REGULATION AND FUNCTION OF PLC GAMMA 1 IN SMALL INTESTIN

细胞生长和分化的控制是由一个信号调节的 由生长因子结合跨膜蛋白启动的转导级联反应 含有酪氨酸激酶活性的受体。 几乎没有 机械信息可用于解释不同的促有丝分裂和 表皮生长因子给药的非促有丝分裂反应 肠子 受体激酶调控的研究进展 和底物的相互作用是src同源结构域命名为SH 2 和SH 3是调节序列，决定蛋白质的特异性- 信号转导过程中的蛋白质相互作用。 磷脂酰肌醇特异性磷脂酶C γ-1（PLC γ 1）是一种 EGFR酪氨酸激酶的最佳表征底物。 我们检测到 蛋白质表达、酪氨酸磷酸化和PLC γ 1的活性， 断奶时大鼠小肠收缩最大。 本建议的主要目的是描述 表达、生长因子调控和PLC γ 1等的功能 SH 2/SH 3结构域信号转导蛋白。 以一种独特的，支持性的 环境，并获得所有必要的试剂和设备， 研究，PI将在申办者的审查和指导下寻求 研究PLC γ 1活性的重要调节机制， 肠，PLC γ 1对分化和功能的影响， 肠细胞系和其他SH 2/SH 3结构域蛋白的酪氨酸 肠内激酶信号通路。 这个项目的模型 将是哺乳和断奶大鼠小肠，进行体外试验 在IEC-6和CaCo-2细胞系中。 杆状病毒表达的组合 系统、免疫沉淀、交联、活性测定和Western 并且北方印迹将用于研究潜在的结合蛋白， 酪氨酸磷酸化和去磷酸化事件和表达， PLC γ 1和其他SH 2/SH 3结构域蛋白的生长因子调节。 将开发一个模型，用于研究PLC γ 1在 肠，使用经尿道诱导的表达载体，所述表达载体含有 将开发反义PLC γ 1 mRNA用于IEC-6中的转染 细胞系 该模型的成功开发将为 为研究其他信号转导蛋白的作用机制奠定了基础。 未来的研究。

Control of cell growth and differentiation is regulated by a signal transduction cascade initiated by growth factor binding to transmembrane receptors containing tyrosine kinase activity. There is little mechanistic information available to explain the diverse mitogenic and non-mitogenic responses to epidermal growth factor administration in the intestine. The current understanding of the regulation of receptor kinase and substrate interaction is that src homologous domains designated SH2 and SH3 are regulatory sequences, determining the specificity of protein- protein interaction in the signal transduction process. Phosphotidylinositol specific phospholipase C gamma-1 (PLCgamma1) is the best characterized substrate of EGFR tyrosine kinase. We have detected protein expression, tyrosine phosphorylation and activity of PLCgamma1 in the rat small intestine to be greatest during weaning. The principal objective of this proposal is to characterize the expression, growth factor regulation and function of PLCgamma1 and other SH2/SH3 domain signal transduction proteins. In a unique, supportive environment, with access to all reagents and equipment necessary for these studies, the PI under the review and guidance of the sponsor will seek to study the significant regulatory mechanisms of PLCgamma1 activity in the intestine, the effect of PLCgamma1 on differentiation and function in an intestinal cell line and other SH2/SH3 domain proteins in the tyrosine kinase signaling pathway in the intestine. The model for this project will be the suckling and weanling rat small intestine with in vitro assays in IEC-6 and CaCo-2 cell lines. A combination of a baculovirus expression system, immunoprecipitation, cross-linking, activity assays, and Western and Northern blotting will be used to study potential binding proteins, tyrosine phosphorylation and dephosphorylation events and expression in growth factor regulation of PLCgamma1 and other SH2/SH3 domain proteins. A model will be developed for the study of PLCgamma1 function in the intestine using a hormonally induced expression vector containing antisense PLCgamma1 mRNA will be developed for transfection in the IEC-6 cell line. The successful development of this model will provide the mechanistic basis for the study of other signal transduction proteins in future studies.