Insulin-like Growth Factor I Receptor Signaling

胰岛素样生长因子 I 受体信号转导

• 批准号: 6756196
• 负责人: S P NISSLEY
• 金额: --
• 依托单位: DIVISION OF CLINICAL SCIENCES - NCI
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/6756196
• 关键词: athymic mouse; biological signal transduction; cell line; cell transformation; gene expression; gene targeting; genetic library; genetically modified animals; growth factor receptors; human genetic material tag; insulinlike growth factor; phosphorylation; protein structure function; receptor binding; tissue /cell culture; transfection; yeast two hybrid system

In addition to being important for normal fetal and postnatal growth, there is increasing evidence that insulin-like growth factors I and II (IGF-I, IGF-II) also support the growth of certain cancers. Biologic responses to IGF-I and IGF-II are signaled by the IGF-I receptor. Therefore, we are focusing our research effort on understanding signaling by the IGF-I receptor. We have used the yeast two-hybrid system to identify new binding partners for the IGF-I receptor. We have inserted the cytoplasmic domain of the IGF-I receptor into the LexA DNA binding vector. Poly A mRNA was prepared from the human osteosarcoma cell line, MG-63, and given to a commercial laboratory for preparation of a cDNA library in the yeast two-hybrid activation domain vector. The library screen identified IGF-I receptor binding partners that we and others had previously identified (p85 regulatory subunit of PI3-K, p66 Shc, Grb10, 14-3-3 beta and zeta, SOCS-1, SH2-B/PSM, RACK-1) as well as new interactors (STAT5a, GIPC, Ril, and 4 clones that are not fully characterized). We are focusing on GIPC which represented 25% of the most strongly interacting clones. GIPC did not interact with the insulin receptor in the yeast two hybrid assay. GIPC is a 333 amino acid protein with a central SH2 domain. Mutational analysis in the yeast two-hybrid system showed that GIPC PDZ domain binds to the carboxy tail of the IGF-I receptor. GIPC dimerizes and the dimerization domain has been shown to include the amino terminal domain as well as a portion of the PDZ domain. IGF-I receptor constructs that have been mutated to prevent GIPC binding have been transfected into IGF-I receptor null mouse embryo fibroblasts in order to explore the function of GIPC in IGF-I receptor signaling. We have developed a mouse monoclonal antibody (4G11) directed against the human IGF-I receptor. This monoclonal antibody blocks the binding of radiolabeled IGF-I to MCF-7 breast cancer cells and MG-63 osteosarcoma cells and downregulates the receptor.

除了对正常胎儿和出生后的生长很重要外，越来越多的证据表明胰岛素样生长因子I和II（IGF-I，IGF-II）也支持某些癌症的生长。对IGF-I和IGF-II的生物学应答由IGF-I受体发出信号。因此，我们的研究重点是了解IGF-I受体的信号传导。 我们已经使用酵母双杂交系统，以确定新的结合伙伴的IGF-I受体。我们已经将IGF-1受体的胞质结构域插入到莱克萨DNA结合载体中。从人骨肉瘤细胞系MG-63中制备Poly A mRNA，并将其提供给商业实验室用于制备酵母双杂交激活结构域载体中的cDNA文库。文库筛选鉴定了我们和其他人先前鉴定的IGF-I受体结合配偶体（PI 3-K的p85调节亚基、p66 Shc、Grb 10、14-3-3 β和zeta、SOCS-1、SH 2-B/PSM、RACK-1）以及新的相互作用物（STAT 5 a、GIPC、Ril和4个未完全表征的克隆）。我们专注于GIPC，它代表了25%的最强相互作用的克隆。在酵母双杂交试验中，GIPC不与胰岛素受体相互作用。GIPC是一种由333个氨基酸组成的蛋白质，具有一个中心SH 2结构域。酵母双杂交系统中的突变分析表明，GIPC PDZ结构域与IGF-I受体的羧基尾结合。GIPC二聚化，并且二聚化结构域已显示包括氨基末端结构域以及PDZ结构域的一部分。已被突变以阻止GIPC结合的IGF-I受体构建体已被转染到IGF-I受体缺失的小鼠胚胎成纤维细胞中，以探索GIPC在IGF-I受体信号传导中的功能。 我们已经开发了一种针对人IGF-I受体的小鼠单克隆抗体（4G 11）。该单克隆抗体阻断放射性标记的IGF-I与MCF-7乳腺癌细胞和MG-63骨肉瘤细胞的结合，并下调受体。