Assessment of aryl hydrocarbon receptor complex interactions using pBEVY plasmids:: expression vectors with bi-directional promoters for use in Saccharomyces cerevisiae
Assessment of aryl hydrocarbon receptor complex interactions using pBEVY plasmids:: expression vectors with bi-directional promoters for use in Saccharomyces cerevisiae
复制标题
DOI：
10.1093/nar/26.15.3577
复制
发表时间：
1998-08-01
影响因子：
14.9
通讯作者：
Hyman, LE
中科院分区：
文献类型：
--
作者：
Miller, CA;Martinat, MA;Hyman, LE
设计了酵母双向表达载体pBEVY（bi-directional expression vectors for yeast），构建了含有组成型启动子和半乳糖诱导型启动子的酵母双向表达载体pBEVY。利用单个pBEVY质粒表达了由人芳烃受体（Ahr）和芳烃受体核转运蛋白（ARNT）组成的异源二聚体转录因子。以往的研究表明，雌激素受体和Ahr/Arnt复合物之间可能发生抑制性串扰，Hsp 90-Ahr复合物的形成对于Ahr介导的信号转导是重要的。在酵母系统中使用pBEVY质粒研究这些蛋白质之间的功能相互作用的证据。在用共表达Ahr、Arnt和雌激素受体的酵母进行的信号传导测定中没有观察到抑制性串扰。相反，在非允许条件下生长的温度敏感的Hsp 90菌株中，Ahr/Arnt介导的信号转导减少了80%。我们得出结论，pBEVY质粒有利于检查酵母模型系统中的多个蛋白质相互作用。
The pBEVY (bi-directional expression vectors for yeast) plasmids were designed with constitutive and galactose-induced bi-directional promoters to direct the expression of multiple proteins in Saccharomyces cerevisiae, Using human estrogen receptor as a test gene, relatively balanced expression levels from each side of a bi-directional promoter were observed. Expression of a functional heterodimeric transcription factor composed of human aryl hydrocarbon receptor (Ahr) and aryl hydrocarbon receptor nuclear translocator (Amt) proteins was accomplished using a single pBEVY plasmid, Previous studies suggest that inhibitory cross-talk between the estrogen receptor and the Ahr/Arnt complex may occur and that Hsp90-Ahr complex formation is important for Ahr-mediated signal transduction. Evidence for functional interaction among these proteins was investigated using pBEVY plasmids in a yeast system. No inhibitory cross-talk was observed in signaling assays performed with yeast that co-expressed Ahr, Arnt and estrogen receptor, In contrast, Ahr/Arnt-mediated signal transduction was reduced by 80% in a temperature-sensitive Hsp90 strain grown under non-permissive conditions. We conclude that pBEVY plasmids facilitate the examination of multiple protein interactions in yeast model systems.