MECHANISMS OF INSULIN AND IGFI RECEPTOR FUNCTION

胰岛素和 IGFI 受体功能的机制

• 批准号: 2143511
• 负责人: THOMAS A. GUSTAFSON
• 金额: $ 10.19万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE
• 依托单位国家: 美国
• 财政年份: 1991
• 资助国家: 美国
• 起止时间: 1991-09-01 至 1996-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2143511
• 关键词: HTC cell; biological signal transduction; cell growth regulation; chimeric proteins; clone cells; fibroblasts; growth factor receptors; hormone binding protein; hormone regulation /control mechanism; insulin; insulin receptor; insulinlike growth factor; mitogens; protein tyrosine kinase; receptor binding; transfection

One of the most challenging problems in biology is to understand how cell growth is controlled. The overall objective of the work proposed here is to understand how the peptide hormones insulin and insulin-like growth factor I (IGFI) modulate cellular effects such as cell proliferation by interacting with specific cell surface receptors. The receptors for insulin (IR) and IGFI(IGFIR) are structurally related cell surface receptors whose function is dependent upon hormone-activated intrinsic receptor tyrosine kinase activity. The second messenger pathways utilized by these receptors are undefined and it is equally unclear whether each receptor is coupled to similar or distinct signalling pathways. The molecular mechanisms by which hormone binding to the extracellular receptor subunit activates the cytoplasmic receptor tyrosine kinase and how this activation is coupled to later events are the key unsolved problems in the understanding of insulin and IGFI action. Clinically, problems related to insulin and the insulin receptor are involved in the pathogenesis of type I and II diabetes which affect 5% of the population in the United States. Excesses or deficiencies of IGFI lead to gigantism and small stature and IGFI have been implicated in the progression of some neoplastic tumors. This proposal asks three related questions, (1) How do the molecular structures of the extracellular portions of the IR and IGFIR determine hormone binding specificity and subsequent receptor kinase activation? (2) Are the overlapping biological effects of insulin and IGFI a reflection of intrinsic differences in signalling potential of the receptors linked to distinct functional pathways or due to differences in the distribution of receptors in different cell types? and (3) What specific receptor domains are involved in transducing and regulating the mitogenic signal from the activated kinase to the second messenger systems? To answer the first question, we will use receptor mutants and chimeras to precisely identify extracellular domains of the IR and IGFIR critical for hormone binding and signal transduction. To address the second question, cell lines which express only the IR or IGFIR and are mitogenically dependent upon either insulin or IGFI in serum-free medium will be transfected with the noncognate receptor cDNA to ask whether the mechanisms of insulin- and IGFI-mediated mitogenesis are similar or distinct. Secondly, to examine the potential for direct signalling between the IR and IGFIR, cell lines will be generated by cotransfection with specific combinations of receptor cDNAs and analyzed for inter-receptor hybrid formation and functional "cross-talk" between the receptors. To address the third question, we will focus on the mitogenic effects of the hormones to assess the functions of mutant and chimeric receptors in specific cell backgrounds to localize regions of each receptor involved in the regulation of cell proliferation. Lastly, we will utilize cell lines whose growth in serum-free medium is dependent upon either insulin or IGFI to develop somatic cell genetic assays of receptor function. The studies proposed here should allow insight into fundamental mechanisms of receptor tyrosine kinase action and enhance the general understanding of growth control by receptor tyrosine kinases.

生物学中最具挑战性的问题之一是了解细胞如何 增长是控制的。 这里提出的工作的总体目标是 了解肽激素胰岛素和胰岛素样生长如何 因子I（IGFI）调节细胞效应，例如通过 与特定的细胞表面受体相互作用。胰岛素的受体 （IR）和IGFI（IGFIR）是与结构相关的细胞表面受体 功能取决于激素激活的内在受体酪氨酸 激酶活性。 这些受体利用的第二个信学途径 不确定，同样尚不清楚每个受体是否耦合到 相似或不同的信号通路。 分子机制 激素与细胞外受体亚基的结合激活 细胞质受体酪氨酸激酶以及这种激活如何耦合 以后的事件是理解胰岛素的关键未解决问题 和IGFI动作。 临床上，与胰岛素和胰岛素有关的问题 受体参与了I型和II糖尿病的发病机理 影响美国5％的人口。 过度或缺陷 IGFI导致了巨大的，身材很小，IGFI被暗示 在某些肿瘤肿瘤的进展中。 该提案提出了三个相关问题，（1）分子如何 IR和IgFir的细胞外部分的结构确定 激素结合特异性和随后的受体激酶激活？ （2） 是胰岛素和IGFI的重叠生物学作用反映 与与 不同的功能途径或由于分布的分布差异 不同细胞类型的受体？ （3）哪些特定受体结构域 参与了转导和调节来自 激活的激酶到第二个Messenger系统？回答第一个 问题，我们将使用受体突变体和嵌合体精确识别 IR和IGFIR的细胞外域对于激素结合至关重要 信号转导。要解决第二个问题，细胞线 仅表达IR或Igfir，并在有丝分裂上取决于任何一个 无血清培养基中的胰岛素或IGFI将被转染 非认知受体cDNA询问胰岛素和 IGFI介导的有丝分裂发生相似或不同。 其次，检查 IR和IGFIR之间的直接信号传导的潜力，细胞系 将通过与受体的特定组合共转染产生 cDNA并分析了受体间杂种和功能 受体之间的“串扰”。为了解决第三个问题，我们将 关注激素的有丝分裂作用以评估 特定细胞背景中的突变体和嵌合受体以定位 每个受体的区域涉及细胞增殖的调节。 最后，我们将利用其在无血清培养基中生长的细胞系 取决于胰岛素或IGFI发展体细胞遗传 受体功能的测定。 这里提出的研究应允许 深入了解受体酪氨酸激酶作用和 增强受体酪氨酸对生长控制的一般理解 激酶。