Molecular Study Of Genes Associated With Systemic Aquired Resistance In Rice

NPR1 (non-expresser of pathogenesis related genes 1) is the master regulator of salicylic acid-mediated systemic acquired resistance. Over-expression of Arabidopsis NPR1 and rice NH1 (NPR1 homolog1)/OsNPR1 in rice results in enhanced resistance. While there are four rice NPR1-paralogs in the rice genome, none have been demonstrated to function in disease resistance. To study rice NPR1 paralog 3, we introduced constructs into rice and tested for effects on resistance to infection by Xanthomonas oryzae pv. oryzae (Xoo), the causal agent of bacterial blight. While over-expression of NH3 using the maize ubiquitin-1 promoter failed to enhance resistance, introduction of an extra copy of NH3 driven by its own promoter (nNT-NH3) resulted in clear, enhanced resistance. Progeny analysis confirms that the enhanced resistance phenotype, measured by Xoo-induced lesion length, is associated with the NH3 transgene. Bacterial growth curve analysis indicates that bacterial populations levels are reduced 10-fold in nNT-NH3 lines compared to control rice lines. The transgenic plants exhibit higher sensitivity to BTH (benzothiadiazole) and INA (2,6-dichloroisonicotinic acid) treatment as measured by increased cell death. Expression analysis of pathogenesis-related (PR) genes showed that nNT-NH3 plants display greatly enhanced induction of PR genes only after treatment with BTH. Our study demonstrates an alternative method to employ a regulatory protein to enhance plant defense. This approach avoids using undesirable constitutive, high-level expression and may prove to be more practical for engineering resistance.NPR1 binds to TGA transcription factors and functions as a transcriptional co-activator after activation by inducers. We have established a rice transient protoplast cell system to study the interaction between NH1/OsNPR1 and NRR proteins. This transient system demonstrates the transcriptional activation activity of NH1 and the ability of NRR to repress NH1-mediated activation. Three NRR homologs (RH1, RH2, and RH3) are identified in rice. RH1, RH2, and RH3 also function to repress NH1-mediated transcriptional activation. NRR, RH1, RH2, and RH3 share sequence similarity in a second region beyond the previously identified NPR1-interacting domain. This second region is required for strong interaction with NH1. Double point mutation W66A/F70A in this NH1-interacting domain severely reduced interaction with NH1, while single point mutation W66A modestly reduced the interaction. Mutation W66A/F70A also greatly reduced the ability to repress NH1-mediated activation and W66A lost most of its ability to repress. These results demonstrate that the ability of NRR to repress NH1-mediated transcriptional activation is tightly correlated with its ability to bind to NH1. Furthermore, this domain in NRR and NRR-like proteins constitutes a novel bipartite (NPR1-interacting +NH1-interacting sequences) protein-protein interacting domain. Importantly, this bipartite domain is widely present plant species, from cereals to castor bean plants, to poplar trees, but absent in Arabidopsis and tobacco.We used bimolecular fluorescence complementation method and transient protoplast cell system to study the protein protein interaction in vito between NH1 family, NRR family, and TGA family members. The results of bimolecular fluorescence complementation assay indicated NH1 and its homologue can bind to each other, so as NRR and its homologue. NH1 family members can interact with NRR family and TGA family members. NRR family members can't interact directly with TGA family members.However, upon with NH1 family members, they can interact with each other. The transient assay results demonstrated NRR family members can repress transcriptional activity from NH1 but activate from NH4 and NH5. And NH1 can activate transcriptional activity for all the TGA family members except TH11.