Characterization of Novel Signaling Pathways Involved in Water Balance Disorders

项目概要/摘要 维持水稳态是肾脏的一项重要功能，对于适应陆地环境至关重要 生活。为了有效地重吸收水，加压素 (VP) 被释放以诱导水通道蛋白 2 (AQP2) 磷酸化 和集合管肾主细胞内的肌动蛋白细胞骨架重塑，这增加了顶端 AQP2 的膜表达。 AQP2 运输失调会导致水平衡紊乱； AQP2 膜表达减少会导致肾性尿崩症 (NDI)，而 AQP2 膜表达增加会导致肾性尿崩症 (NDI) 质膜 AQP2 与 ADH 分泌不当综合征中的液体潴留相关 (SIADH)、充血性心力衰竭和肝硬化。而VP/cAMP/PKA是主要的信号通路 促进 AQP2 膜运输和水重吸收，该过程实际上要复杂得多，并且可以 被其他信号通路诱导或抑制。一种这样的“替代”途径涉及表皮 生长因子受体 (EGFR)，其抑制作用会诱导 AQP2 膜积聚和磷酸化 与 VP 类似，但绕过 V2R、cAMP 和 PKA。因此，这笔赠款的目的是 1) 定义 调节 AQP2 运输的 VP 和 EGFR 信号通路之间的串扰机制； 2） 了解 AQP2 磷酸化对细胞骨架重塑的作用，以及 3) 表征 在充血性心力衰竭患者中观察到的水潴留失调的新信号通路 为了最终设计出缓解临床中遇到的疾病症状的疗法。 培训计划是渐进式的，旨在让 PI 张博士度过初始阶段： 完成并扩展她正在进行的关于 EGF/VP 串扰的令人兴奋的工作（目标 1），然后转向新的 在最终到达目标 3 之前，AQP2 细胞骨架相互作用的方面尚未在实验室中进行检查（目标 2） 关于心/肾相互作用和水平衡，这将完成她的独立之路，并成为 她的第一个 R01 申请的主题。张博士在导师布朗博士的指导下，不仅会聘请 膜生物学计划中的现有技术，但还将获得重要的新技术和 研究培训既来自她的个性化培训计划中列出的课程，也来自她的专家 顾问委员会博士。 Joung（CRISPR/Cas9 基因编辑）和 Nahrendorf（手术和小鼠成像） 心力衰竭）。对于职业生涯发展，除了屡获殊荣的导师布朗博士之外，她 有博士。西尔维·布雷顿和朱迪·巴比特是成功的女性学术科学家的榜样。此外， MGH 肾脏病科主任 Ravi Thadhani 博士将保护她的研究时间，使其免受临床职责的影响，从而允许 完全沉浸在她的研究中。她的导师、她的科学顾问委员会成员、她的部门主管、 麻省总医院和哈佛社区的培育和协作环境，将与精心设计的项目相结合 设计了科学计划来推动她实现成为独立 R01 资助学者的目标 肾病专家，在涉及水调节的器官间通讯方面具有专业知识。

PROJECT SUMMARY/ABSTRACT Maintenance of water homeostasis is a vital function of the kidneys and is essential for adaptation to terrestrial life. To reabsorb water effectively, vasopressin (VP) is released to induce aquaporin-2 (AQP2) phosphorylation and actin cytoskeletal remodeling within kidney principal cells in the collecting ducts, which increases apical membrane expression of AQP2. Dysregulation of AQP2 trafficking results in disorders of water balance; decreased AQP2 membrane expression causes nephrogenic diabetes insipidus (NDI), whereas an increase in plasma membrane AQP2 is associated with fluid retention in the syndrome of inappropriate ADH secretion (SIADH), congestive heart failure and cirrhosis. While VP/cAMP/PKA is the major signaling pathway that facilitates AQP2 membrane trafficking and water reabsorption, the process is in fact far more complex, and can be induced or inhibited by other signaling pathways. One such “alternative” pathway involves the epidermal growth factor receptor (EGFR), whose inhibition induces AQP2 membrane accumulation and phosphorylation similar to VP, but bypasses V2R, cAMP and PKA. Therefore, the aims of this grant are to 1) define the mechanism of crosstalk between VP and EGFR signaling pathways that regulate AQP2 trafficking; 2) understand the role of AQP2 phosphorylation on cytoskeletal remodeling, and 3) characterize the roles of novel signaling pathways in dysregulated water retention observed in patients with congestive heart failure, in order to eventually design therapies to alleviate disease symptoms encountered in the clinic. The training plan is progressive and is designed to move the PI Dr. Cheung through an initial stage of completing and expanding her exciting, ongoing work on EGF/VP crosstalk (Aim 1), then moving on to a new aspect of AQP2 cytoskeletal interactions not so far examined in the lab (Aim 2), before finally arriving at Aim 3 on heart/kidney interactions and water balance, which will complete her pathway to independence and be the subject of her first R01 application. Dr. Cheung, supervised by her mentor Dr. Brown, will not only employ existing technologies in the Program in Membrane Biology, but will also acquire important new techniques and research training both from courses listed in her personalized training plan, and from the experts on her advisory committee Drs. Joung (CRISPR/Cas9 gene editing) and Nahrendorf (surgery and mouse imaging for heart failure). For professional career development, in addition to her award-winning mentor Dr. Brown, she has Drs. Sylvie Breton and Jodie Babitt as successful female academic scientist role models. Moreover, the MGH Nephrology Division chief, Dr. Ravi Thadhani will protect her research time from clinical duties, allowing full immersion in her research. Her mentor, members of her scientific advisory committee, her Division chief, the nurturing and collaborative environment in the MGH and Harvard community, will combine with a carefully designed scientific plan to propel her toward her goal of becoming an independent R01 funded academic nephrologist with expertise in inter-organ communications involved in water regulation.