Protein Phosphatase Regulation of the Principal Renal Sodium Transporter NHE3

简介(由申请人提供)：Ion Alexandru Bobulescu博士是一名内科科学家，最近被任命为德克萨斯大学达拉斯西南医学中心(德克萨斯大学西南分校)教师级别的全职教师职位。他的短期职业目标(未来4年)是严格执行K01申请中概述的培训和职业发展计划，使用现有的和高度创新的技术进行拟议的实验，以确定蛋白磷酸酶2A(PP2A)在调节肾脏钠/质子交换中的作用，然后申请R01机制下的资金，以探索这项工作产生的一个或多个科学机会。Bobulescu博士的长期职业目标是成为钠和血压动态平衡肾脏调节领域的领导者和创新者，使用开创性的方法和技术来解决从分子到模型生物再到人类疾病的基本未解问题。为了实现这些目标，提出了一项全面和详细的职业发展计划，以满足以下培训要求：(1)课程安排。其他几个领域的正式培训被认为是推进候选人研究的关键。(2)技术先进。博布列斯库博士将与他的导师和专家合作者一起实践这项提议的目标，他将学习几种新颖的、最先进的方法，并将获得将这些技术应用于自己的研究问题的能力。(3)辅导式职业发展。Bobulescu博士有一位专家导师将直接指导、监督和评估他的研究，这将大大有利于他在新的和未被研究的信号转导领域通过蛋白质磷酸酶获得相关结果的机会。这款K01的导师Marc Mumby博士是世界知名的蛋白质磷酸酶研究专家。此外，Bobulescu博士意识到，一个成功的独立职业生涯需要的不仅仅是想法、技术技能和良好的出版物记录。运营研究实验室，管理和培养人才，促进合作，成为一名有效的导师和教师，以及能够维持一个强大而富有成效的研究项目，从而在第一个R01之后获得成功的资金，这些都是艰巨的努力-但对于真正的学术成功至关重要。候选人确信，在这些领域，他可以从蒙比博士那里学到很多东西，蒙比博士是一位领先的科学家，也是K01的理想导师。(4)部门。博布列斯库博士之前在临床部门接受的博士后研究培训所提供的以疾病为导向的观点，对他成为内科科学家非常重要。然而，他的培训将极大地受益于一段时间的沉浸式暴露在一个领先的基础科学系的截然不同的环境中。环境：德克萨斯大学西南分校是世界上最重要的研究机构之一，拥有4名诺贝尔奖获得者，36名国家科学院和/或其医学研究所成员，3500多个正在进行的研究项目，每年超过4亿美元的资金，以及超过582,000平方英尺的研究空间，还有更多的正在建设中。Bobulescu博士将主要在Marc Mumby博士的药理学系实验室接受培训，实验室总面积超过2,000平方英尺，具备进行生化、分子生物学和细胞生物学实验所需的所有必要设备。Bobulescu博士还将保留他250平方英尺的实验室空间、90平方英尺的私人办公室和内科教职员工的职位。候选人将有权使用德克萨斯大学西南分校最先进的动物研究设施，以及德克萨斯大学西南乔治·M·奥布莱恩肾脏研究核心中心。德克萨斯大学西南分校的传统是留住最好的博士后研究员，并培养他们的独立职业生涯。这一传统包括候选人的前导师奥森·莫博士，他在1990年之前一直是德克萨斯大学西南分校的博士后研究员，此后一直留在那里，晋升为正教授兼中心主任。研究：这项建议将研究PP2A在调节主要的肾脏钠转运体NHE3中的作用，特别强调肾内多巴胺对NHE3的调节。如果提出的假设是正确的，PP2A将被确立为肾内多巴胺活动和NHE3的主要调节因子。这将直接影响我们对血压调节的生理学和病理生理学的理解，因为多巴胺介导的钠尿受损已在高血压人类中被描述，已被归因于两种高血压啮齿动物模型中肾内多巴胺信号的缺陷，并与肾脏PP2A功能缺陷有关。该提案包括三个独立的目标，这些目标将同时使用各种技术来实现。目的1利用细胞培养和体外实验研究多巴胺对NHE3的抑制是否需要PP2A对NHE3的直接作用。目的2将利用细胞培养、生物化学和先进的电生理技术来测试PP2A对NHE3的去磷酸化是否起到决定NHE3命运的细胞内开关的作用。目的3将使用先进的体内成像和免疫组织化学方法来测试生理刺激对NHE3的体内抑制是否涉及PP2A对NHE3的去磷酸化。通过提出揭示信号通路的分子细节，当信号通路受损时，该通路已与高血压的发病机制有关，该项目显然与NIDDK支持影响公共健康的最严重疾病的研究的使命有关。

DESCRIPTION (provided by applicant): Dr. Ion Alexandru Bobulescu is a physician-scientist recently appointed to a full-time faculty position at the instructor level at the University of Texas Southwestern Medical Center at Dallas (UT Southwestern). His short-term career goals (next 4 years) are to rigorously pursue the training and career development plan outlined in this K01 application, perform the proposed experiments using both established and highly innovative techniques to define the role of Protein Phosphatase 2A (PP2A) in the regulation of renal sodium/proton exchange, and then apply for funding under the R01 mechanism to explore one or more of the scientific opportunities derived from this work. Dr. Bobulescu's long-term career goal is to be a leader and innovator in the field of renal regulation of sodium and blood pressure homeostasis, using pioneering approaches and techniques to address fundamental unanswered questions from molecule to model organism to human disease. To achieve these goals, a thorough and detailed Career Development Plan is proposed to address the following training requirements: (1) COURSEWORK. Additional formal training in several areas is considered critical for the advancement of the candidate's research. (2) ADVANCED TECHNIQUES. Working with his mentor and expert collaborators on a hands-on basis to address the Aims of this proposal, Dr. Bobulescu will learn several novel, state-of-the-art methods, and will gain the ability to apply these techniques for his own research questions. (3) MENTORED CAREER DEVELOPMENT. Dr. Bobulescu's chances of achieving relevant results in the new and understudied field of signal transduction via protein phosphatases will greatly benefit from having an expert mentor who will directly guide, supervise and evaluate his research. The mentor for this K01, Dr. Marc Mumby, is a world-renowned expert in protein phosphatase research. In addition, Dr. Bobulescu is aware of the fact that a successful