Cooperative Mechanisms in IL-4-Induced Gene Expression

IL-4 诱导基因表达的协同机制

• 批准号: 6576428
• 负责人: MICHAEL T BERTON
• 金额: $ 31.77万
• 依托单位: UNIVERSITY OF TEXAS HLTH SCIENCE CENTER
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-12-01 至 2007-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6576428
• 关键词: B lymphocyte; JAK kinase; Retroviridae; biological signal transduction; cell differentiation; cytokine receptors; gene expression; gene rearrangement; genetic transduction; helper T lymphocyte; immunoglobulin genes; interleukin 4; laboratory mouse; mass spectrometry; phosphorylation; protein protein interaction; protein sequence; serine; tissue /cell culture; transcription factor; tyrosine; yeast two hybrid system

DESCRIPTION (provided by applicant): The cytokine IL-4 is the single most important cytokine involved in the development and regulation of humoral immune responses. IL-4 drives the differentiation of naive CD4+ T cells into T helper type 2 (Th2) cells, directs isotype switching in B cells to IgE and IgG4 (IgG1 in mice), and regulates cell growth and death. As such, IL-4 has a pivotal role in regulating inflammatory responses, autoimmune diseases, immune responses to infection, and allergy, including asthma. IL-4 regulates this diverse array of immune functions in part through the activation of specific sets of genes in different cell types. IL-4 induces gene expression by inducing Janus kinase(Jak)-mediated activation of the transcription factor Stat6. Importantly, binding of Stat6 alone to DNA is not sufficient to activate transcription and cooperation of Stat6 with other factors is necessary. Recent evidence suggests that other signaling pathways, including the IRS-2/PI 3-kinase and PKC pathways, may be important for Stat6-dependent gene expression, but how other pathways cooperate with Stat6 is entirely unknown. The long-range goal of this project is to understand how IL-4 signaling regulates gene expression to mediate diverse biological effects. The objective of this application is to elucidate the mechanisms by which the Jak/Stat6 pathway cooperates with other signaling pathways and cellular factors to activate gene expression in response to IL-4. The central hypothesis for the proposed research is that IL-4-responsive gene expression is regulated not only by Jak-mediated tyrosine phosphorylation of Stat6 but also by serine kinase signaling pathways that converge on Stat6 and other required transcription cofactors to promote their cooperative interaction. This hypothesis is supported by our recent demonstration that IL-4 induces serine phosphorylation of Stat6. To test the central hypothesis, this proposal will pursue three specific aims: 1) determine the sites of Star6 serine phosphorylation, the kinetics and stability of serine phosphorylation and the role of IL-4R-mediated signaling events in Stat6 serine phosphorylation; 2) determine the role of IL-4-induced Stat6 serine phosphorylation in the regulation of Stat6-dependent gene expression; and 3) establish the mechanisms by which Stat6 cooperates with other necessary cofactors in the regulation of IL-4-dependent gene expression. Serine phosphorylation sites in Stat6 will be determined by peptide mapping and mass spectrometry. Their role in regulating Stat6 expression and function will be analyzed in Stat6-deficient B cells by retrovirus-mediated transduction. The proposed studies will focus on the regulation of the unrearranged IgG1 immunoglobulin gene in mouse B cells as a model for IL-4-induced gene expression. The proposed studies will significantly advance our current limited understanding of how IL-4 signaling and Stat6 regulate gene expression in lymphocytes as well as in other cell types. A better understanding of how IL-4-mediated signals cooperate to regulate gene expression will also allow development of novel approaches for modulating immune responses and disease processes regulated or influenced by IL-4.

描述（由申请人提供）：细胞因子IL-4是参与体液免疫反应发展和调节的最重要的细胞因子。IL-4驱动原生CD4+ T细胞向辅助T型2 （Th2）细胞的分化，指导B细胞向IgE和IgG4（小鼠为IgG1）的同型转换，并调节细胞生长和死亡。因此，IL-4在调节炎症反应、自身免疫性疾病、感染免疫反应和过敏（包括哮喘）中起着关键作用。在某种程度上，IL-4通过激活不同细胞类型中的特定基因来调节多种免疫功能。IL-4通过诱导Janus kinase（Jak）介导的转录因子Stat6的激活来诱导基因表达。重要的是，Stat6单独与DNA结合并不足以激活转录，需要Stat6与其他因子的合作。最近的证据表明，其他信号通路，包括IRS-2/PI 3-激酶和PKC通路，可能对Stat6依赖的基因表达很重要，但其他途径如何与Stat6合作完全未知。该项目的长期目标是了解IL-4信号如何调节基因表达以介导多种生物效应。本申请的目的是阐明Jak/Stat6通路与其他信号通路和细胞因子合作激活基因表达以响应IL-4的机制。本研究的中心假设是，il -4应答基因的表达不仅受jak介导的Stat6酪氨酸磷酸化的调控，还受丝氨酸激酶信号通路的调控，丝氨酸激酶信号通路聚集在Stat6和其他所需的转录辅助因子上，促进它们的协同相互作用。我们最近的研究表明，IL-4诱导Stat6丝氨酸磷酸化，支持了这一假设。为了验证中心假设，本提案将追求三个具体目标：1)确定Star6丝氨酸磷酸化的位点，丝氨酸磷酸化的动力学和稳定性以及il - 4r介导的信号事件在Stat6丝氨酸磷酸化中的作用；2)确定il -4诱导的Stat6丝氨酸磷酸化在Stat6依赖性基因表达调控中的作用；3)建立Stat6协同其他必要的辅助因子调控il -4依赖性基因表达的机制。Stat6中的丝氨酸磷酸化位点将通过肽图谱和质谱测定。它们在Stat6缺失B细胞中调控Stat6表达和功能的作用将通过逆转录病毒介导的转导进行分析。拟开展的研究将重点关注小鼠B细胞中未重排IgG1免疫球蛋白基因的调控，作为il -4诱导基因表达的模型。这些研究将极大地促进我们目前对IL-4信号和Stat6如何调节淋巴细胞以及其他细胞类型中的基因表达的有限理解。更好地了解IL-4介导的信号如何协同调节基因表达，也将有助于开发新的方法来调节由IL-4调节或影响的免疫反应和疾病过程。