A Sec-Dependent Secretory Protein Of The Huanglongbing-Associated Pathogen Suppresses Hypersensitive Cell Death In Nicotiana Benthamiana

"Candidatus Liberibacter asiaticus" (CLas) is a phloem-restricted Gram-negative bacterium that is the causal agent of citrus huanglongbing (HLB). In this study, we identified a CLas-encoded Sec-dependent secretory protein CLIBASIA_04405 that could contribute to the pathogenicity of this bacterium. The gene expression level of CLIBASIA_04405 was significantly higher in citrus than in psyllids. Transient overexpression of the mature CLIBASIA_04405 protein (m4405) in Nicotiana benthamiana leaves could suppress hypersensitive response (HR)-based cell death and H2O2 accumulation triggered by the mouse BAX and the Phytophthora infestans INF1. An alanine-substitution mutagenesis assay revealed the essential of amino acid clusters EKR45-47 and DE64-65 in cell death suppression. Challenge inoculation of the transgenic N. benthamiana-expressing m4405 with Pseudomonas syringae DC3000 Delta hopQ1-1 demonstrated the greatly reduced bacterial proliferation. Remarkably, transcriptome profiling and RT-qPCR analysis disclosed that the gene expression of six small heat shock proteins (sHSPs), a set of plant defense regulators, were significantly elevated in the transgenic m4405 lines compared with those in wild-type N. benthamiana. In addition, the transgenic m4405 lines displayed phenotypes of dwarfism and leaf deformation. Altogether, these data indicated that m4405 was a virulence factor of CLas.