Chung PJ, Jung H, Choi YD, Kim JK.

BMC Genomics. 2018 Jan 12;19(1):40. doi: 10.1186/s12864-017-4367-1.

Abstract

BACKGROUND:

Plant stress responses and mechanisms determining tolerance are controlled by diverse sets of genes. Transcription factors (TFs) have been implicated in conferring drought tolerance under drought stress conditions, and the identification of their target genes can elucidate molecular regulatory networks that orchestrate tolerance mechanisms.

RESULTS:

We generated transgenic rice plants overexpressing the 4 rice TFs, OsNAC5, 6, 9, and 10, under the control of the root-specific RCc3 promoter. We showed that they were tolerant to drought stress with reduced loss of grain yield under drought conditions compared with wild type plants. To understand the molecular mechanisms underlying this tolerance, we here performed chromatin immunoprecipitation (ChIP)-Seq and RNA-Seq analyses to identify the direct target genes of the OsNAC proteins using the RCc3:6MYC-OsNAC expressing roots. A total of 475 binding loci for the 4 OsNAC proteins were identified by cross-referencing their binding to promoter regions and the expression levels of the corresponding genes. The binding loci were distributed among the promoter regions of 391 target genes that were directly up-regulated by one of the OsNAC proteins in four RCc3:6MYC-OsNAC transgenic lines. Based on gene ontology (GO) analysis, the direct target genes were related to transmembrane/transporter activity, vesicle, plant hormones, carbohydrate metabolism, and TFs. The direct targets of each OsNAC range from 4.0-8.7% of the total number of up-regulated genes found in the RNA-Seq data sets. Thus, each OsNAC up-regulates a set of direct target genes that alter root system architecture in the RCc3:OsNAC plants to confer drought tolerance. Our results provide a valuable resource for functional dissection of the molecular mechanisms of drought tolerance.

CONCLUSIONS:

Many of the target genes, including transmembrane/transporter, vesicle related, auxin/hormone related, carbohydrate metabolic processes, and transcription factor genes, that are up-regulated by OsNACs act as the cellular components which would alter the root architectures of RCc3:OsNACs for drought tolerance.

See https://www.ncbi.nlm.nih.gov/pubmed/29329517

Fig. 1

Construction of RCc3:nMYC:OsNAC or RCc3:OsNAC:cMYC transgenic rice plants. a Map of the binary vector used for generating RCc3:OsNAC5, 6, 9, and 10 transgenic rice lines. RB, right border; pRCc3, a root specific promoter [48]; tNOS, NOS terminator; Bar, phosphinothricin acetyltransferase gene, an herbicide resistance gene used as a selection marker; LB, left border. Orange color indicates nMYC (N-terminal portion of 6MYC protein) or cMYC (C-terminal portion of 6MYC protein). b Translational expression of OsNAC5, 6, 9, and 10 proteins fused cMYC or nMYC, as measured by Western blot analysis using an anti-MYC antibody. Protein extracts from roots of twenty-day-old nontransgenic wild type (NT, Oryza sativa L. japonica cv. Illmi) and RCc3: OsNAC5, 6, 9, and 10 transgenic seedlings (10 μg per lane) were subjected to SDS-PAGE (polyacrylamide gel electrophoresis) and Western blot analysis. Coomassie blue staining is shown to confirm equal loading. Red figure, definite overexpressed 6MYC fused OsNAC protein