Functional Analysis Of Receptor-like Protein Kinases On Plants Responses To N-acylhomoserine Lactones(AHLs)

Gram-negative bacteria can secrete N-acyl-homoserine lactone?AHLs?as signal molecules to communicate each other for coordinating the collective activities in population.Previous studies in our laboratory proved that AHLs are recognized by bacteria as well be perceived by plants resulting in changes of physiological processes in plant.However,little is known about the mechanism by which plant transduce AHL in plant cells.The membrane-associated receptor-like protein?RLK?can perceive the extracellular signals and transduce the signals into cells by action of the intracellular kinase domain.The aim of this study is to investigate the involvement of two RLK in plant response to bacterial AHLs.1.Involvement of leucine-rich repeats receptor-like kinase?LRR-RLK?and lectin receptor-like kinases?LecRLK?in regulation of plant primary root growth by 3OC6-HSL.Root elongation analysis revealed that treatment with 1?mol·L^-11 3OC6-HSL significantly promote the primary root growth of wild-type Arabidopsis.However,the stimulatory effects of 3OC6-HSL on root growth were diminished in lrr and rlk mutant plants and the lrr mutant root length was significantly lower than that of wild-type after treatment with3OC6-HSL.Real Time PCR results showed that 1?mol·L^-1 3OC6-HSL can significantly enhance the transcripts of LRR and RLK genes in wild-type Arabidopsis thaliana.These results indicate that LRR and RLK receptor protein kinases might be involved in3OC6-HSL-mediated primary roo growth of Arabidopsis.2.Involvement of LRR and RLK in regulation of plant disease resistance by3OC6-HSL.The results showed that preconditioning with 10?mol·L^-11 3OC8-HSL for 48h significantly reduced the colonization of pathogens Pseudomonas syringae DC3000 in the leaves of Arabidopsis.No increase was observed in leaves as the plants inoculated with P.syringae DC3000 were cultivated longer,indicating that 3OC8-HSL played an important role in plant resistance against pathogens DC3000 in wild-type Arabidopsis.However,the induction of resistance toward P.syringae DC3000 was no longer detected in lrr and rlk mutant after treatment with 3OC8-HSL,and the number of pathogen colonization was higher in leaves of mutant rlk than that of wild-type Arabidopsis.Real Time PCR results showed that 3OC8-HSL up-regulated the transcript of LRR and RLK gene.After infection of Arabidopsis pathogen DC3000,RLK gene expression was significantly up-regulated,but LRR expression did not change significantly.When treated with 3OC8-HSL before inoculation of DC3000,the expression of RLK gene was further increased while there was still no significant change in LRR expression.These data showed that LRR and RLK are responsible to 3OC8-HSL.RLK was involved in the progress of regulating the plant disease resistance by 3OC8-HSL.3.The prokaryotic expression vectors of LRR and RLK were constructed.LRR and RLK genes were linked with the prokaryotic expression vector pET-28a,constructing recombinant BL21?pET-28a-LRR,pET-28a-RLK?containing His-tag.The experiment laied the foundation for studying the receptor dynamics between LRR?RLK and AHLs.