SteC is a Salmonella kinase required for SPI-2-dependent F-actin remodelling.
Salmonella enterica serovar Typhimurium (S. Typhimurium) replicates inside mammalian cells within membrane-bound compartments called Salmonella-containing vacuoles. Intracellular replication is dependent on the activities of several effector proteins translocated across the vacuolar membrane by the Salmonella pathogenicity island 2 (SPI-2)-type III secretion system (T3SS). This is accompanied by the formation in the vicinity of bacterial vacuoles of an F-actin meshwork, thought to be involved in maintaining the integrity of vacuolar membranes. In this study, we investigated the function of the SPI-2 T3SS effector SteC. An steC mutant strain was not defective for intracellular replication or attenuated for virulence in mice. However, the steC mutant was defective for SPI-2-dependent F-actin meshwork formation in host cells, although the vacuolar membranes surrounding mutant bacteria appeared to be normal. Expression of SteC in fibroblast cells following transfection caused extensive rearrangements of the F-actin cytoskeleton. Sequence analysis identified amino acid similarity between SteC and the human kinase Raf-1. A His-tagged SteC fusion protein had kinase activity in vitro and a point mutant lacking kinase activity was unable to induce F-actin rearrangements in vivo. We conclude that SPI-2-dependent F-actin meshwork formation depends on the kinase activity of SteC, which resembles more closely eukaryotic than prokaryotic kinases.
Poh
John
J
Department of Molecular Microbiology and Infection, Flowers Building, London SW7 2AZ, UK.
Odendall
Charlotte
C
Spanos
Ad
A
Boyle
Cliona
C
Liu
Mei
M
Freemont
Paul
P
Holden
David W
DW
D000199:Actins / Q000378:metabolism*; D000818:Animals; D002460:Cell Line; D004847:Epithelial Cells / Q000382:microbiology; D017353:Gene Deletion; D006801:Humans; D007425:Intracellular Membranes / Q000502:physiology; D008264:Macrophages / Q000382:microbiology; D051379:Mice; D016254:Mutagenesis, Insertional; D017354:Point Mutation; D011494:Protein Kinases / Q000235:genetics / Q000302:isolation & purification / Q000502:physiology*; D012486:Salmonella typhimurium / Q000201:enzymology* / Q000254:growth & development / Q000472:pathogenicity; D017421:Sequence Analysis; D017386:Sequence Homology, Amino Acid; D014617:Vacuoles / Q000382:microbiology; D014774:Virulence / Q000235:genetics
D000199:Actins; D011494:Protein Kinases; C526376:SteC protein, Salmonella typhimurium