Exploring the roles of Gi and S1Pr1 in endothelial barrier function in vivo

项目总结/摘要。这个职业发展奖的目标是博士. Wilsbacher 发展必要的技能，成功地成为一个独立的物理学家，科学家在学术中心。 博士威尔斯巴赫获得了医学博士学位和博士西北大学医学科学家培训项目 在大学里，她研究了驱动昼夜节律的遗传和分子机制。她 在加州大学旧金山分校弗朗西斯科接受内科和心脏病学培训。威尔斯巴赫医生 目前的研究目的是探讨正常生理过程中血管屏障功能的机制 她的长期职业目标是弥合昼夜生理学和内皮细胞之间的差距， 功能，以帮助促进心血管生物学和疾病的机制的理解。 培训计划包括由世界级的G蛋白研究者肖恩·考夫林博士指导， 在止血、血栓形成和血管完整性中的GPCR信号传导， 培训年轻调查员独立的记录。将由一名专家提供额外指导 一组科学家是GPCR信号传导（Mark von Zastrow博士）、炎症（Donald McDonald）和蛋白质化学（James威尔斯博士）。优秀的研究设施和设备， available.该培训计划还包括先进的教学课程，重点是细胞信号， 膜运输，蛋白质相互作用和显微镜;出席系列研讨会;参与和 在地方和国家会议上作报告;指导编写手稿和赠款。 治疗或预防炎症需要彻底了解内皮细胞的机制， 屏障功能这项研究计划旨在揭示调节血管完整性的信号机制， 特别是通过鞘氨醇-1-磷酸受体1（S1Pr1）和Gi信号传导。目标1研究角色 内皮细胞Gi和S1Pr1信号在屏障功能中的作用。具体来说，基因小鼠模型， - 内皮中的百日咳毒素（其抑制Gi），在内皮中表达百日咳不敏感的Gi，或 将测试在成人内皮中有条件地缺失S1Pr1的人的血管通透性的变化。Aim 2用途 一种新的GPCR激活检测系统，用于探测S1Pr1在体内何时何地被激活， 炎症转录报告GPCR激活的新报告系统是一种创新的 该工具将广泛适用于不同组织中的其他GPCR。目的3探讨小窝是否影响 S1Pr1-Gi偶联，以及S1Pr1在小窝中的定位是否影响抑制蛋白介导的内化。 Wilsbacher博士的职业发展奖提案包括一个有前途的候选人，杰出的 培训环境，严格的培训计划和令人兴奋的研究计划，其中包括一个创新的新的 该工具侧重于与人类健康和疾病直接相关的主题。在完成这项工作后， Wilsbacher博士应该具备作为一名独立调查员取得成功所需的技能。

PROJECT SUMMARY/ABSTRACT. The goal of this Career Development Award is for Dr. Wilsbacher to develop the skills necessary to successfully become an independent physician-scientist at an academic center. Dr. Wilsbacher earned her M.D. and Ph.D. degrees in the Medical Scientist Training Program at Northwestern University, where she investigated genetic and molecular mechanisms that drive circadian rhythms. She trained in Internal Medicine and Cardiology at the University of California, San Francisco. Dr. Wilsbacher's current research objective is to investigate mechanisms of vascular barrier function during normal physiology and disease, and her long-term career goal is to bridge the gap between circadian physiology and endothelial function to help advance understanding of mechanisms governing cardiovascular biology and disease. The training plan includes mentorship by Dr. Shaun Coughlin, a world-class investigator of G protein- coupled receptor (GPCR) signaling in hemostasis, thrombosis, and vascular integrity who has an established record of training young investigators to independence. Additional guidance will be provided from an expert group of scientists who are leaders in GPCR signaling (Dr. Mark von Zastrow), inflammation (Dr. Donald McDonald), and protein chemistry (Dr. James Wells). Outstanding research facilities and equipment are readily available. The training plan also incorporates advanced didactic coursework that focuses on cell signaling, membrane trafficking, protein interactions, and microscopy; attendance at seminar series; participation and presentation in local and national conferences; and mentored guidance with manuscript and grant preparation. Treating or preventing inflammation requires thorough understanding of the mechanisms of endothelial barrier function. The research proposal aims to uncover signaling mechanisms that regulate vascular integrity, particularly through sphingosine-1-phosphate receptor 1 (S1Pr1) and Gi signaling. Aim 1 investigates the role of endothelial Gi and S1Pr1 signaling in barrier function. Specifically, genetic mouse models that express pertussis toxin (which inhibits Gi) in endothelium, express a pertussis-insensitive Gi in endothelium, or conditionally delete S1Pr1 in adult endothelium will be tested for changes in vascular permeability. Aim 2 uses a new GPCR activation detection system to probe when and where S1Pr1 is activated in vivo during inflammation. The new reporter system, which transcriptionally reports on GPCR activation, is an innovative tool that will be widely applicable to other GPCRs in diverse tissues. Aim 3 probes whether caveolae influence S1Pr1-Gi coupling, and whether S1Pr1 localization in caveolae affects ¿arrestin-mediated internalization. Dr. Wilsbacher's Career Development Award proposal comprises a promising candidate, outstanding training environment, rigorous training plan, and exciting research proposal that includes an innovative new tool and focuses on a topic with direct relevance to human health and disease. At the completion of this Award, Dr. Wilsbacher should have the skills necessary to succeed as an independent investigator.