Drought-induced protein 19(Di19) is a Cys2/His2 zinc-finger protein that functions in plant growth and development and in tolerance to abiotic stresses.Gm PUB21,an E3 ubiquitin ligase,negatively regulates drought and ...Drought-induced protein 19(Di19) is a Cys2/His2 zinc-finger protein that functions in plant growth and development and in tolerance to abiotic stresses.Gm PUB21,an E3 ubiquitin ligase,negatively regulates drought and salinity response in soybean.We identified potential interaction target proteins of Gm PUB21by yeast two-hybrid c DNA library screening,Gm Di19-5 as a candidate.Bimolecular fluorescence complementation and glutathionine-S-transferase pull-down assays confirmed the interaction between Gm Di19-5 and Gm PUB21.Gm Di19-5 was induced by Na Cl,drought,and abscisic acid(ABA) treatments.Gm Di19-5 was expressed in the cytoplasm and nucleus.Gm Di19-5 overexpression conferred hypersensitivity to drought and high salinity,whereas Gm Di19-5 silencing increased drought and salinity tolerance.Transcripts of ABA-and stress response-associated genes including Gm RAB18 and Gm DREB2A were downregulated in Gm Di19-5-overexpressing plants under drought and salinity stresses.ABA decreased the protein level of Gm Di19-5 in vivo,whereas Gm PUB21 increased the decrease of Gm Di19-5 after exogenous ABA application.The accumulation of Gm PUB21 was also inhibited by Gm Di19-5.We conclude that Gm PUB21 and Gm Di19-5 collaborate to regulate drought and salinity tolerance via an ABA-dependent pathway.展开更多
Soybean mosaic virus(SMV) is one of the most devastating viral pathogens of soybean(Glycine max(L.) Merr). In total, 22 Chinese SMV strains(SC1–SC22) have been classified based on the responses of 10 soybean cultivar...Soybean mosaic virus(SMV) is one of the most devastating viral pathogens of soybean(Glycine max(L.) Merr). In total, 22 Chinese SMV strains(SC1–SC22) have been classified based on the responses of 10 soybean cultivars to these pathogens. However, although several SMVresistance loci in soybean have been identified, no gene conferring SMV resistance in the resistant soybean cultivar(cv.) Kefeng No.1 has been cloned and verified. Here, using F_(2)-derived F_(3)(F_(2:3)) and recombinant inbred line(RIL) populations from a cross between Kefeng No.1 and susceptible soybean cv. Nannong 1138-2, we localized the gene in Kefeng No.1 that mediated resistance to SMV-SC3 strain to a 90-kb interval on chromosome 2. To study the functions of candidate genes in this interval, we performed Bean pod mottle virus(BPMV)-induced gene silencing(VIGS). We identified a recombinant gene(which we named R_(SC3)K) harboring an internal deletion of a genomic DNA fragment partially flanking the LOC100526921 and LOC100812666 reference genes as the SMV-SC3 resistance gene.By shuffling genes between infectious SMV DNA clones based on the avirulent isolate SC3 and virulent isolate 1129, we determined that the viral protein P3 is the avirulence determinant mediating SMV-SC3 resistance on Kefeng No.1. P3 interacts with RNase proteins encoded by R_(SC3)K, LOC100526921, and LOC100812666. The recombinant R_(SC3)K conveys much higher anti-SMV activity than LOC100526921 and LOC100812666, although those two genes also encode proteins that inhibit SMV accumulation, as revealed by gene silencing in a susceptible cultivar and by overexpression in Nicotiana benthamiana. These findings demonstrate that R_(SC3)K mediates the resistance of Kefeng No.1 to SMV-SC3 and that SMV resistance of soybean is determined by the antiviral activity of RNase proteins.展开更多
