The dual role of Xanthomonas effector XccXynB in plant cell wall deconstruction and immunity activation.

The GH10 endoxylanase XccXynB from Xanthomonas campestris pv. campestris (Xcc) functions as a PAMP, triggering PTI responses and enhancing disease resistance in plants, in addition to its polysaccharide degradation capability. Plant pathogens of the Xanthomonas genus are capable of infecting a wide array of economically important plant species. Among their virulence factors, the GH10 family xylanase, XynB, is typically responsible for the predominant extracellular xylanase activity. However, the detailed biochemical characteristics of this enzyme in most Xanthomonas pathogens remain insufficiently understood. In this study, we conducted a comprehensive characterization of the enzymatic and immunogenic properties of XynB from Xanthomonas campestris pv. campestris (Xcc). We found that XccXynB functions as an endoxylanase with a Km of 16.93 ± 4.93 (mg/mL) and Kcat of 1.42 ± 0.17 (min⁻1), primarily producing xylooligosaccharides (XOSs) ranging from xylobiose (X2) to xylohexaose (X6). Site-directed mutagenesis confirmed that residues E138 and E254 are essential for its catalytic activity. Furthermore, we found that XccXynB can act as a pathogen-associated molecular pattern (PAMP) in the model plants N. benthamiana and A. thaliana, triggering immune responses such as reactive oxygen species (ROS) burst comparable to the elicitor Flg22, without inducing cell death. Heterologous expression of XccXynB in A. thaliana led to constitutive immune activation and significantly enhanced disease resistance, with the in planta bacterial population of Pst DC3000 being reduced to ~ 80%. Taken together, these findings could provide a rationale for developing novel strategies against Xanthomonas diseases by targeting the conserved xylanase function or utilizing the protein as an immunogen.