Study On The Function Of Zoysia Japonica Transcription Factor Gene ZjDREB 1.4

Abiotic stresses,such as high temperature,low temperature,drought,and salinity,are important factors that affect distribution and ornamental value of turfgrass.Therefore,the resistance to abiotic stresses and its underlying mechanism have been an important objectives in turfgrass research.Zoysiagrass(Zoysia Willd.)is a key warm-season turf grass species in China,which has tolerance to multiple stresses including drought,heat,saline/alkali,and diseases.The molecular mechanism of its stress tolerance is worthy of study.DREB1(dehydration-responsive element binding protein 1)can response to a variety of abiotic stresses and enhance tolerance of plants to stresses by regulating the expression of a batch of genes.The function of DREB transcription factors in zoysiagrass has not been studied systematically.In this study,a DREB 1-like transcription factor gene from Zoysia japonica Steud.,designated ZjDREBl.4,was reported.The main results are as follows:(1)The coding region of ZjDREBl.4 is 681 bp in length and has no intron.It encodes a protein with 226 amino acids and has an AP2 domain flanked by sequence characteristic of DREB1 proteins.In the phylogenetic tree of the DREB1 homolog of Gramineous grasses,the ZjDREB1.4 gene belongs to the?a branch that is selectively expanded in warm-season species.Results of qRT-PCR indicated that the expression of ZjDREB1.4 was induced by high temperature,low temperature and drought stress,and significantly induced by cold stress.(2)The fusion gene ZjDREBl.4-GFP was constructed and expressed transiently in tobacco(Nicotiana tobaccum)epidermal cells and expressed permanently in Arabidopsis thaliana.The product of the fusion protein was primarily targeted to the nucleus,and a small proportion of the product was also observed in cytoplasm.(3)Yeast one-hybrid assay indicated that ZjDREB1.4 showed strong transactivation activity but weak binding to the DRE with the core sequence of ACCGAC.Gel shift assay showed that the ZjDREB1.4 protein bound to GCCGAC more preferentially than to ACCGAC.(4)ZjDREB1.4 was cloned into a plant expression vector and transformed into Arabidopsis by Agrobacterium immersion method.Transgenic Arabidopsis plants constitutively expressing ZjDREB1.4 showed similar growth to the wild-type with only a 3-8 day delay in bolting,but had higher survival rates under high temperature or freezing temperature stress.(5)The mechanism of ZjDREBl.4 in enhancing resistance to stress was investigated from the aspects of transcription regulation and osmoregulation by qRT-PCR and measuring physiological parameters.Under normal conditions,the overexpression of ZjDREB1.4 in Arabidopsis modulated the expression of 1,153 genes,among which about 15%of CBF-regulon were included.The levels of proline and soluble sugars increased moderately.In summary,ZjDREBl.4 is a member of A-1 subgroup of DREB subfamily.It can bind to GCCGAC more preferentially than to ACCGAC and function in stress tolerance.The gene profile regulated by ZjDREBl.4 when over-expressed in Arabidopsis is different from that of CBF-regulon,and no obvious adverse effect on growth and development was observed in the transgenic plants.The potential usage of the gene in plant breeding warrants further study.