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）：一些细胞内细菌病原体，如李斯特菌，立克次体和志贺氏菌，依靠基于肌动蛋白的运动性从感染细胞传播到邻近细胞并在宿主内传播。遗传和生化研究表明，基于肌动蛋白的运动依赖于细菌表面产生的毒力因子，这些毒力因子与宿主细胞骨架因子相互作用，包括ARP2/3复合物。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。我们使用这种新开发的方法筛选人类激酶，并鉴定出CK1和CK2是李斯特菌传播所需的宿主细胞激酶。我们最近证明，与WASP和WAVE2类似，ActA对ARP2/3复合物的亲和力是由ck2介导的磷酸化调节的，我们将这一概念称为调控模仿。在这篇文章中，我们提出了遗传和生化的方法来探索CK1在李斯特菌肌动蛋白运动（Aim1）调控中的作用。此外，我们建议将用于研究李斯特菌传播的方法扩展到几种显示肌动蛋白运动的细菌病原体，包括志贺氏菌和立克次体（Aim2）。这些探索性研究可能为支持李斯特菌、志贺氏菌和立克次体发病机制以及参与肌动蛋白细胞骨架调节的基本过程提供重要和新颖的见解。