independent career requires much more than ideas, technical skills, and a good track record of publications. Running a research laboratory, managing and nurturing people, fostering collaborations, being an effective mentor and teacher, and being able to maintain a strong and productive research program resulting in successful funding beyond the first R01, are all formidable endeavors- but vital for true academic success. The candidate is certain that he has a lot to learn in these areas from Dr. Mumby, a leading scientist and ideal mentor for this K01. (4) DEPARTMENT. The disease-oriented perspective offered by Dr. Bobulescu's previous postdoctoral research training in a clinical department has been very important for his formation as a physician scientist. However, his training would greatly benefit from a period of immersive exposure to the vastly different environment of a leading basic science department. ENVIRONMENT: UT Southwestern is one of the world's foremost research institutions, with four Nobel prize winners, 36 members of the National Academy of Sciences and/or its Institute of Medicine, more than 3,500 research projects under way with more than $400 million in annual funding, as well as more than 582,000 sq ft of research space with more under construction. Dr. Bobulescu will train primarily in the laboratory of Dr. Marc Mumby in the Department of Pharmacology, including over 2,000 sq ft of total laboratory space, well equipped with all necessary items needed to carry out biochemical, molecular biological, and cell biological experiments. Dr. Bobulescu will also retain his 250 sq ft laboratory space, private 90 sq ft office and faculty position in the Department of Internal Medicine. The candidate will have access to UT Southwestern's state of the art animal research facility, as well as to the UT Southwestern George M. O'Brien Kidney Research Core Center. UT Southwestern has a tradition of retaining the best postdoctoral fellows and fostering their independent careers. This tradition includes the candidate's former mentor, Dr. Orson Moe, who was a postdoctoral fellow at UT Southwestern until 1990 and has remained there ever since, rising to full Professor and Center Director. RESEARCH: This proposal will study the role of PP2A in the regulation of the principal renal sodium transporter NHE3, with special emphasis on the regulation of NHE3 by intra-renal dopamine. If the proposed hypotheses are correct, PP2A will be established as a major regulator of intrarenal dopamine action and of NHE3. This will have a direct impact on our understanding of the physiology and pathophysiology of blood pressure regulation, since impaired dopamine-mediated natriuresis has been described in hypertensive humans, has been attributed to defective intra-renal dopamine signaling in two rodent models of hypertension, and has been linked to defective renal PP2A function. The proposal includes three independent aims that will be pursued simultaneously using a variety of techniques. Aim 1 will use cell culture and in vitro experiments to test whether the inhibition of NHE3 by dopamine requires the direct action of PP2A on NHE3. Aim 2 will use cell culture, biochemistry and advanced electrophysiological techniques to test whether dephosphorylation of NHE3 by PP2A acts as an intracellular switch determining the fate of NHE3. Aim 3 will use advanced in vivo imaging and immunohistochemistry to test whether the inhibition of NHE3 in vivo by physiologic stimuli involves NHE3 dephosphorylation by PP2A. By proposing to uncover the molecular details of a signaling pathway that, when impaired, has been implicated in the pathogenesis of hypertension, this project is clearly relevant to the mission of NIDDK to support research on the most serious diseases affecting public health.