Regulation and function of mammalian HSF4 in vivo

描述(申请人提供)：随着最近一代缺乏hsf1或hsf2的小鼠的出现，哺乳动物热休克转录因子(Hsf)的功能变得越来越清楚。对这些动物模型的分析结果令人震惊，表明Hsf在细胞凋亡、分化和发育以及维持组织内稳中所起的作用。为了了解Hsf在体内的功能，我们培育了Hsf1和hsf2基因敲除小鼠，现在正在努力创造不表达HSF4的小鼠。HSFL的破坏阐明了其在调节细胞凋亡、控制耐热性和雌性不育性方面的重要性。相比之下，hsf2基因敲除的小鼠在中枢神经系统发育和精子生成减少方面表现出缺陷。我们进一步的证据表明，HSFL和hsf2的缺失会导致协同效应和缺陷，这些效应和缺陷比单个HSF缺陷小鼠表现出的更严重。这些缺陷与大脑和行为异常、精子发生完全中断、男性不育以及其他因素有关。与Hsf1和Hsf2相比，对哺乳动物HSF家族中仅存的成员HSF4的了解相对较少。HSF4有几个区别于其他HSF的特征。它被表达为两种选择性剪接变体，一种是转录激活因子，另一种是转录抑制因子，这两种变体在不同的组织中差异表达。我们发现HSF4是磷酸化的，并且已经鉴定出一种新的磷酸酶(命名为DSPH4)，它包含双特异性酪氨酸/丝氨酸/苏氨酸磷酸酶的特征基序，与HSF4结合并调节其磷酸化状态、DNA结合能力和核定位。这代表了细胞外信号调节HSF4活性的一种可能机制。由于Hsf1和Hsf2具有不同的功能、调控机制和靶基因，但在功能上存在一定的相互依赖性，因此有理由推测Hsf4与Hsf1和Hsf2也有一些重叠的功能，但在维持细胞内环境平衡方面也有其独特的作用。这项拨款申请中概述的研究旨在了解HSF4在分子水平上的调控，并不仅研究HSF4的单独功能，而且还研究HSF4在体内与HSFL和Hsf2功能的协同作用。这一建议的具体目的是：1.了解哺乳动物细胞中HSF4调控的分子机制。2.探讨HSF4在动物模型中的作用。3.了解HSF4剪接变异体在动物模型中的功能。

DESCRIPTION (provided by applicant): The functions of the mammalian heat shock transcription factors (Hsfs) are becoming increasingly clear with the recent generation of mice lacking the hsfl or hsf2. The results generated from analyses of these animal models are striking and indicative of the role of Hsfs in apoptosis, differentiation and development as well as maintenance of tissue homeostasis. To understand the functions of Hsfs in vivo, we have generated hsfl and hsf2 knockout mice and are now working to create mice that do not express hsf4. Disruption of hsfl illuminates its importance for regulation of apoptosis and control of thermotolerance and female sterility. In contrast, the hsf2 knockout mouse exhibits defects in development of the central nervous system and reduction in spermatogenesis. We further show evidence that deletion of both hsfl and hsf2 leads to synergistic effects and defects that are more severe than those manifested in individual hsf-deficient mice. These defects are associated with brain and behavioral abnormalities, and complete disruption of spermatogenesis and male sterility, as well as others. Compared to Hsfl and Hsf2, relatively little is known about the only remaining member of the mammalian Hsf family, Hsf4. Several features distinguish Hsf4 from other Hsfs. It is expressed as two alternatively spliced variants with one form being a transcriptional activator and the other a transcriptional repressor and these two variants are differentially expressed in different tissues. We show that Hsf4 is phosphorylated and a novel phosphatase (named DSPH4) containing signature motifs of the dual specificity tyrosine/serine/threonine phosphatases has been identified that binds to Hsf4 and modulates its phosphorylation status, DNA binding ability and nuclear localization. This represents a possible mechanism for the regulation of Hsf4 activity by extracellular signals. Because Hsfl and Hsf2 possess distinct functions, regulatory mechanisms, and target genes, but show some functional interdependency, it is reasonable to postulate that Hsf4 will also have some overlapping function with Hsfl and Hsf2, but also its own unique roles in maintaining cellular homeostasis. The studies outlined in this grant application are directed to understand the regulation of Hsf4 at the molecular level and to examine the function of Hsf4 not only alone, but also in coordination with Hsfl and Hsf2 functions in vivo. The Specific Aims of this proposal are: 1. To understand the molecular mechanisms of Hsf4 regulation in mammalian cells. 2. To investigate the function of Hsf4 in animal models. 3. To understand the function of Hsf4 splice variants in animal models.