Investigation of the role of insulin receptor in chromosome stability.

项目摘要 多细胞生物发展受体介导的信号转导，由细胞外生长因子启动， 增殖。胰岛素是一种生长因子，高胰岛素血症可促进和维持肿瘤的发生 增长胰岛素受体（IR）定位于细胞表面质膜，无论是在代谢组织细胞中还是在代谢组织细胞中。 高度增殖的细胞，如免疫细胞和癌细胞。IR激活两个下游信号通路， PI 3 K-AKT途径和MAPK途径，以调节细胞代谢、增殖和生长。尽管 IR信号在系统稳态中的有趣发现，营养信号如何维持染色体 稳定性仍然不确定。我们知识上的这一差距是我们理解 胰岛素在细胞增殖和分化中的作用及其对人体健康的影响，如高胰岛素血症， 与包括2型糖尿病和癌症在内的各种疾病有关。关键纺锤体检查点蛋白MAD 2 形成有丝分裂检查点复合物（MCC），并确保染色体分离的保真度。我们最近 研究表明MAD 2与IR结合，通过组装MCC样复合物募集网格蛋白接头复合物， 并促进IR内吞作用。在我们未发表的结果中，我们发现， 小鼠增加免疫细胞的非整倍体，并促进T细胞淋巴瘤。这些结果表明 在有丝分裂和IR信号转导中，纺锤体检查点调节因子和IR相互调节。IR和胰岛素- 类生长因子1受体（IGF 1 R）是高度同源的受体酪氨酸激酶（RTK）。IGF 1 R不 与MAD 2结合，其内吞机制和信号转导结果与IR不同。 定义了IR和IGF 1 R的不同激活机制，表明配体特异性诱导的结构 差异可能影响RTKs的内吞作用和下游信号传导。在我们研究 的纺锤体检查点蛋白的IR信号，我们还进行了公正的，系统的全基因组损失 使用CRISPR-Cas9的功能研究。我们发现了新的基因，增加或减少表面的水平， IR和IR信号。在这里，我们建议联合收割机在小鼠遗传学，细胞生物学，生物化学， 基因组学和cryo-EM来确定IR在有丝分裂中的功能，以及IR信号传导的激活机制 细胞增殖和新陈代谢的关系我们在未来五年的目标是探索以下问题：（1） IR如何确保有丝分裂中染色体的准确分离，IR的生理功能是什么 （2）IR如何选择性激活PI 3 K-AKT与MAPK信号分支;以及（3）其他什么 因子是细胞增殖和分化中IR功能所必需的？总体而言，拟议的研究 将促进我们对胰岛素生理作用的功能，调节和机制的理解。 细胞增殖和分化。此外，我们的研究可能会作为进一步翻译的基础。 高胰岛素血症和2型糖尿病的研究和未来的治疗与增加的风险有关 用于某些癌症，也可以用于癌症免疫疗法。

PROJECT SUMMARY Multicellular organisms develop receptor-mediated signal transduction initiated by extracellular growth factors to proliferate. Insulin has long been known as a growth factor, and hyperinsulinemia can promote and sustain tumor growth. Insulin receptor (IR) localizes to the cell surface plasma membrane both in metabolic tissue cells and in highly proliferative cells such as immune cells and cancer cells. IR activates two downstream signaling pathways, the PI3K-AKT pathway and the MAPK pathway, to regulate cell metabolism, proliferation, and growth. Despite intriguing findings for IR signaling in systemic homeostasis, how the nutrient signaling maintains chromosome stability still remains uncertain. This gap in our knowledge presents a key barrier to our understanding of the function of insulin in cell proliferation and differentiation and its impact on human health, as hyperinsulinemia is associated with various diseases including type 2 diabetes and cancer. The key spindle checkpoint protein MAD2 forms a mitotic checkpoint complex (MCC) and ensures the fidelity of chromosome segregation. Our recent studies showed that MAD2 binds to IR, recruits the clathrin adaptor complex by assembling an MCC-like complex, and promotes IR endocytosis. In our unpublished results, we found that disruption of IR-MAD2 interaction in mice increases aneuploidy in the immune cells and promotes T-cell lymphoma. These results suggest that the spindle checkpoint regulators and IR mutually regulate each other in both mitosis and IR signaling. IR and insulin- like growth factor 1 receptor (IGF1R) are highly homologous receptor tyrosine kinases (RTKs). IGF1R does not bind to MAD2, and its endocytosis mechanism and signaling outcomes are different from that of IR. We recently defined the distinct activation mechanisms of IR and IGF1R, suggesting that ligand-specific induced structural differences might affect the endocytosis and downstream signaling of RTKs. In parallel to our studies on the role of spindle checkpoint proteins in IR signaling, we have also performed unbiased, systematic genome-wide loss of function studies using CRISPR-Cas9. We identified novel genes that increase or decrease the levels of surface IR and IR signaling. Here, we propose to combine approaches in mouse genetics, cell biology, biochemistry, genomics and cryo-EM to determine the function of IR in mitosis, as well as activation mechanisms of IR signaling for cell proliferation vs. metabolism. Our goals over the next five years are to explore the following questions: (1) how does IR ensure accurate chromosome segregation in mitosis, and what is the physiological function for IR in cell division; (2) how does IR selectively activate the PI3K-AKT vs. MAPK signaling branch; and (3) what other factors are required for IR function in cell proliferation and differentiation? Collectively, the proposed research will advance our understanding of the function, regulation, and mechanism(s) of insulin action in physiological cell proliferation and differentiation. Furthermore, our studies will likely serve as a basis for further translational research and future therapeutics as hyperinsulinemia and type 2 diabetes are associated with increased risks for certain cancers and may also be harnessed for cancer immunotherapies.