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      OsWRKY67 positively regulates blast and bacteria blight resistance by direct activation of PR genes in rice

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          Abstract

          Background

          WRKY proteins are one of the largest gene families and are well-known for their regulatory roles in many aspects of plant development, including plant response to both biotic and abiotic stresses. Although the roles of WRKY proteins in leaf blast resistance have been well-documented in rice, their functions in panicle blast, the most destructive type of blast disease, are still largely unknown.

          Results

          Here, we identified that the transcription of OsWRKY67 was strongly activated by leaf and panicle blast infection. OsWRKY67 is ubiquitously expressed and sub-localized in the nucleus. Rice plants overexpressing OsWRKY67 showed quantitatively enhanced resistance to leaf blast, panicle blast and bacterial blight. In contrast, silencing of OsWRKY67 increased the susceptibility to blast and bacterial blight diseases. RNA-seq analysis indicated that OsWRKY67 induces the transcription of a set of defense-related genes including the ones involved in the salicylic acid (SA)-dependent pathway. Consistent with this, the OsWRKY67-overexpressing plants accumulated higher amounts of endogenous SA, whereas lower endogenous SA levels were observed in OsWRKY67-silenced plants relative to wild-type Nipponbare plants before and after pathogen attack. Moreover, we also observed that OsWRKY67 directly binds to the promoters of PR1a and PR10 to activate their expression.

          Conclusions

          These results together suggest the positive role of OsWRKY67 in regulating rice responses to leaf blast, panicle blast and bacterial blight disease. Furthermore, conferring resistance to two major diseases makes it a good target of molecular breeding for crop improvement in rice.

          Electronic supplementary material

          The online version of this article (10.1186/s12870-018-1479-y) contains supplementary material, which is available to authorized users.

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          Most cited references28

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          A highly efficient rice green tissue protoplast system for transient gene expression and studying light/chloroplast-related processes

          Background Plant protoplasts, a proven physiological and versatile cell system, are widely used in high-throughput analysis and functional characterization of genes. Green protoplasts have been successfully used in investigations of plant signal transduction pathways related to hormones, metabolites and environmental challenges. In rice, protoplasts are commonly prepared from suspension cultured cells or etiolated seedlings, but only a few studies have explored the use of protoplasts from rice green tissue. Results Here, we report a simplified method for isolating protoplasts from normally cultivated young rice green tissue without the need for unnecessary chemicals and a vacuum device. Transfections of the generated protoplasts with plasmids of a wide range of sizes (4.5-13 kb) and co-transfections with multiple plasmids achieved impressively high efficiencies and allowed evaluations by 1) protein immunoblotting analysis, 2) subcellular localization assays, and 3) protein-protein interaction analysis by bimolecular fluorescence complementation (BiFC) and firefly luciferase complementation (FLC). Importantly, the rice green tissue protoplasts were photosynthetically active and sensitive to the retrograde plastid signaling inducer norflurazon (NF). Transient expression of the GFP-tagged light-related transcription factor OsGLK1 markedly upregulated transcript levels of the endogeneous photosynthetic genes OsLhcb1, OsLhcp, GADPH and RbcS, which were reduced to some extent by NF treatment in the rice green tissue protoplasts. Conclusions We show here a simplified and highly efficient transient gene expression system using photosynthetically active rice green tissue protoplasts and its broad applications in protein immunoblot, localization and protein-protein interaction assays. These rice green tissue protoplasts will be particularly useful in studies of light/chloroplast-related processes.
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            Arabidopsis WRKY33 transcription factor is required for resistance to necrotrophic fungal pathogens.

            Plant WRKY transcription factors are key regulatory components of plant responses to microbial infection. In addition to regulating the expression of defense-related genes, WRKY transcription factors have also been shown to regulate cross-talk between jasmonate- and salicylate-regulated disease response pathways. The two pathways mediate resistance against different types of microbial pathogens, and there are numerous reports of antagonistic interactions between them. Here we show that mutations of the Arabidopsis WRKY33 gene encoding a WRKY transcription factor cause enhanced susceptibility to the necrotrophic fungal pathogens Botrytis cinerea and Alternaria brassicicola concomitant with reduced expression of the jasmonate-regulated plant defensin PDF1.2 gene. Ectopic over-expression of WRKY33, on the other hand, increases resistance to the two necrotrophic fungal pathogens. The wrky33 mutants do not show altered responses to a virulent strain of the bacterial pathogen Pseudomonas syringae, although the ectopic expression of WRKY33 results in enhanced susceptibility to this pathogen. The susceptibility of WRKY33-over-expressing plants to P. syringae is associated with reduced expression of the salicylate-regulated PR-1 gene. The WRKY33 transcript is induced in response to pathogen infection, or treatment with salicylate or the paraquat herbicide that generates activated oxygen species in exposed cells. WRKY33 is localized to the nucleus of plant cells and recognizes DNA molecules containing the TTGACC W-box sequence. Together, these results indicate that pathogen-induced WRKY33 is an important transcription factor that regulates the antagonistic relationship between defense pathways mediating responses to P. syringae and necrotrophic pathogens.
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              Rice WRKY45 plays a crucial role in benzothiadiazole-inducible blast resistance.

