Characterization of Novel Signaling Pathways Involved in Water Balance Disorders

项目概要/摘要 维持水平衡是肾脏的重要功能，对于适应陆地环境至关重要。 生活为了有效地重吸收水分，释放加压素（VP）以诱导水通道蛋白-2（AQP 2）磷酸化 以及集合管中肾脏主细胞内的肌动蛋白细胞骨架重塑， AQP 2的膜表达。AQP 2运输的失调导致水平衡紊乱; AQP 2膜表达减少导致肾源性尿崩症（NDI），而AQP 2膜表达增加导致尿崩症（NDI）。 在ADH分泌不当综合征中，质膜AQP 2与液体潴留相关 （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AQP 2膜运输和水的重吸收，这个过程实际上要复杂得多， 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），其抑制诱导AQP 2膜蓄积和磷酸化 类似于VP，但绕过V2 R、cAMP和PKA。因此，该补助金的目的是：1）确定 调节AQP 2运输的VP和EGFR信号通路之间的串扰机制; 2） 了解AQP 2磷酸化在细胞骨架重塑中的作用，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， 以便最终设计治疗方法来减轻临床上遇到的疾病症状。 培训计划是渐进式的，旨在帮助PI张医生完成以下初始阶段： 完成和扩大她的令人兴奋的，正在进行的工作EGF/VP串扰（目标1），然后转移到一个新的 AQP 2细胞骨架相互作用的一个方面，在最终达到目标3之前，尚未在实验室中进行研究（目标2） 心脏/肾脏的相互作用和水平衡，这将完成她的独立之路， 第一次申请R 01。张博士在她的导师布朗博士的指导下，不仅将雇用 现有的技术在膜生物学计划，但也将获得重要的新技术， 研究培训，既从课程中列出了她的个性化培训计划，并从专家对她的 顾问委员会Jorge博士（CRISPR/Cas9基因编辑）和Najordorf博士（手术和小鼠成像， 心力衰竭）。对于专业的职业发展，除了她的获奖导师布朗博士，她 西尔维布雷东和朱迪·巴比特博士是成功的女性学术科学家的榜样。而且 MGH肾脏科主任Ravi Thadhani博士将保护她的研究时间不受临床职责的影响， 全身心投入她的研究她的导师，她的科学顾问委员会成员，她的部门主管， 在MGH和哈佛社区的培育和合作环境，将结合联合收割机， 设计了科学计划，以推动她朝着成为一个独立的R 01资助的学术目标 肾脏科医生，擅长水调节中的器官间通信。

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.