The major outer sheath protein of Treponema denticola selectively inhibits Rac1 activation in murine neutrophils.
Treponema denticola major outer sheath protein (Msp) inhibits neutrophil chemotaxis in vitro, but key regulatory mechanisms have not been identified. Because the Rac small GTPases regulate directional migration in response to chemoattractants, the objective was to analyse the effects of Msp on formyl-methionyl-leucyl-phenylalanine (fMLP)-mediated neutrophil polarization and Rac activation in murine neutrophils. Msp pretreatment of neutrophils inhibited both polarization and chemotactic migration in response to fMLP. Activation of small GTPases was measured by p21 binding domain (PBD) pulldown assays, followed by Western analysis, using monoclonal anti-Rac1, anti-Rac2, anti-cdc42 and anti-RhoA antibodies. Enriched native Msp selectively inhibited fMLP-stimulated Rac1 activation in a concentration-dependent manner, but did not affect Rac2, cdc42 or RhoA activation. Murine neutrophils transfected with vectors expressing fluorescent probes PAK-PBD-YFP and PH-AKT-RFP were used to determine the effects of Msp on the localization of activated Rac and PI3 kinase products. Real-time confocal images showed that Msp inhibited the polarized accumulation of activated Rac and PI3-kinase products upon exposure to fMLP. The findings indicate that T. denticola Msp inhibition of neutrophil polarity may be due to the selective suppression of the Rac1 pathway.
Magalhães
Marco A O
MA
CIHR Group in Matrix Dynamics and Dental Research Institute, University of Toronto, Faculty of Dentistry, Toronto, Ontario, Canada.
Sun
Chun Xiang
CX
Glogauer
Michael
M
Ellen
Richard P
RP
D000818:Animals; D001426:Bacterial Proteins / Q000378:metabolism / Q000502:physiology*; D016764:Cell Polarity / Q000187:drug effects; D002633:Chemotaxis / Q000187:drug effects; D004789:Enzyme Activation; D051379:Mice; D009240:N-Formylmethionine Leucyl-Phenylalanine / Q000494:pharmacology; D009479:Neuropeptides / Q000378:metabolism*; D009504:Neutrophils / Q000187:drug effects / Q000378:metabolism*; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D018272:Porins / Q000378:metabolism / Q000502:physiology*; D048373:Treponema denticola / Q000378:metabolism*; D020744:rac GTP-Binding Proteins / Q000378:metabolism*; D020830:rac1 GTP-Binding Protein
D001426:Bacterial Proteins; D009240:N-Formylmethionine Leucyl-Phenylalanine; D009479:Neuropeptides; D019869:Phosphatidylinositol 3-Kinases; D018272:Porins; D020744:rac GTP-Binding Proteins; D020830:rac1 GTP-Binding Protein; C484602:Rac1 protein, mouse; C099361:major outer sheath protein, Treponema