Bonner J J, Heyward S, Fackenthal D L
Department of Biology, Indiana University, Bloomington 47405.
Mol Cell Biol. 1992 Mar;12(3):1021-30. doi: 10.1128/mcb.12.3.1021-1030.1992.
The heat shock transcription factor (HSF) of the yeast Saccharomyces cerevisiae is posttranslationally modified. At low growth temperatures, it activates transcription of heat shock genes only poorly; after shift to high temperatures, it activates transcription readily. In an effort to elucidate the mechanism of this regulation, we constructed a series of HSF-VP16 fusions that join the HSF DNA-binding domain to the strong transcriptional activation domain from the VP16 gene of herpes simplex virus. Replacement of the endogenous C-terminal transcriptional activation domain with that of VP16 generates an HSF derivative that exhibits behavior reminiscent of HSF itself: low transcriptional activation activity at normal growth temperature and high activity after heat shock. HSF can thus restrain the activity of the heterologous VP16 transcriptional activation domain. To determine what is required for repression of activity at low temperature, we deleted portions of HSF from this HSF-VP16 fusion to map the regulatory domain. We also isolated point mutations that convert the HSF-VP16 fusion into a constitutive transcriptional activator. We conclude that the central, evolutionarily conserved domain of HSF, encompassing the DNA-binding and multimerization domains, contains a major determinant of temperature-dependent regulation.
酿酒酵母的热休克转录因子（HSF）存在翻译后修饰。在低温生长条件下，它对热休克基因转录的激活作用很差；而在转移至高温后，它能轻易激活转录。为了阐明这种调控机制，我们构建了一系列HSF-VP16融合蛋白，将HSF的DNA结合结构域与单纯疱疹病毒VP16基因的强转录激活结构域连接起来。用VP16的C端转录激活结构域替换内源性C端转录激活结构域，产生了一种HSF衍生物，其表现类似于HSF自身：在正常生长温度下转录激活活性低，热休克后活性高。因此，HSF能够抑制异源VP16转录激活结构域的活性。为了确定低温下抑制活性所需的条件，我们从这种HSF-VP16融合蛋白中删除了部分HSF，以定位调控结构域。我们还分离出了能将HSF-VP16融合蛋白转化为组成型转录激活剂的点突变。我们得出结论，HSF的中央保守结构域，包括DNA结合结构域和多聚化结构域，包含温度依赖性调控的主要决定因素。
