Cytoskeletal regulation of intestinal barrier function and response to pathogens

描述（由申请人提供）：候选人是胃肠病学家，对基础科学研究有很强的承诺，对宿主-病原体相互作用背景下的上皮细胞生物学有特殊的兴趣。候选人的长期职业目标是成为一名独立资助的肠道生物学基础科学研究者，并为我们理解哺乳动物上皮屏障完整性在稳态和与肠道微生物相互作用过程中的调节机制做出根本性贡献。候选人的短期职业目标是：1)精通分子、细胞和生物化学技术，这些技术有助于在体外和体内机械地研究细胞骨架动力学；2)获得在概念和技术上前沿的实验设计方法方面的专业知识，以新的方式阐明宿主-病原体相互作用；3)在培养免疫学、上皮生物学和微生物学专家之间创造性和高度跨学科合作方面积累专业经验；4)提供足够的初步数据和出版物，作为竞争性R01资助的基础，作为学术医学研究事业的基石。该项目的总体主题是了解调节肌动蛋白细胞骨架的因素如何影响肠上皮内细胞间连接的稳定性和动态功能。根尖连接复合体（紧密连接和粘附连接）是肠通透性的主要调节器，必须能够快速微调其闭塞性或漏性。神经Wiskott-Aldrich综合征蛋白（N-WASP）是肌动蛋白成核分子家族的典型成员，在被多种宿主或病原体编码的因子激活后，与Arp2/3复合物相互作用，促进肌动蛋白细丝的时空限制性组装。这一过程是多种细胞过程的核心，但越来越多的证据表明，极化上皮细胞系和果蝇表明，N-WASP在调节涉及细胞间连接维持的动态蛋白质运输事件中起着核心和特定的作用，沿着这条线，人们越来越多地认识到病原体，如EPEC和C. rodentium，专门针对N-WASP调节肌动蛋白细胞骨架并破坏肠道屏障功能。这个的具体目标

DESCRIPTION (provided by applicant): The candidate is a gastroenterologist with an extremely strong commitment to basic science research, and a specific interest in epithelial cell biology in the context of host-pathogen interactions. The candidate's long- term career goal is to become an independently funded investigator in the basic science of intestinal biology, and to make fundamental contributions to our understanding of the mechanisms that regulate mammalian epithelial barrier integrity during homeostasis and during interaction with intestinal microbes. The candidate's short-term career goals are 1) to become proficient in molecular, cellular and biochemical techniques useful for mechanistically investigating cytoskeletal dynamics both in vitro and in vivo; 2) to acquire expertise in conceptually and technologically cutting-edge approaches to experimental design that can elucidate host- pathogen interactions in novel ways; 3) to develop professional experience in fostering creative and highly interdisciplinary collaborations between experts in immunology, epithelial biology and microbiology; 4) to produce sufficient preliminary data and publications that will serve as the basis for competitive R01 funding as the cornerstone of a career in academic medical research. The overall theme of this project is to understand how factors that regulate the actin cytoskeleton influence the stability and dynamic function of intercellular junctions within the intestinal epithelium. The apical junction complex (the tight junction and adherens junction) is the major regulator of intestinal permeability and must be capable of rapidly fine-tuning its occlusiveness or leakiness. The Neural Wiskott-Aldrich Syndrome protein (N-WASP) is the canonical member of a family of actin nucleating molecules that, upon activation by a variety of host- or pathogen-encoded factors, interact with the Arp2/3 complex and promote the spatiotemporally restricted assembly of actin filaments. This process is central to diverse cellula processes, but there is growing evidence from polarized epithelial cell lines and Drosophila that N-WASP plays a central and specific role in regulating the dynamic protein trafficking events involved in intercellular junction maintenance, and along this line, there has been increasing recognition of pathogens, such as EPEC and C. rodentium, that specifically target N-WASP to modulate the actin cytoskeleton and disrupt intestinal barrier function. The specific goals of this study are to test two fundamental hypotheses: 1) N-WASP is an effector molecule that balances both the delivery and removal of AJC proteins through an interaction with the Par3/Par6/aPKC polarity complex, and 2) AE pathogens require N-WASP to induce epithelial barrier dysfunction, and the translocated effector protein EspF targets host N-WASP to enhance removal of AJC proteins during infection. The proposed work will be pursued within the context of an intensive and formalized career development program, which will allow the candidate to acquire expertise in both classic and leading edge molecular and cell biological approaches to studying intestinal biology and bacterial interactions, and will take advantage of the highly collaborative and complementary co-mentorship of Scott Snapper (immune and epithelial biology) and John Leong (microbiology). In addition, a formal research advisory committee, consisting of a diverse group of researchers at MGH and Harvard Medical School with deep expertise in immunology, gut histopathology, and junction biology, has been established and has overseen the candidate's progress since July 2010. The research environment, which includes the MGH Gastrointestinal Unit and the Harvard Digestive Diseases Center at Children's Hospital Boston, will provide an intellectually enriching, technically resourceful and collaborative atmosphere which will catalyze the candidate's scientific productivity. At the conclusion of the award period, the candidate will be well positioned for a career as an independent, R01-funded investigator.