Abstract
BackgroundPlant 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.ResultsWe 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.ConclusionsMany 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.
Highlights
Plant stress responses and mechanisms determining tolerance are controlled by diverse sets of genes
Our results reveal the regulatory networks governed by OsNAC Transcription factors (TFs) that contribute to drought-tolerance mechanisms, providing a foundation for the development of drought tolerant crops
Identification of genes differentially regulated in RCc3: 6MYCOsNAC roots using RNA sequencing (RNA-Seq) To investigate OsNAC regulatory networks, RNA-Seq analyses were performed of roots from 20 d-old nontransformed (NT) and RCc3:6MYC-OsNAC expressing plants grown under normal growth conditions
Summary
Plant stress responses and mechanisms determining tolerance are controlled by diverse sets of genes. Molecular and genomic analyses have revealed many drought-inducible transcription factors (TFs) that regulate the expression of stress-inducible downstream genes [6, 7]. These include TFs belonging to the AP2/ERF, MYB, bZIP and NAC families. It is estimated that 2408 genes encode TF or TF-like proteins in rice (Oryza sativa), based on the Plant Transcription Factor Database v3.0 (http://planttfdb.cbi.pku.edu.cn). Rice has 170 putative OsNAC genes, based on sequence analysis (Plant Transcription Factor Database, http://planttfdb.cbi.pku.edu.cn/), of which 41 and 29 are induced and repressed, respectively, in leaves of reproductive stage under drought stress conditions [16]. Despite their important function in modulating drought tolerance, little is known about the molecular mechanisms associated with OsNAC TFs and their target genes [19]
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