Role of endocytosis in beta-adrenergic receptors signaling in cardiomyocytes

描述（由申请人提供）：项目摘要/摘要<$-肾上腺素能受体（<$AR）在控制心脏收缩的强度和频率方面发挥核心作用，并引起GS-连接的cAMP生成。AR信号传导的紊乱与许多病理状况如高血压和心力衰竭有关。在激动剂诱导的激活后，AR经历功能性脱敏和从质膜（PM）内吞移除。一旦进入细胞，受体以前被认为是功能不活跃的。结合先进的成像平台与最近开发的构象生物传感器，Irannejad博士直接探测了<$2AR及其同源Gs蛋白的激活，并发现活性<$2AR-G <$s复合物不仅限于PM，也存在于内体。作为一名独立的生物医学研究人员，她的长期目标是了解AR内体信号在调节心脏功能中的功能后果。有了这个奖项，Irannejad博士将研究内吞作用在心肌细胞中AR信号传导中的作用。这个R99/R 00奖将使她实现以下职业目标：1）光学显微镜的额外培训，2）心肌细胞生物学的额外培训3）发展技能，进行蛋白质复合物的蛋白质组学研究，4）发展沟通，指导，赠款写作和实验室管理技能，成为成功的，独立的生物医学研究人员。本研究的指导阶段将在加州大学旧金山分校弗朗西斯科进行，由Mark von Zastrow博士指导。在指导阶段，Irannejad博士将使用心肌细胞中的构象生物传感器来识别区室化的AR信号（Aim 1）。接下来，她将阐明内吞作用在心肌细胞中<$1AR与<$2AR激活介导的信号传导和收缩率中的作用（目的2）。Shaun Coughlin博士（UCSF）是加州大学旧金山分校CVRI主任，也是心血管生物学专家，他将担任这一目标的共同导师。为了鉴定核内体中的活性2AR-Gs相互作用组（Aim 3），Irannejad博士将开始使用蛋白质组学方法。Nevan Krogan博士（UCSF），蛋白质复合物的蛋白质组学研究专家将担任合作者和顾问。在独立阶段，Irannejad博士将继续专注于目标2和3，表征<$ARs-Gs介导的信号传导的动力学，了解<$2AR内体信号传导的功能后果，并确定<$2AR-Gs内体相互作用。该奖项将使Irannejad博士能够阐明AR区室化信号及其在调节心脏功能方面的功能重要性，并可能开发新的更好的药物来治疗病理性心脏功能。

DESCRIPTION (provided by applicant): Project Summary/Abstract ¿-adrenergic receptors (¿ARs) play a central role in controlling the strength and frequency of cardiac contraction, and elicit Gs-linked cAMP generation. Disturbances of ¿AR signaling have been implicated in many pathological conditions such as hypertension and heart failure. Upon agonist-induced activation, ¿ARs are subject to a functional desensitization and endocytic removal from the plasma membrane (PM). Once inside the cells, the receptors were previously thought to be functionally inactive. Combining sophisticated imaging platforms with recently developed conformational biosensors, Dr. Irannejad directly probed activation of ¿2AR and its cognate Gs protein and discovered that active ¿2AR-G¿s complex is not restricted to the PM and is also present at endosomes. Her long-term goal as an independent biomedical researcher is to understand the functional consequence of the ¿AR endosomal signaling in regulating heart function. With this award, Dr. Irannejad will examine the role of endocytosis in ¿ARs signaling in cardiomyocytes. This R99/R00 award will allow her to achieve the following career goal: 1) additional training in optical microscopy, 2) additional training in cardiomyocytes biology 3) develop skills to perform proteomic studies of protein complexes, 4) develop the communication, mentoring, grant-writing, and laboratory management skills necessary to become and successful, independent biomedical researcher. The mentored phase of this research will be carried out at the University of California, San Francisco under the guidance of Dr. Mark von Zastrow. During the mentored phase, Dr. Irannejad will use the conformational biosensors in cardiomyocytes to indentify compartmentalized ¿ARs signaling (Aim1). Next, she will elucidate the role of endocytosis in signaling and contraction rate mediated by ¿1AR versus ¿2AR activation in cardiomyocytes (Aim 2). Dr. Shaun Coughlin (UCSF) director of CVRI at UCSF and an expert in cardiovascular biology will serve as co-mentor for this Aim. To indentify active ¿2AR-Gs interactomes at the endosome (Aim 3), Dr. Irannejad will begin using proteomic approach. Dr. Nevan Krogan (UCSF), experts in proteomic studies of protein complexes will serve as collaborator and advisor. During the independent phase, Dr. Irannejad will continue to focus on Aim 2 and 3, characterize the dynamics of ¿ARs-Gs mediated signaling, understand the functional consequence of the ¿2AR endosomal signaling, and identifying ¿2AR-Gs interactomes at the endosome. This award will enable Dr. Irannejad to elucidate the ¿ARs compartmentalize signaling and its functional importance in regulating heart function and potentially developing new and better drugs for the treatment of pathological cardiac function.