OsWRKY114 Negatively Regulates Drought Tolerance by Restricting Stomatal Closure in Rice.

The WRKY family of transcription factors plays a pivotal role in plant responses to biotic and abiotic stress. The WRKY Group III transcription factor OsWRKY114 is a positive regulator of innate immunity against Xanthomonas oryzae pv. oryzae; however, its role in abiotic stress responses is largely unknown. In this study, we showed that the abundant OsWRKY114 transcripts present in transgenic rice plants are reduced under drought conditions. The overexpression of OsWRKY114 significantly increased drought sensitivity in rice, which resulted in a lower survival rate after drought stress. Moreover, we showed that stomatal closure, which is a strategy to save water under drought, is restricted in OsWRKY114-overexpressing plants compared with wild-type plants. The expression levels of PYR/PYL/RCAR genes, such as OsPYL2 and OsPYL10 that confer drought tolerance through stomatal closure, were also markedly lower in the OsWRKY114-overexpressing plants. Taken together, these results suggest that OsWRKY114 negatively regulates plant tolerance to drought stress via inhibition of stomatal closure, which would otherwise prevent water loss in rice.

Identification of the Capsicum baccatum NLR Protein CbAR9 Conferring Disease Resistance to Anthracnose.

Anthracnose is caused by Colletotrichum species and is one of the most virulent fungal diseases affecting chili pepper (Capsicum) yield globally. However, the noble genes conferring resistance to Colletotrichum species remain largely elusive. In this study, we identified CbAR9 as the causal locus underlying the large effect quantitative trait locus CcR9 from the anthracnose-resistant chili pepper variety PBC80. CbAR9 encodes a nucleotide-binding and leucine-rich repeat (NLR) protein related to defense-associated NLRs in several other plant species. CbAR9 transcript levels were induced dramatically after Colletotrichum capsici infection. To explore the biological function, we generated transgenic Nicotiana benthamiana lines overexpressing CbAR9, which showed enhanced resistance to C. capsici relative to wild-type plants. Transcript levels of pathogenesis-related (PR) genes increased markedly in CbAR9-overexpressing N. benthamiana plants. Moreover, resistance to anthracnose and transcript levels of PR1 and PR2 were markedly reduced in CbAR9-silenced chili pepper fruits after C. capsici infection. Our results revealed that CbAR9 contributes to innate immunity against C. capsici.

The Capsicum baccatum-Specific Truncated NLR Protein CbCN Enhances the Innate Immunity against Colletotrichum acutatum.

Chili pepper (Capsicumannuum) is an important fruit and spice used globally, but its yield is seriously threatened by anthracnose. Capsicum baccatum is particularly valuable as it carries advantageous disease resistance genes. However, most of the genes remain to be identified. In this study, we identified the C. baccatum-specific gene CbCN, which encodes a truncated nucleotide-binding and leucine-rich repeat protein in the anthracnose resistant chili pepper variety PBC80. The transcription of CbCN was greater in PBC80 than it was in the susceptible variety An-S after Colletotrichum acutatum inoculation. In order to investigate the biological function of CbCN, we generated transgenic tobacco lines constitutively expressing CbCN. Notably, CbCN-overexpressing transgenic plants exhibited enhanced resistance to C. acutatum compared to wild-type plants. Moreover, the expression of pathogenesis-related (PR) genes was remarkably increased in a CbCN-overexpressing tobacco plants. In order to confirm these results in chili pepper, we silenced the CbCN gene using the virus-induced gene silencing system. The anthracnose resistance and expressions of PR1, PR2, and NPR1 were significantly reduced in CbCN-silenced chili peppers after C. acutatum inoculations. These results indicate that CbCN enhances the innate immunity against anthracnose caused by C. acutatum by regulating defense response genes.

Identification of a novel NPR1 homolog gene, OsNH5N16, which contributes to broad-spectrum resistance in rice.

Over half of the earth's population consumes rice as the primary food crop for dietary calories. However, severe loss of rice yield occurs due to bacterial blight caused by Xanthomonas oryzae pv. oryzae (Xoo) and bakanae disease caused by Fusarium fujikuroi (F. fujikuroi). Therefore, broad-spectrum resistance (BSR) to these pathogens is essential for rice cultivation. The Nonexpressor of Pathogenesis-Related Genes1 (NPR1), which is related to the signal molecule salicylic acid (SA) and the expression of pathogenesis-related (PR) genes, is a key regulator of systemic acquired resistance (SAR). Although five NPR1 homologs (NHs) have been identified in rice thus far, their cellular and biological functions remain largely unexplored. In this study, we identified a novel rice NH gene from Oryza sativa L. cv. Dongjin. The genetic variation of single nucleotide polymorphisms in OsNH5 caused a single amino acid substitution of asparagine for serine at residue 16. OsNH5N16 was mainly located in the nucleus, and its transcription was induced by Xoo. We generated transgenic rice lines constitutively expressing OsNH5N16 to investigate its function. Plants that overexpressed OsNH5N16 displayed enhanced BSR to Xoo and F. fujikuroi compared with wild varieties, and the transcription of PR genes such as OsPR1, GLUC, and CHIT2 was considerably upregulated. Moreover, we revealed that SA increases the transcription of OsNH5N16 and the promoter activity of OsPR1 regulated by OsNH5N16. These results showed that OsNH5N16 enhances BSR by regulating the expression of PR genes related to SAR and it is controlled by SA at the transcriptional and post-translational levels. This is the first report on the innate immune response conferring BSR associated with NH5.

Rice glutaredoxin GRXS15 confers broad-spectrum resistance to Xanthomonas oryzae pv. oryzae and Fusarium fujikuroi.

Rice is a particularly widely consumed food crop globally, but its yield is seriously damaged by bacterial blight due to Xanthomonas oryzae pv. oryzae (Xoo) and bakanae disease due to Fusarium fujikuroi (F. fujikuroi). However, broad-spectrum resistance (BSR) to both Xoo and F. fujikuroi remains largely elusive. In this study, we showed that rice monothiol glutaredoxin GRXS15 localizes in mitochondria and the nucleus, and its transcription is induced by Xoo. Transgenic rice lines constitutively expressing OsGRXS15 showed enhanced disease resistance to Xoo and F. fujikuroi, while CRISPR/Cas9-based knockout mutants showed reduced resistance compared with the wild-type plants. The transcription of pathogenesis-related (PR) genes was significantly induced in OsGRXS15-expressing plants. The rice transcription factor OsWRKY65 was identified as a binding partner, and it directly interacted with OsGRXS15 in the nucleus. Moreover, we revealed that the interaction of OsGRXS15 and OsWRKY65 results in the upregulation of OsPR1. These results suggested that OsGRXS15 interacts with transcription factors, and it confers BSR through regulating the expression of genes related to pathogen response. This is the first report on the nuclear function associated with the monothiol glutaredoxin GRXS15.