Soybean mosaic virus (SMV) causes severe yield losses and seed quality reduction in soybean (Glycine max) production worldwide. Rsc4 from cultivar Dabaima is a dominant genetic locus for SMV resistance, and its mappin...Soybean mosaic virus (SMV) causes severe yield losses and seed quality reduction in soybean (Glycine max) production worldwide. Rsc4 from cultivar Dabaima is a dominant genetic locus for SMV resistance, and its mapping interval contains three Nucleotide-binding domain Leucine-rich Repeat containing (NLR) candidates (Rsc4-1, Rsc4-2, and Rsc4-3). The NLR-type resistant proteins were considered as important intracellular pathogen sensors in the previous studies. In this research, based on transient expression assay in Nicotiana benthamiana leaves, we found that the longest transcript of Rsc4-3 is sufficient to induce resistance response to SMV;and CRISPR/Cas9-mediated Rsc4-3 knockout in resistant cultivar Dabaima compromised the resistance. These indicate that Rsc4-3 confers resistance to SMV. Interestingly, Rsc4-3 encodes a cell wall localized NLR-type resistant protein (Rsc4-3). The internal polypeptide region responsible for apoplastic targeting of Rsc4-3 and the putative palmitoylation sites on the N-terminus are essential for the resistance response. Furthermore, we showed that viral-encoded cylindrical inclusion (CI) protein partially localizes to the cell wall and can interact with Rsc4-3. Virus-driven or transient expression of CI protein of avirulent SMV strains is enough to induce resistance response in the presence of Rsc4-3, suggesting that CI is the avirulent gene for Rsc4-3 mediated resistance. Our work exhibited a case of NLR recognizing virus in the apoplast and provided a simple and effective method for identifying resistant genes against SMV infection.展开更多
基金supported by the National Key Research and Development Program of China (2022YFF1001500)the Open Competition Project of Seed Industry Revitalization of Jiangsu Province (JBGS[2021]060)+3 种基金the Core Technology Development for Breeding Program of Jiangsu Province (JBGS-2021-014)China Agriculture Research System of MOF and MARA (CARS-04)the Jiangsu Collaborative Innovation Center for Modern Crop Production (JCIC-MCP)Collaborative Innovation Center for Modern Crop Production Co-sponsored by Province and Ministry (CIC-MCP)。
文摘Drought-induced protein 19(Di19) is a Cys2/His2 zinc-finger protein that functions in plant growth and development and in tolerance to abiotic stresses.Gm PUB21,an E3 ubiquitin ligase,negatively regulates drought and salinity response in soybean.We identified potential interaction target proteins of Gm PUB21by yeast two-hybrid c DNA library screening,Gm Di19-5 as a candidate.Bimolecular fluorescence complementation and glutathionine-S-transferase pull-down assays confirmed the interaction between Gm Di19-5 and Gm PUB21.Gm Di19-5 was induced by Na Cl,drought,and abscisic acid(ABA) treatments.Gm Di19-5 was expressed in the cytoplasm and nucleus.Gm Di19-5 overexpression conferred hypersensitivity to drought and high salinity,whereas Gm Di19-5 silencing increased drought and salinity tolerance.Transcripts of ABA-and stress response-associated genes including Gm RAB18 and Gm DREB2A were downregulated in Gm Di19-5-overexpressing plants under drought and salinity stresses.ABA decreased the protein level of Gm Di19-5 in vivo,whereas Gm PUB21 increased the decrease of Gm Di19-5 after exogenous ABA application.The accumulation of Gm PUB21 was also inhibited by Gm Di19-5.We conclude that Gm PUB21 and Gm Di19-5 collaborate to regulate drought and salinity tolerance via an ABA-dependent pathway.