              Benzothiadiazole (BTH) is a so-called plant activator and protects plants from diseases by activating the salicylic acid (SA) signaling pathway. By microarray screening, we identified BTH- and SA-inducible WRKY transcription factor (TF) genes that were upregulated within 3 h after BTH treatment. Overexpression of one of them, WRKY45, in rice (Oryza sativa) markedly enhanced resistance to rice blast fungus. RNA interference-mediated knockdown of WRKY45 compromised BTH-inducible resistance to blast disease, indicating that it is essential for BTH-induced defense responses. In a transient expression system, WRKY45 activated reporter gene transcription through W-boxes. Epistasis analysis suggested that WRKY45 acts in the SA signaling pathway independently of NH1, a rice ortholog of Arabidopsis thaliana NPR1, which distinguishes WRKY45 from known Arabidopsis WRKY TFs. Two defense-related genes, encoding a glutathione S-transferase and a cytochrome P450, were found to be regulated downstream of WRKY45 but were not regulated by NH1, consistent with the apparent independence of the WRKY45- and NH1-dependent pathways. Defense gene expression in WRKY45-overexpressed rice plants varied with growth conditions, suggesting that some environmental factor(s) acts downstream of WRKY45 transcription. We propose a role for WRKY45 in BTH-induced and SA-mediated defense signaling in rice and its potential utility in improving disease resistance of rice, an importance food resource worldwide.
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                Author and article information

                Contributors
                295426650@qq.com
                382600311@qq.com
                shijuan@agrogene.ac.cn
                278208621@qq.com
                y60382035@tom.com
                Jingfangnihao@163.com
                szhanggz@tom.com
                otest@163.com
                yang2004009@163.com
                maoxingxue@qq.com
                gzzhuxy@tom.com
                lbgz1009@163.com
                Journal
                BMC Plant Biol
                BMC Plant Biol
                BMC Plant Biology
                BioMed Central (London )
                1471-2229
                26 October 2018
                26 October 2018
                2018
                : 18
                : 257
                Affiliations
                [1 ]GRID grid.488205.3, Guangdong Key Laboratory of New Technology in Rice Breeding, Rice Research Institute, Guangdong Academy of Agricultural Sciences, ; Guangzhou, 510640 China
                [2 ]ISNI 0000 0004 1790 4137, GRID grid.35155.37, School of Plant Science and Technology, , Huazhong Agricultural University, ; Wuhan, 430070 China
                [3 ]ISNI 0000 0001 0561 6611, GRID grid.135769.f, Agro-biological Gene Research Center, Guangdong Academy of Agricultural Sciences, ; Guangzhou, 510640 China
                [4 ]ISNI 0000 0001 0561 6611, GRID grid.135769.f, Guangdong Key Laboratory of New Technology in Plant Protection, Plant Protection Research Institute, Guangdong Academy of Agricultural Sciences, ; Guangzhou, 510640 China
                Article
                1479
                10.1186/s12870-018-1479-y
                6204034
                30367631
                042cd6bb-02f9-4eb7-af9b-de511865f950
                © The Author(s). 2018

                Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License ( http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver ( http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.

                History
                : 20 January 2018
                : 10 October 2018
                Funding
                Funded by: NSFC-IRRI project
                Award ID: 31461143019
                Award Recipient :
                Funded by: Science and Technology Project of Guangdong Province
                Award ID: 2015B020231002
                Award ID: 2016A050502030
                Award ID: 2014B070706013
                Award Recipient :
                Funded by: the Natural Science Foundation of Guangdong Province
                Award ID: 2014A030310489
                Award Recipient :
                Funded by: the Common Technical Innovation Team of Guangdong Province on Agricultural Seed Industry
                Award ID: 2017LM2148
                Award Recipient :
                Categories
                Research Article
                Custom metadata
                © The Author(s) 2018

                Plant science & Botany
                oswrky67,leaf blast,panicle blast,bacterial blight,salicylic acid
                Plant science & Botany
                oswrky67, leaf blast, panicle blast, bacterial blight, salicylic acid

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