Functional Analysis Of Two DREB Type Transcription Repressors In Plant

Dehydration-responsive-element binding protein (DREB)-type transcription factors are well known to play important roles in adaptation to abiotic stress in plant. The mechanisms by which DREB-type transcription factors activate stress-induced gene expression have been relatively well studied. However, little is known about how DREB-type transcriptional repressors modulate plant stress responses. In this study, we report the functional analysis of two DREB-type transcriptional repressors.Cotton dehydration-responsive element (DRE) binding protein 1 (GhDBP1) was firstly identified as a DREB-type transcriptional repressor. In this study, we demonstrate that GhDBP1's transcriptional repression domain is located at the C-terminus and is known as an ERF-associated amphiphilic repression (EAR)-motif. Furthermore, the amino acid residues aspartic acid (D), leucine (L), asparagine (N), and proline (P) are conserved in its EAR-motif, and they are found to be necessary for repression through mutational analysis. In addition, our promoter assays demonstrate a dehydration-induced and rehydration-repressed expression pattern of GhDBP1. Transgenic Arabidopsis plants overexpressing GhDBP1 are more sensitive to high salinity stress and appeare to down-regulate the expression levels of the stress-induced effector genes.For the further study of DREB-type transcriptional repressors, RAP2.1, a transcription factor from Arabidopsis is also intensively studied. RAP2.1 possesses an AP2/ERF domain that binds to DRE elements and an EAR-motif, as the repression domain located at the C-terminus of the protein. Expression of RAP2.1 is strongly induced by drought and cold stress via an ABA-independent pathway. Arabidopsis plants overexpressing RAP2.1 show enhanced sensitivity to cold and drought stresses, while rap2.1-1 and rap2.1-2 T-DNA insertion alleles result in reduced sensitivity to these stresses. The reduced stress sensitivity of the plant containing the rap2.1 allele can be genetically complemented by the expression of RAP2.1, but not by the expression of EAR-motif-mutated RAP2.1. Furthermore, chromatin immunoprecipitation (ChIP) analysis has identified Responsive to desiccation/Cold-regulated (RD/COR) genes as downstream targets of RAP2.1 in vivo. Stress-induced expression of the RD/COR genes is repressed by overexpression of RAP2.1 and is increased in plants expressing the rap2.1 allele. In addition, RAP2.1 can negatively regulate its own expression by binding to DREs present in its own promoter.The functional analysis of two DREB-type transcriptional repressors, GhDBP1 from cotton and RAP2.1 from Arabidopsis, suggested that DREB-type transcriptional repressors are involved in the precise regulation of expression of stress-related genes, acting to keep stress responses under tight control, so that the self-inflicted damage caused by a runaway stress response can be prevented. This study enlarges our knowledge about plant DREBs, and also provides more information about the negative regulation of stress signal transduction in plant.