Abstract
Erwinia carotovora subsp. atroseptica is an enterobacterial phytopathogen causing economically significant soft rot disease. Pathogenesis is mediated by multiple secreted virulence factors, many of which are secreted by the type II (Out) secretion system. DsbA catalyzes the introduction of disulfide bonds into periplasmic and secreted proteins. In this study, the extracellular proteome (secretome) of wild type E. carotovora subsp. atroseptica SCRI1043, and dsbA and out mutants, was analyzed by spectral counting mass spectrometry. This revealed that dsbA inactivation had a huge impact on the secretome and identified diverse DsbA- and Out-dependent secreted proteins, representing known, predicted, and novel candidate virulence factors. Further characterization of the dsbA mutant showed that secreted enzyme activities, motility, production of the quorum-sensing signal, and virulence were absent or substantially reduced. The impact of DsbA on secreted virulence factor production was mediated at multiple levels, including impacting on the Out secretion system and the virulence gene regulatory network. Transcriptome analyses revealed that the abundance of a broad, but defined, set of transcripts, including many virulence factors, was altered in the dsbA mutant, identifying a new virulence regulon responsive to extracytoplasmic conditions. In conclusion, DsbA plays a crucial, multifaceted role in the pathogenesis of E. carotovora subsp. atroseptica.
Highlights
The soft rot erwinias are Gram-negative, enteric bacteria and economically important plant pathogens
Their primary virulence characteristic is the coordinated production of large amounts of multiple secreted plant cell wall-degrading enzymes (PCWDEs),3 which leads to the breakdown of plant tissue and soft rot disease [2]
We found that DsbA is required for the proper production of almost all secreted virulence factors in E. carotovora subsp. atroseptica SCRI1043 and that its effect on virulence factor production is mediated at multiple levels, from transcript to protein secretion
Summary
The soft rot erwinias are Gram-negative, enteric bacteria and economically important plant pathogens. Bution of the former effect seems likely as, in general, the out Spectral Counting Mass Spectrometry Reveals Multiple DsbA- mutant (missing some but not all of the major secreted prodependent and Out-dependent Proteins in the Secretome of E. teins) showed an intermediate level of cellular proteins between carotovora subsp.
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