鑑定與研究青枯病菌效應蛋白 RipBJ 之植物交互作用蛋白質

Abstract

Ralstonia solanacearum (Rs) causes bacterial wilt (BW), a vascular disease leading to severe global crop losses. However, information about how virulence-related factors function is still limited. Our previous studies showed that R. solanacearum effector RipBJ is present in low/medium-virulence strains and it plays a crucial negative role in virulence. RipBJ expression in tomato and Nicotiana benthamiana enhances plant defense against distinct important pathogens by modulating multiple plant defense signaling pathways. Furthermore, tomato NADPH oxidase SlWfi1 interacts with RipBJ, and plays a positive role in RipBJ-triggered effects on plants. This study aimed to elucidate the mechanism underlying the RipBJ-triggered defense by studying its interacting proteins in plants. Bimolecular fluorescence complementation (BiFC) assays showed that the interacting domains of RipBJ and SlWfi1 are 1-47 aa and 1-233 aa, respectively. Overexpression of SlWfi1 in N. benthamiana promotes RipBJ-induced plant cell death. Silencing of the homolog of SlWfi1 in N. benthamiana, NbRbohB, might affect the accumulation of RipBJ in planta. In vitro NADPH oxidase assay showed that RipBJ1-47 is sufficient to prolong the enzymatic activity of SlWfi1. Expression of SlWfi1 was differentially expressed in tomato root and stem-base after R. solanacearum infection. Furthermore, SlHCF164, SlCYP1 and SlCAT2 are candidate interacting proteins of RipBJ identified by Co-IP-LC-MS/MS. BiFC assays revealed these proteins may interact with RipBJ. Overexpression of SlHCF164 in N. benthamiana promoted RipBJ-induced plant cell death, and silencing of the homolog of SlWfi1 in N. benthamiana, NbCysP6, reduced RipBJ-induced plant cell death. Although the authentic interactions between RipBJ and these proteins remains to be determined further, SlHCF164 and NbCysP6 have a positive role in RipBJ-induced plant cell death. These results are expected to shed light on mechanisms involved in R. solanacearum virulence and plant defense.