title: RNAi screen of Salmonella invasion shows role of COPI in membrane targeting of cholesterol and Cdc42.
abstract: The pathogen Salmonella Typhimurium is a common cause of diarrhea and invades the gut tissue by injecting a cocktail of virulence factors into epithelial cells, triggering actin rearrangements, membrane ruffling and pathogen entry. One of these factors is SopE, a G-nucleotide exchange factor for the host cellular Rho GTPases Rac1 and Cdc42. How SopE mediates cellular invasion is incompletely understood. Using genome-scale RNAi screening we identified 72 known and novel host cell proteins affecting SopE-mediated entry. Follow-up assays assigned these 'hits' to particular steps of the invasion process; i.e., binding, effector injection, membrane ruffling, membrane closure and maturation of the Salmonella-containing vacuole. Depletion of the COPI complex revealed a unique effect on virulence factor injection and membrane ruffling. Both effects are attributable to mislocalization of cholesterol, sphingolipids, Rac1 and Cdc42 away from the plasma membrane into a large intracellular compartment. Equivalent results were obtained with the vesicular stomatitis virus. Therefore, COPI-facilitated maintenance of lipids may represent a novel, unifying mechanism essential for a wide range of pathogens, offering opportunities for designing new drugs.
authors.affiliation: Institute of Microbiology, D-BIOL, ETH Zürich, Zürich, Switzerland.
authors.forename: Benjamin
authors.initials: B
authors.lastname: Misselwitz
authors.forename: Sabrina
authors.initials: S
authors.lastname: Dilling
authors.forename: Pascale
authors.initials: P
authors.lastname: Vonaesch
authors.forename: Raphael
authors.initials: R
authors.lastname: Sacher
authors.forename: Berend
authors.initials: B
authors.lastname: Snijder
authors.forename: Markus
authors.initials: M
authors.lastname: Schlumberger
authors.forename: Samuel
authors.initials: S
authors.lastname: Rout
authors.forename: Manuel
authors.initials: M
authors.lastname: Stark
authors.forename: Christian
authors.initials: C
authors.lastname: von Mering
authors.forename: Lucas
authors.initials: L
authors.lastname: Pelkmans
authors.forename: Wolf-Dietrich
authors.initials: WD
authors.lastname: Hardt
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism; D002784:Cholesterol / Q000378:metabolism*; D020755:Coat Protein Complex I / Q000235:genetics / Q000378:metabolism*; D004705:Endocytosis; D020662:Guanine Nucleotide Exchange Factors / Q000235:genetics / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D010641:Phenotype; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000737:chemistry; D012480:Salmonella Infections / Q000235:genetics / Q000378:metabolism / Q000382:microbiology; D012486:Salmonella typhimurium / Q000235:genetics / Q000378:metabolism* / Q000472:pathogenicity; D037521:Virulence Factors / Q000378:metabolism; D020764:cdc42 GTP-Binding Protein / Q000235:genetics / Q000378:metabolism*; D020830:rac1 GTP-Binding Protein / Q000378:metabolism
chemical_list: D001426:Bacterial Proteins; D020755:Coat Protein Complex I; D020662:Guanine Nucleotide Exchange Factors; D034741:RNA, Small Interfering; C103987:SopE protein, Salmonella; D037521:Virulence Factors; D002784:Cholesterol; D020764:cdc42 GTP-Binding Protein; D020830:rac1 GTP-Binding Protein