GENE SEQUENCES INVOLVED IN PROLACTIN ACTION

涉及催乳素作用的基因序列

• 批准号: 2440645
• 负责人: LI-YUAN YU-LEE
• 金额: $ 17万
• 依托单位: BAYLOR COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-03-15 至 2002-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2440645
• 关键词: Golgi apparatus; biological signal transduction; cell proliferation; centrosome; cytokine receptors; gel mobility shift assay; hormone receptor; hormone regulation /control mechanism; immunofluorescence technique; nucleic acid sequence; phosphotransferases; prolactin; protein binding; protein localization; protein sequence; protein structure function; regulatory gene; site directed mutagenesis; tissue /cell culture; transcription factor; transfection; tyrosine; yeast two hybrid system

The long term objective of this proposal is to understand how prolactin (PRL) regulates cell proliferation and differentiation. To study multiple components along the PRL-stimulated mitogenic pathway, we cloned and characterized the PRL receptor (PRL-R), a member of the hematopoietin/cytokine receptor superfamily, and a panel of early response genes (transcription factors, cytokines, cytokine receptors) and delayed early response genes, such as RNUDC. The structure and function of two of these genes will be analyzed: the PRL-R which mediates the mitogenic effects of PRL, and RNUDC, whose synthesis is a result of PRL-stimulated cellular proliferation. AIM number 1 analyzes PRL-R domains involved in interacting with signaling proteins. How the PRL-R intracellular domain tyrosine residues serve as docking sites for recruiting the transcription factors Stat1 and Stat5 will be examined by mutagenesis, transfection, gel sift and in vitro interaction assays. The significance of PRL-R interaction with the enzyme 2',5- oligoadenylate synthetase (OAS), which was identified as a PRL-R interacting protein by yeast two-hybrid genetics, will be addressed. PRL-R truncations will be used to determine the site of PRL-R/OAS interaction, and OAS sense and anti-sense constructs will be used to address potential OAS function in PRL-R signal transduction. AIM number 2 addresses RNUDC structure and function. Genetic complementation studies in Aspergillus nidulans have shown that rat RNUDC is a functional homologue of a nuclear movement protein in the fungus; however, its function in mammalian cells is unclear. Our localization of RNUDC staining to the centrosome/Golgi region suggests the exciting possibility that RNUDC may play a role in centrosome/Golgi structure and/or function. A series of biochemical, pharmacological, genetic and immunofluorescence microscopy studies are outlined to determine the cellular localization of RNUDC, and to address potential RNUDC function by sense and anti-sense strategies in mammalian cells. Finally, the identity of a 50 kDa RNUDC interacting protein will be determined by microsequencing. These combined studies should provide new insights into the signaling mechanisms, signaling molecules, and gene sequences involved in PRL regulation of cell proliferation. On a broader scale, these studies may elucidate important steps in cellular growth controls and provide insights into neuroendocrine-immune system interactions.

本提案的长期目标是了解催乳素 (PRL)调节细胞增殖和分化。研究 沿着PRL刺激的促有丝分裂通路，我们 克隆并鉴定了PRL受体（PRL-R）， 促红细胞生成素/细胞因子受体超家族和一组早期 应答基因（转录因子、细胞因子、细胞因子受体） 和延迟早期反应基因，如RNUDC。 结构和 将分析其中两个基因的功能：PRL-R， 介导催乳素和RNUDC的促有丝分裂作用，RNUDC的合成是 PRL刺激细胞增殖的结果。 AIM 1号分析 参与与信号蛋白相互作用的PRL-R结构域。 如何 PRL-R胞内结构域酪氨酸残基作为对接位点 为了募集转录因子Stat 1和Stat 5， 通过诱变、转染、凝胶筛选和体外相互作用检查 分析。 PRL-R与2 '，5- 寡腺苷酸合成酶（OAS），被鉴定为PRL-R 相互作用蛋白的酵母双杂交遗传学，将得到解决。 PRL-R截短将用于确定PRL-R/OAS的位点 相互作用，并且OAS正义和反义构建体将用于 探讨OAS在PRL-R信号转导中的潜在功能。 AIM编号 2.说明了区域城市发展方案的结构和职能。 遗传互补 对构巢曲霉的研究表明，大鼠RNUDC是一种 真菌中核运动蛋白的功能同源物; 然而，其在哺乳动物细胞中的功能尚不清楚。 我们的本地化 RNUDC对中心体/高尔基体区域的染色表明， RNUDC可能在中心体/高尔基体结构中发挥作用 和/或功能。 一系列生化、药理、遗传和 免疫荧光显微镜研究概述，以确定 RNUDC的细胞定位，并解决潜在的RNUDC功能 通过哺乳动物细胞中的正义和反义策略。 最后 50 kDa RNUDC相互作用蛋白的同一性将通过 微测序 这些综合研究应该提供新的见解 信号机制、信号分子和基因序列 参与PRL调节细胞增殖。 在更广泛的范围内， 这些研究可以阐明细胞生长控制的重要步骤 并提供神经内分泌免疫系统相互作用的见解。