Growth Hormone Regulation of SH2-B

说明(申请人提供)：将JAK家族酪氨酸激酶鉴定为生长激素受体(GHR)和细胞因子受体超家族的许多其他成员的受体相关信号分子，是我们理解GH作用的细胞基础的重要一步。在GH结合时，GHR相关的JAK2被激活，并在JAK2本身和GHR内磷酸化酪氨酸。这些磷酸酪氨酸形成了多种信号分子的结合位点，JAK2-GHR复合体的募集和/或激活启动了导致对GH的不同生理反应的信号通路。虽然一些信号分子与GHR中的磷酸酪氨酸结合，但对JAK2中与磷酸酪氨酸结合的信号分子知之甚少，尽管JAK2中存在49个酪氨酸。Sh2-B是为数不多的已知JAK2结合蛋白之一。作为对GH的响应，SH2-B与JAK2结合，被JAK2磷酸化，并增强GH诱导的JAK2的激活。SH2-B独立于其激活JAK2的能力，增强了对肌动蛋白细胞骨架和细胞运动的生长激素调节。最后，SH2-B穿梭于细胞质和细胞核之间，促进叉头转录因子的核输出。这一建议的目的是验证SH2-B是一种多功能适配器/支架蛋白的假设，它增强JAK2的GH激活，增强对肌动蛋白细胞骨架的GH调节，并调节细胞核内的GH反应。目的1研究SH2-B激活JAK2的机制。目的2将进一步确定SH2-B在生长激素介导的肌动蛋白细胞骨架变化中的作用。与肌动蛋白细胞骨架调控有关的多种蛋白质已被初步鉴定为SH2-Bβ结合蛋白。它们与SH2-B的结合将得到证实，并确定SH2-B对其功能和亚细胞位置的影响。目标3将研究SH2-B是否通过将蛋白质送入或送出细胞核来调节转录。最后，Aim 4将使用各种蛋白质耗竭技术来确定SH2-B在生长激素作用中的生理作用，包括siRNA、SH2-B“小鼠和来自SH2-B-/-小鼠的小鼠胚胎成纤维细胞和组织。这些研究将深入了解SH2-B在生长激素和其他细胞因子、激素和生长因子(如干扰素-伽马、胰岛素、神经生长因子)功能中的作用，这些因素也利用SH2-B作为信号蛋白。这种洞察力与理解GH调节身体生长和代谢的机制(S)有关，其他SH2-B激活配体有助于、预防和/或缓解各种疾病的症状，包括各种癌症、糖尿病、多发性硬化症和免疫系统疾病。

DESCRIPTION (provided by applicant): Identification of JAK family tyrosine kinases as receptor-associated signaling molecules for growth hormone (GH) receptors (GHR) and the many other members of the cytokine receptor superfamily was a major step forward in our understanding of the cellular basis for GH actions. Upon GH binding, GHR-associated JAK2 is activated and phosphorylates tyrosines within JAK2 itself and GHR. These phosphotyrosines form binding sites for a variety of signaling molecules whose recruitment to and/or activation by JAK2-GHR complexes initiates the signaling pathways that lead to the diverse physiological responses to GH. While some signaling molecules bind to phosphotyrosines within GHR, little is known about the signaling molecules that bind to phosphotyrosines within JAK2, despite the presence of 49 tyrosines within JAK2. SH2-B is one of the few known JAK2 binding proteins. In response to GH, SH2-B binds to JAK2, is phosphorylated by JAK2 and enhances GH-induced activation of JAK2. Independent of its ability to activate JAK2, SH2-B enhances GH regulation of the actin cytoskeleton and cell motility. Finally, SH2-B shuttles between the cytoplasm and the nucleus, and enhances the nuclear export of forkhead transcription factors. The aim of this proposal is to test the hypothesis that SH2-B is a multi-functional adapter/scaffolding protein that enhances GH activation of JAK2, enhances GH regulation of the actin cytoskeleton and regulates GH responses in the nucleus. Aim 1 lexamines the mechanism by which SH2-B activates JAK2. Aim 2 will further define the role of SH2-B in GH-mediated changes in the actin cytoskeleton. Multiple proteins implicated in the regulation of the actin cytoskeleton have been tentatively identified as SH2-Bbeta binding proteins. Their binding to SH2-B will be confirmed and the effect of SH2-B on their function and subcellular location determined. Aim 3 will examine whether SH2-B regulates transcription by shuttling proteins into or out of the nucleus. Finally, Aim 4 will use a variety of protein depletion techniques to identify the physiological role for SH2-B in GH action, including siRNA, SH2-B" mice, and murine embryo fibroblasts and tissues from SH2-B -/- mice. These studies will provide insight into the role of SH2-B in the function of GH and other cytokines, hormones and growth factors (e.g. interferon-gamma, insulin, nerve growth factor) that also utilize SH2-B as a signaling protein. Such insight is relevant to understanding the mechanism(s) by which GH regulates body growth and metabolism, and other SH2-B activating ligands contribute to, prevent and/or alleviate symptoms of a variety of diseases including various cancers, diabetes, multiple sclerosis, and diseases of the immune system.