基金supported by the National Key R&D Program of China (2021YFD1201604)Jiangsu Collaborative Innovation Center for Modern Crop Production (JCIC-MCP)+1 种基金the Core Technology Development for Breeding Program of Jiangsu Province (JBGS-2021-014)China Agriculture Research System of MOF and MARA (No. CARS-04)。
文摘Soybean mosaic virus(SMV) is one of the most devastating viral pathogens of soybean(Glycine max(L.) Merr). In total, 22 Chinese SMV strains(SC1–SC22) have been classified based on the responses of 10 soybean cultivars to these pathogens. However, although several SMVresistance loci in soybean have been identified, no gene conferring SMV resistance in the resistant soybean cultivar(cv.) Kefeng No.1 has been cloned and verified. Here, using F_(2)-derived F_(3)(F_(2:3)) and recombinant inbred line(RIL) populations from a cross between Kefeng No.1 and susceptible soybean cv. Nannong 1138-2, we localized the gene in Kefeng No.1 that mediated resistance to SMV-SC3 strain to a 90-kb interval on chromosome 2. To study the functions of candidate genes in this interval, we performed Bean pod mottle virus(BPMV)-induced gene silencing(VIGS). We identified a recombinant gene(which we named R_(SC3)K) harboring an internal deletion of a genomic DNA fragment partially flanking the LOC100526921 and LOC100812666 reference genes as the SMV-SC3 resistance gene.By shuffling genes between infectious SMV DNA clones based on the avirulent isolate SC3 and virulent isolate 1129, we determined that the viral protein P3 is the avirulence determinant mediating SMV-SC3 resistance on Kefeng No.1. P3 interacts with RNase proteins encoded by R_(SC3)K, LOC100526921, and LOC100812666. The recombinant R_(SC3)K conveys much higher anti-SMV activity than LOC100526921 and LOC100812666, although those two genes also encode proteins that inhibit SMV accumulation, as revealed by gene silencing in a susceptible cultivar and by overexpression in Nicotiana benthamiana. These findings demonstrate that R_(SC3)K mediates the resistance of Kefeng No.1 to SMV-SC3 and that SMV resistance of soybean is determined by the antiviral activity of RNase proteins.
基金This work was supported by the National Natural Science Foundation of China(31571690,31770164)the Fundamental Research Funds for the Central Universities(KYT201801)+3 种基金Program for Changjiang Scholars and Innovative Research Team in University(PCSIRT_17R55)the National Soybean Industrial Technology System of China(CARS-004)Jiangsu Collaborative Innovation Center for Modern Crop Production(JCIC-MCP),the National Key R&D Program of China(2017YFD0101501)the Natural Science Foundation of Jiangsu Province(BK20180039).
文摘Soybean mosaic virus (SMV) causes severe yield losses and seed quality reduction in soybean (Glycine max) production worldwide. Rsc4 from cultivar Dabaima is a dominant genetic locus for SMV resistance, and its mapping interval contains three Nucleotide-binding domain Leucine-rich Repeat containing (NLR) candidates (Rsc4-1, Rsc4-2, and Rsc4-3). The NLR-type resistant proteins were considered as important intracellular pathogen sensors in the previous studies. In this research, based on transient expression assay in Nicotiana benthamiana leaves, we found that the longest transcript of Rsc4-3 is sufficient to induce resistance response to SMV;and CRISPR/Cas9-mediated Rsc4-3 knockout in resistant cultivar Dabaima compromised the resistance. These indicate that Rsc4-3 confers resistance to SMV. Interestingly, Rsc4-3 encodes a cell wall localized NLR-type resistant protein (Rsc4-3). The internal polypeptide region responsible for apoplastic targeting of Rsc4-3 and the putative palmitoylation sites on the N-terminus are essential for the resistance response. Furthermore, we showed that viral-encoded cylindrical inclusion (CI) protein partially localizes to the cell wall and can interact with Rsc4-3. Virus-driven or transient expression of CI protein of avirulent SMV strains is enough to induce resistance response in the presence of Rsc4-3, suggesting that CI is the avirulent gene for Rsc4-3 mediated resistance. Our work exhibited a case of NLR recognizing virus in the apoplast and provided a simple and effective method for identifying resistant genes against SMV infection.
