RNAi screen reveals an AbI kinase-dependent host cell pathway involved in Pseudomonas aeruginosa internalization

• Type of Material: Article
• Authors: Julia F. Pielage, University of California San Francisco; Kimberly R. Powell, Emory University; Daniel Kalman, Emory University; Joanne N. Engel, University of California San Francisco
• Language: English
• Date: 2008-03-01
• Publisher: Public Library of Science
• Publication Version: Final Published Version
• Copyright Statement: © 2008 Pielage et al.
• License: Creative Commons Attribution 4.0 International
• Final Published Version (URL): http://dx.doi.org/10.1371/journal.ppat.1000031
• Title of Journal or Parent Work: PLoS Pathogens
• ISSN: 1553-7366
• Volume: 4
• Issue: 3
• Start Page: e1000031
• End Page: e1000031
• Grant/Funding Information: This work was supported by NIH grants AI065902 (to JNE), AI42806 (to JNE), and AI056067-01 (to DK).
• Keywords: VIROLOGY; Virology; EPITHELIAL-CELLS; ESCHERICHIA-COLI; MICROBIOLOGY; GUANINE-NUCLEOTIDE EXCHANGE; SIGNAL-TRANSDUCTION; EXOT CONTRIBUTES; GTPASE-ACTIVATING PROTEIN; Parasitology; Life Sciences & Biomedicine; TYROSINE KINASES; RIBOSYLTRANSFERASE SUBSTRATE-SPECIFICITY; PARASITOLOGY; Science & Technology; MEDIATED APOPTOSIS; EXOENZYME S; Microbiology
• Research Categories: Health Sciences, Pathology; Health Sciences, Immunology; Biology, Microbiology

Abstract

Internalization of the pathogenic bacterium Pseudomonas aeruginosa by non-phagocytic cells is promoted by rearrangements of the actin cytoskeleton, but the host pathways usurped by this bacterium are not clearly understood. We used RNAi-mediated gene inactivation of ∼80 genes known to regulate the actin cytoskeleton in Drosophila S2 cells to identify host molecules essential for entry of P. aeruginosa. This work revealed Abl tyrosine kinase, the adaptor protein Crk, the small GTPases Rac1 and Cdc42, and p21-activated kinase as components of a host signaling pathway that leads to internalization of P. aeruginosa. Using a variety of complementary approaches, we validated the role of this pathway in mammalian cells. Remarkably, ExoS and ExoT, type III secreted toxins of P. aeruginosa, target this pathway by interfering with GTPase function and, in the case of ExoT, by abrogating P. aeruginosa-induced Abl-dependent Crk phosphorylation. Altogether, this work reveals that P. aeruginosa utilizes the Abl pathway for entering host cells and reveals unexpected complexity by which the P. aeruginosa type III secretion system modulates this internalization pathway. Our results furthermore demonstrate the applicability of using RNAi screens to identify host signaling cascades usurped by microbial pathogens that may be potential targets for novel therapies directed against treatment of antibiotic-resistant infections.