A Glycoside Hydrolase Family 99-Like Domain-Containing Protein Modifies Outer Membrane Proteins to Maintain Xanthomonas Pathogenicity and Viability in Stressful Environments

Protein glycosylation is an essential process that plays an important role in proteome stability, protein structure, and protein function modulation in eukaryotes. However, in bacteria, especially plant pathogenic bacteria, similar studies are lacking. Here, we investigated the relationship between protein glycosylation and pathogenicity by using Xanthomonas oryzae pv. oryzae, the causal agent of bacterial leaf blight in rice, as a well-defined example. In our previous work, we identified a virulence-related hypothetical protein, PXO_03177, but how this protein regulates X. oryzae pv. oryzae virulence has remained unclear. BLAST analysis showed that most homologous proteins of PXO_03177 are glycoside hydrolase family 99-like domain-containing proteins. In the current study, we found that the outer membrane integrity of Delta PXO_03177 appeared to be disrupted. Extracting the outer membrane proteins (OMPs) and performing comparative proteomics and sodium dodecyl sulphate-polyacrylamide gel electrophoresis gel staining analyses revealed that PXO_03177 deletion altered the protein levels of 13 OMPs. Western blot analyses showed that the protein level and glycosylation modification of PXO_02523, a related OmpA family-like protein, was changed in the Delta PXO_03177 mutant background strain. Additionally, the interaction between PXO_03177 and PXO_02523 was confirmed by coimmunoprecipitation. Both PXO_03177 and PXO_02523 play important roles in regulating pathogen virulence and viability in stressful environments. This work provides the first evidence that protein glycosylation is necessary for the virulence of plant pathogenic bacteria.

Automated summary

This study investigated the relationship between protein glycosylation and pathogenicity using Xanthomonas oryzae pv. oryzae as a model. The deletion of PXO_03177 altered the protein levels of outer membrane proteins and affected outer membrane integrity. The study provides evidence that protein glycosylation is essential for the virulence of plant pathogenic bacteria.