Characterization Of Key Domains For Avrrxol An Effector To Suppress Nonhost Resistance And To Trigger Rxol-mediated Resistance

Bacterial blight and bacterial leaf streak, caused by Xanthomonas oryzae pv. oryzae (Xoo)and X. oryzae pv. oryzicola (Xoc) respectively, are two most important bacterial diseases onrice. Although they have more than90%similarity on the genomic DNA, the infectioncharacteristics of Xoo and Xoc are very different: Xoo is the plant vascular-located pathogenand Xoc mainly colonizes the intercellular spaces of mesophyll cells. At present, more than30major resistance genes have been obtained for Xoo, but no one for Xoc has been identified inrice except Rxo1gene in maize, which was found to induce HR by recognizing cognateAvrRxo1. Rxo1was also found to resist Xoc after mobolised into rice. The application ofRxo1gives us a good opportunity to resist Xoc, however, the resistance resources are alsolargely limited. Meanwhile, Makino et al (2006) found that Xoc could suppress the plantdefense mediated by the recognition between AvrXa10and Xa10, which suggests anothercertain effectors can play a suppressor role in Xoc.In this study, by using a nonhost imcompatible system between PXO99a and Nicotianabenthaniana on which a fast HR can be triggered, we screened the PXO99a library habouringgenomic DNA of Xoc strain RS105. It is amazing that the clone containing avrRxo1wasfound to suppress the HR induced by PXO99a on N. benthaniana (NB). Then five avrRxo1alleles were cloned from different strains of Xoc. Their sequences show to cause diversity atthe amino acid level derived from nucleotide substitutions ranging from8to11. But they areidentified to habor the two certain functions: be recognized by Rxo1to trigger HR on B73,and to suppress PXO99a-mediated HR on NB. To detect the main domains controling bothfunctions, a series of truncated peptides of this effector were conducted from bothNH2-terminus and carboxy-termius. The results suggested that C-terminus was very importantfor AvrRxo1. Even the last amino acid residue deletion will result in lossing the suppressionability with AvrRxo1, the ability of recognition with Rxo1will fail from deletion of the lastthree amino acid residues. In addition, the putative ATPase site and nuclear location site (NLS)were also identified to be required for both functions as well as AvrRxo1C-terminus.The results of infection on rices by PXO99a carrying all the truncated peptides andimportant point mutants of AvrRxo1made us surprised that AvrRxo1suppresses thepathogenicity of PXO99a on rice varieties. Detection of bacterial growth on resistant andsusceptible cultivars made clear that amount of PXO99a carrying AvrRxo1is much less thanwild strain. Coincidentally, recognition domains with Rxo1are consistent with thepathogenicity suppressor’s domains, which suggests AvrRxo1probably stimulates rice resistance mediated by a Rxo1-like resistance protein. In the other words, AvrRxo1was alsoregarded as a “defeated” virulence factor which promoted the rice to form a specificresistance mechnism during the evolutionary process. Further evidence was provided bytrangenic rice. Overexpressing avrRxo1improves resistance to RS105and PXO99a in rice.The lesion lengths resulted from RS105infection on positive transgenic lines are shortenedranging from0.5to0.7cm and approximately3cm from PXO99a. Moreover, according toqRT-PCR test, the expression of resistance related genes PR1a and PR10was upregulated intransgenic plants. Together we concluded that AvrRxo1was a defeated pathogen virulencefactor which can elicit a non-Rxo1triggered resistance on rice.In conclusion, we have characterised the new function of AvrRxo1that it can suppressthe nonhost resistance mediated by interaction between PXO99a and NB, and identified somekey domains required for its function. More interestingly, we found that this effector cantrigger a Rxo1-like plant defense in rice. These results will help to extensive application oncrop disease improvement in the future.