Comparative proteomic analysis of Xanthomonas citri ssp citri periplasmic proteins reveals changes in cellular envelope metabolism during in vitro pathogenicity induction

Citrus canker is a plant disease caused by gram-negative bacteria from the genus Xanthomonas, and the most virulent species is Xanthomonas citri subsp.citri (XAC), which attacks a wide range of citrus hosts. Differential proteomic analysis of periplasm-enriched fraction was performed for XAC cells grown in pathogenicity-inducing (XAM-M) and pathogenicity non-inducing (NB) media, using two-dimensional electrophoresis combined with liquid chromatography-tandem mass spectrometry. Among the 40 proteins identified, transglycosylase was detected in a highly abundant spot in XAC cells grown under inducing condition. Additional upregulated proteins related to cellular envelope metabolism included glucose-1-phosphate thymidylyltransferase, dTDP-4-dehydrorhamnose 3,5-epimerase, and peptidyl-prolyl cis-trans isomerase. Phosphoglucomutase and superoxide dismutase proteins, known to be involved in pathogenicity in other Xanthomonas species or organisms, were also detected. Western blot and quantitative real-time PCR analyses for transglycosylase and superoxide dismutase confirmed that these proteins are upregulated under inducing condition, consistent with the proteomic results. Multiple spots for the 60 kDa chaperonin and glyceraldehyde-3-phosphate dehydrogenase were identified, suggesting the presence of post-translational modifications. We propose that substantial alterations in the cellular envelope metabolism occur during the XAC infectious process, which are related to several aspects, from defense against reactive oxygen species to exopolysaccharide synthesis. Our results provide new candidates of virulence-related proteins, whose abundance correlates with the induction of pathogenicity and virulence genes like hrpD6, hrpG, hrpB7, hpa1, and hrpX. The results present new potential targets against XAC to be investigated in further functional studies. This article is protected by copyright. All rights reserved.