GROWTH HORMONE REGULATION OF SH2B

在理解膜受体信号转导方面最令人兴奋的最新进展之一是鉴定了JAK家族的酪氨酸激酶作为生长激素(GH)受体和细胞因子/造血素受体超家族的许多其他成员的受体相关信号分子。生长激素结合后，生长激素受体相关的JAK2被激活，并使JAK2自身和GH受体内的酪氨酸磷酸化。这些磷酸酪氨酸形成了多种信号分子的结合部位，JAK2-GH受体复合体的募集和/或激活启动了信号通路，最终导致对GH的不同生理反应。一些信号分子与生长激素受体内的磷酸酪氨酸结合。然而，尽管JAK2中存在多个(49)酪氨酸，但对JAK2中与磷酸酪氨酸结合的信号分子知之甚少。利用酵母双杂交系统鉴定了一个新的SH2-B剪接变异体(SH2-Bbeta)作为JAK2结合蛋白。作为对GH的反应，SH2-Bbeta与JAK2结合，并被酪氨酸磷酸化。前期的研究还表明，SH2-Bβ刺激JAK2的自磷酸化，并介导GH诱导的肌动蛋白聚合的变化。这项提议的目的是验证这样的假设，即SH2-Bbeta既作为JAK2的激活剂发挥作用，也作为适配器分子招募到激活的JAK2-GH受体复合体，信号分子调节细胞骨架。具体地说，JAK2与SH2-Bβ结合和酪氨酸磷酸化的机制将被阐明。将确定JAK2-SH2-Bβ复合体形成所需的JAK2和SH2-Bβ区域，以及JAK2响应GH而被JAK2磷酸化的SH2-Bβ内的酪氨酸。SH2-Bbeta增强生长激素刺激的JAK2活性的能力将被检测。将识别与SH2-Bβ富含脯氨酸的N末端一半以及SH2-Bβ中的磷酸化酪氨酸结合的信号分子。最后，位于SH2-Bbeta下游的细胞反应将被识别，最初的重点是识别可能介导GH诱导的细胞骨架变化的途径。这些研究将深入了解SH2-Bbeta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、血小板衍生生长因子)功能中的作用，这些因素也利用SH2-Bbeta作为信号分子。这种洞察力与理解GH调节身体生长和代谢的机制有关，其他SH2-Bbeta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、各种发育异常以及免疫和造血系统疾病。

One of the most exciting recent advances in understanding signal transduction by membrane receptors is the identification of the JAK family of tyrosine kinases as receptor-associated signaling molecules for growth hormone (GH) receptors and the many other members of the cytokine/hematopoietin receptor superfamily. Upon GH binding, GH-receptor-associated JAK2 is activated and phosphorylates tyrosines within JAK2 itself and GH receptor. These phosphotyrosines form binding sites for a variety of signaling molecules whose recruitment to and/or activation by JAK2-GH receptor complexes initiates the signaling pathways that ultimately lead to the diverse physiological response to GH. Some signaling molecules bind to phosphotyrosines within GH receptor. However, little is known about the signaling molecules that bind to phosphotyrosines within JAK2, despite the presence of multiple (49) tyrosines within JAK2. The yeast 2-hybrid system was used to identify a new splicing variant of SH2-B (SH2- Bbeta) as a JAK2 binding protein. In response to GH, SH2-Bbeta binds to JAK2 and is tyrosyl phosphorylated. Prelim studies also suggest that SH2-Bbeta stimulates JAK2 autophosphorylation and mediates GH induced changes in actin polymerization. The aim of this proposal is to test the hypothesis that SH2-Bbeta functions both as an activator of JAK2 and as an adapter molecule that recruits to activated JAK2-GH receptor complexes signaling molecules that regulate the cytoskeleton. Specifically, the mechanism by which JAK2 binds to and tyrosyl phosphorylates SH2- Bbeta will be elucidated. Regions of JAK2 and SH2-Bbeta required for JAK2-SH2-Bbeta complex formation and tyrosines within SH2- Bbeta that are phosphorylated by JAK2 in response to GH will be determined. The ability of SH2-Bbeta to enhance GH-stimulated JAK2 activity will be examined. Signaling molecules that bind to both the proline-rich, N-terminal half of SH2-Bbeta and to the phosphorylated tyrosines in SH2-Bbeta will be identified. Finally, cellular responses that lie downstream of SH2-Bbeta will be identified, with an initial emphasis on identifying pathways that may mediate GH- induced changes in the cytoskeleton. These studies will provide insight into the role of SH2-Bbeta in the function of GH and other cytokines, hormones and growth factors (e.g. interferon-gamma, insulin, platelet-derived growth factor) that also utilize SH2-Bbeta as a signaling molecule. Such insight is relevant to understanding the mechanisms by which GH regulates body growth and metabolism, and other SH2-Bbeta activating ligands contribute to, prevent and/or alleviate symptoms of a variety of diseases, including various cancers, diabetes, multiple sclerosis, various developmental abnormalities, and diseases of the immune and hematopoietic systems.