Role of TFII-I in signal transduction by growth factors

最近，TFII-I已成为导致c-fos启动子的信号转导通路的重要组成部分。虽然TFII-I最初被认为是一种可以与启动子启动子结合的因子，但现在很清楚，它也在信号转导中发挥直接作用。在淋巴细胞中，它被发现与细胞质BTK酪氨酸激酶相关，并被它磷酸化。TFII-I的半缺失与人类的神经发育障碍Williams-Beuren综合征(WBS)密切相关。我们已经证明，TFII-I与c-fos启动子的上游调控元件结合，并在体内与血清反应因子(SRF)和STAT家族蛋白形成复合体。过表达TFII-I可增强c-fos启动子的激活，显性阴性TFII-I则抑制c-fos启动子的激活。我们发现TFII-I的酪氨酸磷酸化受血清生长因子的调节，JAK2激酶在这一调节中起着重要的作用。此外，我们还发现TFII-I还受RhoA和MAP激酶途径的调控，并通过其D结构域直接与ERK和p38结合。这些发现表明，TFII-I是血清/RhoA/SRF途径中的一个关键转录调节因子，用于即刻早期基因表达。在这项研究中，我们将研究TFII-I在核内信号转导中的作用。我们将确定蛋白质中关键的磷酸化位点，并将确定调节丝氨酸、苏氨酸和酪氨酸上TFII-I磷酸化的途径。然后，我们将确定这些磷酸化位点在TFII-I活性中的作用，包括DNA结合、蛋白质结合、反式激活和核转位。此外，我们将产生TFII-I及其相关因子的突变，这些突变将使这些相互作用分离，并使我们能够评估TFII-I与血清反应因子的相互作用，以及MAP激酶在其功能中的作用。我们还将评估TFII-I在RhoA-SRF途径中作为信号中间体的可能作用。这些研究结果将有助于阐明和理解一个可能与人类疾病有关的重要的新信号转导分子。规格。目标1：绘制TFII-I的磷酸化位点图，分析磷酸化在TFII-I功能中的作用。具体目标2：确定特定蛋白质相互作用在TFII-I功能中的作用。具体目标3：确定MAP激酶和RhoA信号通路在TFII-I调控中的作用。具体目标4：确定JAK激酶对TFII-I的调节机制。特异性目标5：确定内源性c-fos基因的诱导是否需要TFII-I。

TFII-I has recently emerged as an important component of the signal transduction pathways that lead to the c-fos promoter. Though TFII-I was initially identified as a factor that can bind to promoter initiator elements, it is now clear that it also plays a direct role in signal transduction. In lymphocytes, it has been found to be associated with the cytoplasmic BTK tyrosine kinase and is phosphorylated by it. Hemizygousity for TFII-I is closely linked to the neurodevelopmental disorder Williams-Beuren syndrome (WBS) in humans. We have shown that TFII-I binds to upstream regulatory elements of the c-fos promoter and forms in vivo complexes with the serum response factor (SRF) and the STAT family proteins. Overexpression of TFII-I enhances c-fos promoter activation and dominant negative TFII-I inhibits c-fos promoter activation. We have found that the tyrosine phosphorylation of TFII-I is regulated by serum growth factors and that the JAK2 kinase is important for this regulation. In addition, we have found that TFII-I is also regulated by the RhoA and MAP kinase pathways and associates directly with ERK and p38 through its D domain. These findings suggest that TFII-I is a key transcriptional mediator in the serum/RhoA/SRF pathway for immediate early gene expression. In this proposal, we will investigate the role of TFII-I in signal transduction to the nucleus. We will identify critical phosphorylation sites in the protein and will determine the pathways which regulate phosphorylation of TFII-I on both serine threonine and tyrosine. We will then determine the role of these phosphorylation sites in TFII-I activity including DNA binding, protein binding, transactivation, and nuclear translocation. Moreover, we will generate mutations in TFII-I and its associated factors that will disassociate these interactions and allow us to assess the role of TFII-I interactions with serum response factor and MAP kinases in its function. We will also evaluate the possible role of TFII-I as a signaling intermediate in the RhoA- SRF pathway. The results of these studies will help to elucidate and understand an important new signal transduction molecule which may be involved in human disease. Spec. Aims Specific Aim number 1: To map the Phosphorylation sites in TFII-I and analyze the role of Phosphorylation in TFII-I function. Specific Aim number 2: To determine the role of specific protein interactions in TFII-I function. Specific Aim number 3: To determine the role of MAP kinase and RhoA signaling pathways in TFII-I regulation. Specific Aim number 4: To determine the mechanism of regulation of TFII-I by of JAK kinases. Specific Aim number 5: To determine whether TFII-I is required for the induction of endogenous c-fos gene.