STRUCTURE-FUNCTION ANALYSIS OF PIT-1/PIT-1BETA ISOFORM

PIT-1/PIT-1BETA异构体的结构功能分析

• 批准号: 6176883
• 负责人: SCOTT E DIAMOND
• 金额: $ 8.56万
• 依托单位: UNIVERSITY OF KENTUCKY
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-09-15 至 2002-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6176883
• 关键词: circular dichroism; gene expression; nuclear magnetic resonance spectroscopy; pituitary neoplasms; prolactin; protein isoforms; protein structure function; transcription factor

Prolatin-secreting lactotroph adenomas (prolactinomas), the most common human pituitary tumors, cause infertility, osteoporosis, loss of vision and loss of pituitary functions. Prolactin gene expression, tightly linked to lactotroph-specific function, is governed by Pit-1, a pituitary-specific POU- homeodomain transcription factor, that transduces cAMP/PKA and Ras signaling pathways. The Pit-1beta isoform contains the 26 amino-acid beta- domain, whose sequence specifies its role as a cell-type specific molecular switch. Pit-1beta blocks basal and Ras-stimulated prolactin promoter activity in GH4 pituitary cells, yet enhances basal and PKA-simulated prolactin promoter activity in HeLa non-pituitary cells. The goal of this proposal is to determine the precise molecular mechanism(s) by which the Pit-1 beta-domain dictates isoform-specific transcriptional responses. To this end, I plan to use molecular, biochemical and structural approaches to dissect the mechanistic structure-function relationships of the Pit-1 beta-domain, in the context of the distinct biological responses we have defined for Pit-1 versus Pit-1beta. I hypothesize that Pit-1 isoform-specific responses are dictated by specific amino acids brought to the Pit-1beta TAB by the beta domain insertion, and that the primary-structural changed created by the beta-domain induce three- dimensional structural changes in Pit-1/Pit-1beta, which may be responsible for the functional consequences of the beta-domain insertion. I came to UCHSC to join Dr. Gutierrez-Hartmann's laboratory in order to apply the molecular approaches to structure-function questions that I learned as a graduate student to the important field of molecular endocrinology. My work has lead to one publication, a review, a submitted manuscript and an internationally presented abstract on this exciting subject. This experience has helped to define an area of research of particular interest to me with potential for future independent development. Dr. Gutierrez-Hartmann's laboratory, where I will be carrying out the proposed work, is actively involved in molecular endocrinology research, and has an outstanding record of productivity. This award will allow me to advance my work on the Pit-1/Pit-1beta system so that I will be able to gain promotion to an independent tenure-tract position and to win the competitive funding necessary for me to successfully pursue that goal.

泌乳素催乳素腺瘤（催乳素瘤）是最常见的人类垂体肿瘤，会导致不孕、骨质疏松、视力丧失和垂体功能丧失。催乳素基因表达与泌乳素特异性功能紧密相关，由 Pit-1 控制，Pit-1 是一种垂体特异性 POU 同源域转录因子，可转导 cAMP/PKA 和 Ras 信号通路。 Pit-1beta 同工型包含 26 个氨基酸的 β 结构域，其序列指定了其作为细胞类型特异性分子开关的作用。 Pit-1beta 阻断 GH4 垂体细胞中基础和 Ras 刺激的催乳素启动子活性，但增强 HeLa 非垂体细胞中基础和 PKA 模拟的催乳素启动子活性。该提案的目标是确定 Pit-1 β 结构域决定异构体特异性转录反应的精确分子机制。为此，我计划在我们为 Pit-1 与 Pit-1beta 定义的不同生物反应的背景下，使用分子、生化和结构方法来剖析 Pit-1 beta 结构域的机械结构-功能关系。我假设 Pit-1 同工型特异性反应是由通过 β 结构域插入带到 Pit-1beta TAB 的特定氨基酸决定的，并且 β 结构域产生的初级结构变化会诱导 Pit-1/Pit-1beta 中的三维结构变化，这可能是造成 β 结构域插入的功能后果的原因。我来到 UHSC 加入 Gutierrez-Hartmann 博士的实验室，是为了将我作为研究生学到的结构功能问题的分子方法应用到分子内分泌学的重要领域。我的工作导致了关于这个令人兴奋的主题的一份出版物、一篇评论、一份提交的手稿和一份在国际上发表的摘要。这段经历帮助我确定了一个我特别感兴趣的研究领域，该领域具有未来独立发展的潜力。我将在 Gutierrez-Hartmann 博士的实验室开展拟议的工作，该实验室积极参与分子内分泌学研究，并拥有出色的生产力记录。该奖项将使我能够推进 Pit-1/Pit-1beta 系统的工作，以便我能够晋升到独立的终身职位，并赢得成功实现这一目标所需的竞争性资金。

项目成果

Repression of the prolactin promoter: a functional consequence of the heterodimerization between Pit-1 and Pit-1 beta.

催乳素启动子的抑制：Pit-1 和 Pit-1 beta 之间异二聚化的功能结果。

• DOI: 10.1677/jme.1.01678
• 发表时间: 2005
• 期刊: Journal of molecular endocrinology.
• 作者: Sporici,RA;Hodskins,JS;Locasto,DM;Meszaros,LB;Ferry,AL;Weidner,AM;Rinehart,CA;Bailey,JC;Mains,IM;Diamond,SE
• 通讯作者: Diamond,SE