The global transcription factor Clp exerts positive regulatory effects in the walnut bacterial black spot pathogen, Xanthomonas arboricola pv. juglandis

Walnut blight caused by the bacterium Xanthomonas arboricola pv. juglandis (Xaj) is one of the most common diseases of walnut (Juglans spp.), resulting in serious yield decline and significant economic losses. Crp-like protein (Clp) is an important global regulatory transcription factor in bacteria. In this study, we sought to elucidate the role of Clp in the pathogenicity of Xaj strain DW3F3 and the associated regulatory mechanism. The results indicated that clp gene deficiency significantly reduced the pathogenicity of Xaj DW3F3 in walnut without affecting the growth of the bacterium. We found that Clp positively regulates biofilm formation, extracellular polysaccharide production, exoenzyme secretion, and motility of Xaj, which was consistent with the transcript levels of virulence factor-encoding genes. However, overexpression of clp does not enhance the expression of all virulence genes, it may inhibit the expression of a part of virulence factor-related genes. EMSA assay further showed that Clp specifically binds to the promoters of these genes and regulates their expression, and CD spectra test certified that the ligand of Clp was c-di-GMP. Our findings contribute to the in-depth understanding of the pathogenic mechanism of Xaj and highlight the potential of Clp as a drug target for the development of agents to prevent and control walnut diseases.