Growth Hormone Regulation of SH2B1

描述（由申请人提供）：最近令人兴奋的遗传研究表明支架蛋白SH 2B 1与人类疾病有关。SH 2B 1的人类突变与严重的早发性肥胖相关，伴有不成比例的严重胰岛素抵抗，大脑发育受损，在某些情况下，身材矮小。这种表型（与SH 2B 1-/-小鼠的表型惊人相似）与SH 2B 1是多种配体（包括生长激素（GH）、瘦素和胰岛素）的信号分子一致。SH 2B 1被募集到JAK 2中，是GH调节肌动蛋白细胞骨架的关键组分，是细胞增殖、分化和迁移的先决条件。然而，SH 2B 1促进GH诱导的肌动蛋白细胞骨架变化的机制在很大程度上仍然未知。长期目标是了解SH 2B 1促进GH和其他配体功能的机制，并深入了解GH未被识别的细胞作用; SH 2B 1的新功能;以及支架蛋白如SH 2B 1如何将高度调节和动态复合物从膜结合受体移动到细胞中的多个，通常是远程位点（例如，局灶性粘连、板状伪足、细胞质、细胞核）。本提案的目的是了解SH 2B 1如何调节“信号复合物”的组装和运动，以及这些功能的失调是否会导致细胞功能缺陷，最终导致人类肥胖、胰岛素抵抗、神经系统疾病和生长抑制。中心假设是SH 2B 1促进SH 2B 1“信号复合物”的形成和/或移动到细胞中的特定位置（例如，pm，板状伪足，细胞质，细胞核）。SH 2B 1的这些功能增强了GH对肌动蛋白细胞骨架的调节、GH诱导的运动和细胞核中的GH反应。它们对于正常的人类生理学是至关重要的，使得它们由于突变而受损有助于在人类患者中观察到的显著表型。在强有力的初步数据的指导下，这一假设将通过追求三个特定的目标进行测试：1）阐明SH 2B 1可逆地定位于质膜，细胞质和细胞核的机制。2)确定SH 2B 1在调节GH诱导的肌动蛋白细胞骨架、细胞运动和细胞内室之间蛋白质运输变化中的作用。3)确定与人类疾病相关的SH 2B 1突变的功能后果。实验将使用生物化学，免疫学，细胞生物学和成像技术标准在实验室的组合。还将使用最先进的显微镜、光活化探针、蛋白质组学、细胞系以及来自SH 2B 1-/-小鼠的原代巨噬细胞和MEF。SH 2B 1的核定位信号是介导核输入和质膜结合的双重功能基序，其由附近丝氨酸的磷酸化调节，这一概念是创新的。拟议的研究是重要的，因为它将提供深入了解SH 2B 1在GH和许多其他配体（例如，瘦素、胰岛素、神经生长因子），其利用SH 2B 1作为信号蛋白。 公共卫生相关性：这些研究将深入了解多功能衔接蛋白SH 2B 1在生长激素和许多其他细胞因子，激素和生长因子（例如瘦素，胰岛素，神经生长因子）的功能中的作用，这些因子使用SH 2B 1来介导它们在细胞中的作用。这种见解对于我们理解生长激素如何在体内产生其众所周知的多种效应至关重要，包括身体生长和代谢，以及其他SH 2B 1活化配体有助于预防和/或减轻各种疾病的症状，包括肥胖症，各种癌症，糖尿病，多发性硬化症和免疫系统疾病。

DESCRIPTION (provided by applicant): Exciting recent genetic studies implicate the scaffold protein SH2B1 in human disease. Human mutations in SH2B1 are associated with severe early onset obesity with disproportionately severe insulin resistance, impaired brain development, and, in some cases, short stature. This phenotype (strikingly similar to that of SH2B1-/- mice) is consistent with SH2B1 being a signaling molecule for multiple ligands, including growth hormone (GH), leptin, and insulin. SH2B1 is recruited to JAK2 and is a critical component in GH regulation of the actin cytoskeleton, a prerequisite for cellular proliferation, differentiation and migration. However, the mechanism by which SH2B1 facilitates GH-induced changes in the actin cytoskeleton remains largely unknown. The long-term goal is to understand the mechanism by which SH2B1 contributes to the function of GH and other ligands, and to gain insight into unrecognized cellular actions of GH; new functions of SH2B1; and how scaffold proteins such as SH2B1 move highly regulated and dynamic complexes from membrane-bound receptors to multiple, often remote, sites in the cell (e.g., focal adhesions, lamellipodia, cytoplasm, nucleus). The objective of the current proposal is to understand how SH2B1 regulates assembly and movement of "signaling complexes" and whether dysregulation of these functions contributes to defects in cellular function and ultimately human obesity, insulin-resistance, neurologic disorders, and suppressed growth. The central hypothesis is that SH2B1 facilitates the formation and/or movement of SH2B1 "signaling complexes" to specific locations in the cell (e.g., pm, lamellipodia, cytoplasm, nucleus) in response to GH. These functions of SH2B1 enhance GH regulation of the actin cytoskeleton, GH-induced motility, and GH responses in the nucleus. They are critical to normal human physiology, such that their impairment due to mutation contributes to the dramatic phenotype observed in human patients. Guided by strong preliminary data, this hypothesis will be tested by pursuing three specific aims: 1) Elucidate the mechanism by which SH2B1 reversibly localizes to the plasma membrane, cytosol and nucleus. 2) Determine the role of SH2B1 in regulating GH-induced changes in the actin cytoskeleton, cell motility, and trafficking of proteins between intracellular compartments. 3) Determine the functional consequences of mutations in SH2B1 associated with human disease. Experiments will use a combination of biochemical, immunologic, cell biology, and imaging techniques standard in the lab. State-of-the-art microscopes, photoactivable probes, proteomics, cell lines and both primary macrophages and MEFs from SH2B1-/- mice, will also be used. The concept that the nuclear localization signal of SH2B1 is a dual function motif that mediates both nuclear import and plasma membrane binding that is regulated by phosphorylation of nearby serines is innovative. The proposed research is significant because it will provide insight into the role of SH2B1 in the function of GH and the numerous other ligands (e.g., leptin, insulin, nerve growth factor) that utilize SH2B1 as a signaling protein. PUBLIC HEALTH RELEVANCE: These studies will provide insight into the role of a multifunctional adapter protein SH2B1 in the function of growth hormone and the many other cytokines, hormones and growth factors (e.g. leptin, insulin, nerve growth factor) that use SH2B1 to mediate their effects in cells. Such insight is critical to our understanding on how growth hormone brings about its well known diverse effects in the body, including body growth and metabolism, and other SH2B1 activating ligands contribute to, prevent and/or alleviate symptoms of a variety of diseases including obesity, various cancers, diabetes, multiple sclerosis, and diseases of the immune system.