Abstract：The APETALA2/ethylene-responsive element binding protein (AP2/EREBP) superfamily is one of the largest and most conserved gene families in plant. It has great contributions in plant growth, development and response to diverse stresses such as extreme temperature (freezing damage, and heat stress), drought, high salinity and pathogen infection. It has also been involved in various hormone-related signal transduction pathways. The AP2/EREBP superfamily could be classified as four subfamilies, i.e., ERF (ethylene-responsive factor), DREB (dehydration-responsive element binding protein), RAV (related to ABI3/VP) and AP2. DREB subfamily genes play an important role in response to drought, high salt and low temperature stress in higher plants. In our previous analysis of Brachypodium distachyon AP2/EREBP gene superfamily, we found that B. distachyon DREB38 gene (BdDREB38) of DREB subfamily was significantly induced by cold, while its expression profile under other stress conditions and the activity of its promoter was still unclear. In this study, the expression profile of BdDREB38 gene under several abiotic stress conditions including cold, drought, NaCl, abscisic acid (ABA), salicylic acid (SA) and H2O2 was detected by qRT-PCR. The result showed that the expression of BdDREB38 gene had no obvious change at each time point of NaCl and H2O2 treatments in contrast with the control (without treatment), but increased gradually with the extension of drought treatment time. Moreover, the expression of BdDREB38 gene was significantly higher than that of control at 1 h point after cold treatment and at 2 h point after ABA treatment, whereas the expression of BdDREB38 gene decreased at 5 h point after SA treatment. These results indicated that the promoter of BdDREB38 gene might be a stress-inducible promoter. To further investigate the structure and function of BdDREB38 promoter in B. distachyon, a 1 510 bp fragment (named as PBdDREB38) at the upstream of this gene was cloned. Plant CARE analysis reveals that this promoter not only consists of the basic cis-elements such as TATA box and CAAT box, but also includes some cis-elements involved in adversity stress and light response, such as LTR (lower temperature response element)、HSE (Heat shock response element), TC-rich repeats (defense and stress response element), SP1(light response element), TCA-element (SA response element) and so on. To investigate the expression profiles of this promoter, it was fused with the β-glucuronidase (GUS) reporter gene in the recombinant expression vector pCAMBIA1381-GUS, and then the constructed expression vector was transformed into tobacco (Nicotiana tabacum) through Agrobacterium-mediated method. GUS staining result showed that BdDREB38 promoter could be significantly induced by drought stress, but was not induced by cold treatment, although BdDREB38 gene enhanced its expression after 2 h cold treatment. This case may be the reason that one regulatory element in the promoter region is not enough to drive the expression of its downstream target genes and needs other remote cis-acting element such as enhancer in the promoter region to promote the binding of transcription factors and finally activates their downstream target genes. This study would provide a theoretical basis for further functional study of BdDREB38 gene and its promoter in B. distachyon.
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