Broad spectrum regulation of non receptor tyrosine kinases by the bacterial ADP ribosyltransferase EspJ

Abstract

Tyrosine phosphorylation is key for signal transduction from exogenous stimuli, including the defence against pathogens . Conversely, p athogens can s ubvert protein phosphorylation to control host immune response s and facilitate invasion and dissemination . The bacterial 23 effector s EspJ and SeoC are injected into host cells though a type III secretion system by entero pathogenic and enterohaemorrhagic Escherichia coli (EPEC and EHEC) , Citrobacter rodentium and Salmonella enterica where they inhibit Src kinase by coupled amidation and ADP - ribosylation . C. rodentium , which is u sed to model EPEC and EHEC infections in human, is a mouse pathogen triggering colonic crypt hyperplasia (CCH) and colitis. Enumeration of bacterial shedding and CCH confirmed that EspJ affects neither tolerance nor resistance to infection. However, compar ing the proteomes of intestinal epithelial cells isolated from mice infected with wildtype C. rodentium or C. rodentium encoding catalytically inactive EspJ revealed that EspJ - induced ADP - ribosylation regulates multiple non - receptor tyrosine kinases in vivo . Investi gati ng the substrate repertoire of EspJ revealed that in HeLa and A549 Src and Csk were significantly targeted; in polarised Caco2 cells EspJ targeted Src and Csk and the Src family kinase (SFK) Yes1, while in differentiated Thp1 EspJ modified Csk, the SFKs Hck and Lyn, the Tec family kinases Tec and Btk, and the adapter tyrosine kinase Syk. Furthermore, Abl (HeLa and Caco2) and Lyn (Caco2) were enriched specifically in the EspJ - containing samples. Biochemical assays revealed that EspJ , the onl y bacterial ADP - ribosyltransferase which targets mammalian kinases, control s immune responses and the Src/Csk signalling axis.