Pathogenic serovars of Salmonella utilise two type-three secretion systems to deliver distinct cohorts of effector proteins into host cells during infection. These effector proteins interact with specific human proteins to subvert normal cellular processes, thus impairing the ability of host cells to respond to the invading bacteria. Our research focuses on the SseK family of Salmonella effectors, which have unknown function and play an unknown role in Salmonella infection. This effector family shows strong sequence homology to the enteropathogenic E. coli effector protein NleB1, which glycosylates several host signalling proteins. Therefore, we predict the SseK effectors are also glycosyltransferases, and the aims of our work are to identify the host substrates of these effectors and to characterise the post-translational modification catalysed by these effectors.
Our work has established that SseK1 and SseK3 function as glycosyltransferases, and catalyse a rare post-translational modification in which the sugar N-acetylglucosamine is transferred to arginine residues of host proteins. By immunofluorescence, SseK1 and its glycosylated substrate were found to be localised throughout the host cell, while SseK3 and its substrate were localised to the host Golgi. Next, we developed an approach combining immunoprecipitation and mass spectrometry to identify the glycosylated host substrates of these effector proteins. SseK1 was found to glycosylate arginine residues of the host protein TRADD, which functions as an adaptor molecule in both inflammatory and cell death signalling pathways. Further, we found that SseK3 glycosylates several components of these same pathways, suggesting the function of this effector family may be to enact a blockade of multiple host signalling pathways. Currently, we are exploring the contribution of these post-translational modifications to inflammatory signalling and cell death during Salmonella infection. Ultimately, this work suggests that Salmonella engages in a broad program of interference with host cell signalling during infection.