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URI permanente para esta coleçãohttps://locus.ufv.br/handle/123456789/11847
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Item Revisiting the soybean GmNAC superfamily(Frontiers in Plant Science, 2018-12) Melo, Bruno P.; Fraga, Otto T.; Silva, José Cleydson F.; Ferreira, Dalton O.; Brustolini, Otávio J. B.; Carpinetti, Paola A.; Machado, Joao Paulo B.; Reis, Pedro A. B.; Fontes, Elizabeth P. B.The NAC (NAM, ATAF, and CUC) genes encode transcription factors involved with the control of plant morph-physiology and stress responses. The release of the last soybean (Glycine max) genome assembly (Wm82.a2.v1) raised the possibility that new NAC genes would be present in the soybean genome. Here, we interrogated the last version of the soybean genome against a conserved NAC domain structure. Our analysis identified 32 putative novel NAC genes, updating the superfamily to 180 gene members. We also organized the genes in 15 phylogenetic subfamilies, which showed a perfect correlation among sequence conservation, expression profile, and function of orthologous Arabidopsis thaliana genes and NAC soybean genes. To validate our in silico analyses, we monitored the stress-mediated gene expression profiles of eight new NAC-genes by qRT-PCR and monitored the GmNAC senescence-associated genes by RNA-seq. Among ER stress, osmotic stress and salicylic acid treatment, all the novel tested GmNAC genes responded to at least one type of stress, displaying a complex expression profile under different kinetics and extension of the response. Furthermore, we showed that 40% of the GmNACs were differentially regulated by natural leaf senescence, including eight (8) newly identified GmNACs. The developmental and stress-responsive expression profiles of the novel NAC genes fitted perfectly with their phylogenetic subfamily. Finally, we examined two uncharacterized senescence-associated proteins, GmNAC065 and GmNAC085, and a novel, previously unidentified, NAC protein, GmNAC177, and showed that they are nuclear localized, and except for GmNAC065, they display transactivation activity in yeast. Consistent with a role in leaf senescence, transient expression of GmNAC065 and GmNAC085 induces the appearance of hallmarks of leaf senescence, including chlorophyll loss, leaf yellowing, lipid peroxidation and accumulation of H 2 O 2 . GmNAC177 was clustered to an uncharacterized subfamily but in close proximity to the TIP subfamily. Accordingly, it was rapidly induced by ER stress and by salicylic acid under late kinetic response and promoted cell death in planta. Collectively, our data further substantiated the notion that the GmNAC genes display functional and expression profiles consistent with their phylogenetic relatedness and established a complete framework of the soybean NAC superfamily as a foundation for future analyses.Item Functional and regulatory conservation of the soybean ER stress-induced DCD/NRP- mediated cell death signaling in plants(BioMed Plant Biology, 2016-07-12) Reis, Pedro A. B.; Carpinetti, Paola A.; Freitas, Paula P.J.; Santos, Eulálio G.D.; Camargos, Luiz F.; Oliveira, Igor H.T.; Silva, José Cleydson F.; Carvalho, Humberto H.; Dal-Bianco, Maximiller; Soares-Ramos, Juliana R.L.; Fontes, Elizabeth P. B.The developmental and cell death domain (DCD)-containing asparagine-rich proteins (NRPs) were first identified in soybean (Glycine max) as transducers of a cell death signal derived from prolonged endoplasmic reticulum (ER) stress, osmotic stress, drought or developmentally-programmed leaf senescence via the GmNAC81/GmNAC30/GmVPE signaling module. In spite of the relevance of the DCD/NRP-mediated signaling as a versatile adaptive response to multiple stresses, mechanistic knowledge of the pathway is lacking and the extent to which this pathway may operate in the plant kingdom has not been investigated. Here, we demonstrated that the DCD/NRP-mediated signaling also propagates a stress-induced cell death signal in other plant species with features of a programmed cell death (PCD) response. In silico analysis revealed that several plant genomes harbor conserved sequences of the pathway components, which share functional analogy with their soybean counterparts. We showed that GmNRPs, GmNAC81and VPE orthologs from Arabidopsis, designated as AtNRP-1, AtNRP-2, ANAC036 and gVPE, respectively, induced cell death when transiently expressed in N. benthamiana leaves. In addition, loss of AtNRP1 and AtNRP2 function attenuated ER stress-induced cell death in Arabidopsis, which was in marked contrast with the enhanced cell death phenotype displayed by overexpressing lines as compared to Col-0. Furthermore, atnrp-1 knockout mutants displayed enhanced sensitivity to PEG-induced osmotic stress, a phenotype that could be complemented with ectopic expression of either GmNRP-A or GmNRP-B. In addition, AtNRPs, ANAC036 and gVPE were induced by osmotic and ER stress to an extent that was modulated by the ER-resident molecular chaperone binding protein (BiP) similarly as in soybean. Finally, as putative downstream components of the NRP-mediated cell death signaling, the stress induction of AtNRP2, ANAC036 and gVPE was dependent on the AtNRP1 function. BiP overexpression also conferred tolerance to water stress in Arabidopsis, most likely due to modulation of the drought-induced NRP-mediated cell death response. Our results indicated that the NRP-mediated cell death signaling operates in the plant kingdom with conserved regulatory mechanisms and hence may be target for engineering stress tolerance and adaptation in crops.