Regulation of bacterial pathogen actin-based motility by host cell kinases

宿主细胞激酶对细菌病原体肌动蛋白运动的调节

• 批准号: 8095076
• 负责人: HERVE F AGAISSE
• 金额: $ 24.83万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-03-01 至 2013-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8095076
• 关键词: Actins; Affect; Affinity; Biochemical; Biological Assay; CSNK1A1 gene; Cell physiology; Cells; Comparative Study; Complex; Computer-Assisted Image Analysis; Cytoskeleton; Cytosol; Defect; Fluorescence Microscopy; Foundations; Genetic; Human; Integration Host Factors; Knowledge; Lead; Listeria; Mediating; Methodology; Molecular; Mutagenesis; N-terminal; Pathogenesis; Phosphorylation; Phosphotransferases; Play; Preventive Intervention; Process; Production; Protein-Serine-Threonine Kinases; RNA Interference; Regulation; Research Design; Rickettsia; Role; Screening procedure; Seminal; Shigella; Site; Substrate Specificity; Surface; Tail; Therapeutic Intervention; Virulence Factors; Work; base; cell motility; design; innovation; insight; microbial; mimicry; novel; novel strategies; pathogen; research study

DESCRIPTION (provided by applicant): Several intracellular bacterial pathogens, such as Listeria ssp, Rickettsia spp. and Shigella spp., rely on actin-based motility to spread from infected cells to neighboring cells and disseminate within their host. Genetic and biochemical studies have revealed that actin-based motility relies on the production of virulence factors at the bacterial surface that interact with host cell cytoskeleton factors, including the ARP2/3 complex. In spite of these seminal contributions, the host cell factors supporting actin-based motility are still poorly understood. To fill this gap in knowledge, we have developed innovative approaches combining automated fluorescence microscopy and computer-assisted image analysis in order to visualize and quantify bacterial pathogen spread. We have used this newly developed approach to screen the human kinome and identified CK1 and CK2 as host cell kinases required for Listeria spread. We recently demonstrated that, similar to WASP and WAVE2, the affinity of ActA for the ARP2/3 complex is modulated by CK2-mediated phosphorylation, a notion that we refer to as regulatory mimicry. In this proposal we propose genetic and biochemical approaches designed to explore the role(s) of CK1 in the regulation of Listeria actin-based motility (Aim1). In addition, we propose to extend the approach used for investigating Listeria spread to several bacterial pathogens displaying actin-based motility, including Shigella and Rickettsia (Aim2). These exploratory studies may provide important and novel insights both into the mechanisms supporting Listeria, Shigella and Rickettsia pathogenesis and into the fundamental processes involved in the regulation of the actin cytoskeleton. PUBLIC HEALTH RELEVANCE: Various intracellular pathogens have evolved the ability to manipulate host cell processes in order to spread from infected cells into the neighboring cells. Most of he host factors contributing to the cell-to-cell spread of pathogens are unknown. In this proposal, we present our plans to determine and investigate the host cell kinases involved in Listeria spp., Shigella spp. and Rickettia spp. actin-based motility. The proposed approach will contribute to our general understanding of the mechanisms underlying microbial pathogenesis and may therefore constitute the foundation for the rational design of preventive and therapeutic interventions.

描述（由申请人提供）：几种细胞内细菌病原体，如李斯特菌属、立克次氏体属。和志贺氏菌属，依靠肌动蛋白为基础的运动性从感染细胞传播到邻近细胞并在其宿主内传播。遗传和生物化学研究表明，肌动蛋白为基础的运动依赖于在细菌表面的毒力因子的生产，与宿主细胞骨架因子，包括ARP 2/3复合物相互作用。尽管这些开创性的贡献，宿主细胞因子支持肌动蛋白为基础的运动仍然知之甚少。为了填补这一知识空白，我们开发了结合自动荧光显微镜和计算机辅助图像分析的创新方法，以可视化和量化细菌病原体的传播。我们已经使用这种新开发的方法来筛选人类激酶组，并确定CK 1和CK 2为李斯特菌传播所需的宿主细胞激酶。我们最近证明，类似于WASP和WAVE 2，ActA对ARP 2/3复合物的亲和力是由CK 2介导的磷酸化调节的，我们称之为调节模拟。在这项提案中，我们提出了遗传和生物化学的方法，旨在探索CK 1在李斯特菌肌动蛋白为基础的运动（Aim 1）的调节作用。此外，我们建议将用于调查李斯特菌传播的方法扩展到几种显示基于肌动蛋白的运动性的细菌病原体，包括志贺氏菌和立克次氏体（Aim 2）。这些探索性的研究可能提供重要的和新的见解支持李斯特菌，志贺氏菌和立克次氏体的发病机制和肌动蛋白细胞骨架的调节所涉及的基本过程。 公共卫生相关性：各种细胞内病原体已经进化出操纵宿主细胞过程的能力，以便从受感染的细胞扩散到邻近细胞中。大多数导致病原体细胞间传播的宿主因素尚不清楚。在这个建议中，我们提出了我们的计划，以确定和调查宿主细胞激酶参与李斯特菌，志贺菌和立克次体属（Rickettia spp.）肌动蛋白为基础的运动。所提出的方法将有助于我们对微生物致病机制的普遍理解，因此可能构成合理设计预防和治疗干预措施的基础。