The Studies Of Drought-resistant Mechanisms Of Rice's Transcription Factor OsbZIP16

One third of the people all over of the world and 60%people of China make rice as their staple food.So insuring the stale and high yield of rice is the basic national policy of China.Arid,which possesses features of high frequency,long duration and a wide range of affected areas,is one of the climate disasters that have the most serious impacts on the stale and high yield of rice.Improving the drought tolerance of rice is an important aspect of rice stress-resistant breeding.Previous genome expression analysis showed that the expression of OsbZIP16 was strongly induced by drought and high sanility stresses.Recent gene chip data also validated that 26 rice bZIP transcription factors including OsbZIP16 were up-regulated under drought and high sanility stresses.These researches indicated that some bZIP transcription factors play roles in drought signal transduction.Through investigating the function of OsbZIP16 in drought response in rice,this research provides a further insight into the drought-resistant mechanisms of rice in order to accumulate theoretic foundation about rice's drought-resistant breeding and involves some primary parts below:1.Clone and sequence analysis of OsbZIP16.This research characterized a basic leucine zipper(bZIP)transcription factor OsbZIP16 involved in rice's drought signal transduction.Previous researchs have predicted 89 rice bZIP transcription factors and classified them into 11 groups according to their DNA-binding specificity and homology within basic lucine zipper region.After cloning OsbZIP16,alignments between OsbZIP16 and several orther functionally analysed group IV bZIP transcription factors showed that their protein sequences had no high homology except for the bZIP region.Two function-known domain,bZIP and NLS,were found in OsbZIP16 protein sequence.2.Validation the OsbZIP16 as a transcription factor.Given the OsbZIP16 is only a transcription factor predicted by bioinformatics.OsbZIP 16's function performenced as a transcription factor should first be validated.Transient expression of fusion protein GFP-OsbZIP16 in onion epidermal cells indicated that OsbZIP16 was localized in the nucleus.A transactivation assay in yeast showed that the N terminal of OsbZIP 16 was sufficient and necessary for its transactivation activity.3.The expression profile analyses of OsbZIP16.Real-time PCR wae used to study the profiles of OsbZIP16.No matter under normal or under drought conditions,constitutive expression of OsbZIP16 was detected in different tissues and organs at both sedling and booting stages and the mature leaves possessed the highest expression level.Under drought and high sanility stress,the expression level of OsbZIP16 was rapidly increased.These results indicated that OsbZIP16 probablely participate the drought signal transduction.4.The analysis of drought tolerance mediated by OsbZIP16 in rice.Through the comparision of drought tolerance between wild type and transgenic plants,it is can be validated that OsbZIP16 is involved in drought signaling.At both the seedling and tillering stages,transgenic rice plants overexpressing OsbZIP16 exhibited significantly improved drought tolerance,which were positively correlated with the expression levels of OsbZIP16.The change of phenotype is the result of the expression profile changes of some related genes,so the expression level changes of some drought tolerance-related genes were detected by real-time PCR.Representative downstream drought-responsive genes showed significantly higher expression levels in transgenic rice plants than wild types under drought conditions but not under normal conditions.5.OsbZIP16 was involved in ABA-dependent drought signal transduction.Given ABA-dependent and ABA-independent drought signal transduction exist in plants,which one OsbZIP16 is involved in should be determined.OsbZIP16 was shown to be induced by exogenous ABA treatment,while overexpression of OsbZIP16 made transgenic plants more sensitive to ABA than wild type plants.No difference in the expression levels of two upstream genes involved in ABA signal transduction was observed between wild type and transgenic plants,but the expression levels of two downstream genes showed significantly higher expression levels in transgenic plants than in wild type plants under drought conditions.These results further indicated that OsbZIP16 is involved in ABA-dependent drought signal transduction,and suggests that the transcription factor functions downstream of SAPK10 and PP2C and upstream of LEA3-1 and RAB16C.6.The analysis of genome-wide expressive regulation of OsbZIP16 and the screen of proteins interacting with OsbZIP16.OsbZIP16 is a transcription factor and regulate a number of downstream genes'expression.So the expression level change of OsbZIP16 definitely causes the expression level changes of numerous downstream genes which are most likely related to drought tolerance in rice.Transcriptome analysis identified a lot of differentially expressed genes between wild types and OsbZIP16 overexpression plants,many of which were clustered in stress-responsive categories of gene ontology,such as antioxidant activity,protein kinase activity and response to oxidative stress.Transcription factors are regulated by or coordinately promte the downstream genes'transcription with orther proteins.Yeast two hybrid systems were used to screen the proteins interacted with OsbZIP16.A total of 23 protein interacted with OsbZIP16 were screened out.Overall,our findings indicate that OsbZIP16 positively regulates the drought tolerance in rice possibly through ABA-dependent signal transduction.