形式与ARNT或同时表达HIF-1α和ARNT的酵母菌株，我们表征了作为显性负突变体并抑制HIF-1活性的HIF-1α片段。
