Shigella is a major cause of diarrhea, dysentery, and mortality annually worldwide. S. flexneri is a CDC/NIAID Category B priority pathogen. It causes disease by invading and spreading through the colonic mucosa. The bacteria enter cells by inducing a process that resembles macropinocytosis. Once within the cell, they move to the cell periphery by means of actin-based motility. At the cell periphery, S. flexneri pushes out against the plasma membrane, forming a protrusion that contains the bacterium at its tip. The protrusion is then engulfed by an adjacent cell, whereupon the cycle of actin-based motility and cell-to-cell spread is repeated. Whereas the mechanisms by which S. flexneri enters cells have been studied extensively, the molecular mechanisms involved in S. flexneri spread from one cell into an adjacent cell are poorly understood. In this application, we propose a detailed investigation of S. flexneri intercellular spread that utilizes both targeted and genomewide approaches. Our targeted approaches will test the hypothesis that the cellular pathway of filopodia extension, which requires diaphanous formins and VASP, is used by S. flexneri to enhance its intercellular spread. Our genome-wide approach will examine the human genome for additional factors required for intercellular spread.
Our specific aims are: 1. Characterize the role of the S. flexneri effector OspE2 and eukaryotic VASP in intercellular spread; 2. Characterize the role of diaphanous formins in the formation of plasma membrane protrusions by S. flexneri;and, 3. Identify and characterize other host factors required for S. flexneri intercellular spread using a genome-wide human siRNA screen; We believe that these approaches will lead to insights not only into the mechanisms of intercellular spread of S. flexneri, but also into fundamental mechanisms of eukaryotic cellular and intercellular processes.

Agency
National Institute of Health (NIH)
Institute
National Institute of Allergy and Infectious Diseases (NIAID)
Type
High Priority, Short Term Project Award (R56)
Project #
3R56AI073967-01A2S1
Application #
7908271
Study Section
Host Interactions with Bacterial Pathogens Study Section (HIBP)
Program Officer
Mills, Melody
Project Start
2009-09-26
Project End
2010-08-31
Budget Start
2009-09-26
Budget End
2010-08-31
Support Year
1
Fiscal Year
2009
Total Cost
$428,793
Indirect Cost
Name
Massachusetts General Hospital
Department
Type
DUNS #
073130411
City
Boston
State
MA
Country
United States
Zip Code
02199
Heindl, Jason E; Saran, Indrani; Yi, Chae-ryun et al. (2010) Requirement for formin-induced actin polymerization during spread of Shigella flexneri. Infect Immun 78:193-203