GENE SEQUENCES INVOLVED IN PROLACTIN ACTION

涉及催乳素作用的基因序列

• 批准号: 6785046
• 负责人: LI-YUAN YU-LEE
• 金额: $ 7.53万
• 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 2004-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6785046
• 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.

该提议的长期目标是了解催乳素 （PRL）调节细胞增殖和分化。学习 沿PRL刺激的有丝分裂途径的多个成分，我们 克隆并表征了PRL受体（PRL-R）， 造血素/细胞因子受体超家族和早期面板 反应基因（转录因子，细胞因子，细胞因子受体） 并延迟了早期反应基因，例如RNUDC。 结构和 这些基因中的两个功能将进行分析：PRL-R， 介导PRL和RNUDC的有丝分裂作用，其合成是一个 PRL刺激的细胞增殖的结果。 目标1分析 与信号蛋白相互作用的PRL-R结构域。 如何 PRL-R细胞内结构域酪氨酸残基充当对接位点 用于招募转录因子STAT1和Stat5将是 通过诱变，转染，凝胶筛和体外相互作用检查 测定。 PRL-R与酶2'，5-相互作用的重要性 少聚二苯基合成酶（OAS），被识别为PRL-R-R 通过酵母双杂交遗传学的相互作用蛋白质将得到解决。 PRL-R截断将用于确定PRL-R/OAS的位置 相互作用，OAS感官和反义构造将用于 解决PRL-R信号转导中潜在的OAS函数。 目标号码 2解决了RNUDC的结构和功能。 遗传互补 在曲霉中的研究表明，大鼠rnudc是一个 真菌中核运动蛋白的功能同源物； 但是，其在哺乳动物细胞中的功能尚不清楚。 我们的本地化 rNUDC染色到中心体/高尔基州地区表明令人兴奋的 RNUDC可能在中心体/高尔基体结构中发挥作用的可能性 和/或功能。 一系列生化，药理学，遗传和 概述了免疫荧光显微镜研究，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理性和反义策略。 最后， 50 kDa rNUDC相互作用蛋白的身份将由 微链试验。 这些合并的研究应提供新的见解 进入信号传导机制，信号分子和基因序列 参与细胞增殖的PRL调节。 在更广泛的规模上 这些研究可能阐明细胞生长控制中的重要步骤 并提供有关神经内分泌 - 免疫系统相互作用的见解。