Transcriptome Analysis Of Rice Young Spike Under High Temperature Stress And Preliminary Functional Characterization Of OsHSP23.7and OsDREBA5-1Genes

Abiotic stresses are the major impediments that restrain plant growth and result in significant reductions in crop yields and quality. Global warming and frequent occurrence of drought seriously affect crop productivity. Rice is the world's most important food crop and a primary source of food for more than half the world's population. As the most important staple food, rice supply means much to the people of South East (SE). To ensure high and stable yields of cereals, screening and cultivating rice varieties with improved tolerance to high temperature and drought are of great significance. Identification of genes/proteins responsive to abiotic stresses is an essential first step towards understanding the molecular mechanisms, and thereby producing transgenic plants with enhanced tolerance to a particular stress.High-throughput gene chip technology is one of the most important methods to study gene functions. In order to study the influence of high temperature on rice young spike development, a22K Rice Oligo Microarray kit was used to analysis the gene expression profiles in young spikes of two rice cultivars (heat-tolerant'996'and heat-susceptible '4628', at microspore mother cell meiosis stage) suffered to three-day high temperature treatment. We found that219and457genes (transcripts) in both '996' and '4628'were commonly up-and down-regulated, respectively. In addition,303and105genes were specifically up-regulated and451and309genes were specifically down-regulated in '996' and'4628', respectively.Among these common differentially expressed genes, the gene groups involving in cell cycle and DNA processing, cell wall/cell structure, primary metabolism and energy and secondary metabolism were significantly overrepresented in down-regulated gene clusters. On the other hand, the gene groups related to regulation/signaling and stress/defense-related were overrepresented in up-regulated gene clusters. These results indicate that long period of high temperature stress seriously affect the normal cellular activities, and activate some stress-responsive and regulation-related genes to adjust to high temperature. Several classes of stress-related genes including HSP, AP2/EREBP, HSF, WRKY and GST were specifically/more up-regulated in '996' or specifically/more down-regulated in '4628', indicating that these genes may play certain roles in rice heat-tolerance.HSPs act as molecular chaperons and play an important role in plant stress tolerance. In our previous study, we found that the expression of OsHSP23.7was up-regulated by high temperature and high salt stresses. In the present study, our data showed that OsHSP23.7-OE transgenic lines demonstrated higher germination ability compared to wild type (WT) plants when subjected to mannitol and NaCl stresses. In addition, OsHSP23.7-OE transgenic rice lines showed significantly lower REC, lower MDA content and higher free proline content than WT under drought and salt stress conditions, and displayed higher survival rate after drought and high salt treatments. These results suggest that overexpression of OsHSP23.7improve tolerance to drought and salt in transgenic rice plants. Real-time PCR analysis suggested that the contribution of OsHSP23.7on stress tolerance is not due to altered expression of SODs and APXs genes.DREBs are a subfamily of AP2/EREBP transcription factor family, which play important role in abiotic stress tolerance in plants.OsDREBA5-1was a nuclear target protein predicted by cellular localization software and confirmed by transient expression assay in onion epidermal cells. Overexpression of OsDREBA5-1leaded to enhanced ABA sensitivity in transgenic rice while OsDREBA5-1-RNAi transgenic rice plants were less sensitive to24-epibrassinolide (eBL). Drought treatment at seedling stage showed that overexpression of OsDREBA5-1enhanced drought tolerance in transgenic rice plants. In addition, overexpression of OsDREBA5-1resulted in increased leaf angle in transgenic rice plants. SEM and paraffin slice analysises revealed that the adaxial cells were longitudinally elongated in the collar of OsDREBA5-1-OE rice plant compared to WT. Real-time PCR assay revealed that the expression levels of several drought responsive genes were more highly up-regulated in OsDREBA5-1overexpression plants than WT under drought condition. Real-time PCR assay also showed that the expression levels of OsDWARF4and the genes involved in cell division and elongation and cell wall remodeling were affected by OsDREBA5-1. Together, these results demonstrate that OsDREBA5-1plays a positive role in drought stress response may through transcriptional regulation of some stress-related genes; OsDREBA5-1is involved in brassinosteroid signal transduction, and may regulate leaf inclination through modifying the expression of genes involved in cell division and elongation and cell wall remodeling.