Functional genomics of human variation to Salmonella invasion

描述(申请人提供)：尽管公共卫生有所改善，疫苗不断进步，许多种类的抗生素也在发展，但传染病仍占全球所有死亡人数的四分之一以上。然而，即使是最具破坏性的大流行，历史表明感染的严重程度和持续时间也有很大的差异。候选人Dennis Ko博士的长期研究目标是确定细菌感染易感性差异的遗传基础。这一知识对于确定为什么一些人对不同的感染具有抵抗力以及开发治疗方法以降低易感人群的死亡率和发病率非常重要。这项提案中的实验将阐明人类遗传差异如何导致伤寒沙门氏菌感染的变异。第一个目标是确定改变伤寒沙门氏菌入侵培养细胞的人类基因差异。这是通过一种新的遗传关联筛查实现的，该筛查使用从数百个基因分型的个体获得的细胞。然后将通过测量RNA干扰对伤寒沙门氏菌入侵的影响来验证筛查揭示的遗传变异。第二个目标是确定已识别的基因变异如何影响伤寒沙门氏菌入侵的细胞机制。荧光显微镜和流式细胞术将用于确定细菌的摄取或细胞内生态位的早期建立是否受到影响。进一步的实验将包括对已知在沙门氏菌入侵中发挥作用的分子的干扰和评估，包括肌醇磷脂、Rho家族GTP酶和分泌的细菌效应物。第三个目标是使用小鼠模型和临床关联数据来评估已识别的疾病差异的相关性。小鼠模型将被用来确定已识别的基因是否影响感染的建立和/或在宿主内的传播。作为这些研究的补充，将通过病例对照遗传关联研究来探讨基因差异与伤寒和其他疾病的相关性。这些拟议实验的总体目标是发现基本的细胞生物学机制以及对健康和疾病至关重要的人类基因变异。Ko博士是塞缪尔·米勒博士实验室的生命科学研究基金会研究员。他获得了斯坦福大学的医学博士学位。他对C型脂质储存障碍的论文研究为遗传学和细胞生物学提供了坚实的基础，他已经将这些学科应用于他目前关于理解人类对细菌感染的变异的研究中。作为一名博士后，他的工作重点是开发一种新的筛查策略，以发现修改沙门氏菌诱导的细胞死亡的基因变异，并确定这些变异如何影响人类健康。高博士计划启动一个独立的研究实验室，研究伤寒沙门氏菌入侵的易感性，目标是将他的重点扩大到宿主和病原体关系双方的变异和适应的重要性。

DESCRIPTION (provided by applicant): Despite improvements in public health, advancements in vaccines, and the development of many classes of antibiotics, infectious disease is still responsible for over a quarter of all deaths worldwide. However, even for the most devastating of pandemics, history demonstrates a large variability in the severity and duration of infection. The long-term research goal of the candidate, Dr. Dennis Ko, is to determine the genetic basis for differences in susceptibility to bacterial infections. This knowledge is important for determining why some individuals are resistant to different infections and in developing therapies to decrease the mortality and morbidity of susceptible individuals. Experiments in this proposal will elucidate how human genetic differences contribute to variation in infection by the bacteria that causes typhoid fever, Salmonella typhi. The first aim is to identify human genetic differences that modify invasion of S. typhi into cultured cells. This is accomplished through a novel genetic association screen using cells obtained from hundreds of genotyped individuals. Genetic variants revealed by the screen will then be validated by measuring the effect of RNA interference on S. typhi invasion. The second aim is to determine how the identified genetic variants affect the cellular mechanism of S. typhi invasion. Fluorescence microscopy and flow cytometry will be used to determine if uptake of the bacteria or early establishment of an intracellular niche is being affected. Further experiments will include perturbation and assessment of molecules known to play roles in Salmonella invasion, including phosphoinositides, Rho-family GTPases, and secreted bacterial effectors. The third aim is to assess the relevance of identified differences on disease using mouse models and clinical association data. Mouse models will be used to determine if the identified genes affect establishment of infection and/or spread within the host. As a complement to these studies, the relevance of the genetic differences upon typhoid fever and other diseases will be addressed through case-control genetic association studies. The overall goal of these proposed experiments is discovery of both basic cell biological mechanism as well as human genetic variation important for health and disease. Dr. Ko, is a Life Sciences Research Foundation fellow in the laboratory of Dr. Samuel Miller. He received MD-PhD degrees from Stanford University. His thesis research on the lipid-storage disorder Niemann-Pick Type C provided a firm foundation in genetics and cell biology, and he has applied these disciplines to his current research on understanding human variation to bacterial infection. As a postdoctoral fellow, his efforts focused on developing a novel screening strategy for discovering genetic variants that modify Salmonella- induced cell death and determining how these variants affect human health. Dr. Ko plans to start an independent research lab studying susceptibility to S. typhi invasion, with the goal of expanding his focus to the importance of variation and adaptation on both sides of the host-pathogen relationship.