Abstract
The hallmark of enteropathogenic Escherichia coli (EPEC) infection is the formation of actin-rich pedestal-like structures, which are generated following phosphorylation of the bacterial effector Tir by cellular Src and Abl family tyrosine kinases. This leads to recruitment of the Nck–WIP–N-WASP complex that triggers Arp2/3-dependent actin polymerization in the host cell. The same phosphorylation-mediated signalling network is also assembled downstream of the Vaccinia virus protein A36 and the phagocytic Fc-gamma receptor FcγRIIa. Here we report that the EPEC type-III secretion system effector EspJ inhibits autophosphorylation of Src and phosphorylation of the Src substrates Tir and FcγRIIa. Consistent with this, EspJ inhibits actin polymerization downstream of EPEC, Vaccinia virus and opsonized red blood cells. We identify EspJ as a unique adenosine diphosphate (ADP) ribosyltransferase that directly inhibits Src kinase by simultaneous amidation and ADP ribosylation of the conserved kinase-domain residue, Src E310, resulting in glutamine-ADP ribose.
Highlights
The hallmark of enteropathogenic Escherichia coli (EPEC) infection is the formation of actinrich pedestal-like structures, which are generated following phosphorylation of the bacterial effector Tir by cellular Src and Abl family tyrosine kinases
We previously reported that opsono-phagocytosis via FcgRIIa could be inhibited by the enteropathogenic Escherichia coli (EPEC) and enterohaemorrhagic E. coli (EHEC) effector EspJ through an unknown mechanism[14]
While the positions of some 1H–15N heteronuclear single quantum coherence (HSQC) peaks for EspJ28-217R79A and EspJ28-217D187A were altered relative to the EspJ28-217 spectra, the excellent spectra dispersion indicates that the mutations have not affected the overall protein structure
Summary
The hallmark of enteropathogenic Escherichia coli (EPEC) infection is the formation of actinrich pedestal-like structures, which are generated following phosphorylation of the bacterial effector Tir by cellular Src and Abl family tyrosine kinases. This leads to recruitment of the Nck–WIP–N-WASP complex that triggers Arp2/3-dependent actin polymerization in the host cell. We show that EspJ inhibits opsono-phagocytosis through inactivation of Src, disrupting phosphorylation of the FcgRIIa. EspJ inhibits Src activity by a unique post-translational modification mechanism involving amidation and adenosine diphosphate (ADP) ribosylation of a key kinase-domain residue, which is conserved across the protein kinase superfamily
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