Laboratory of Molecular Plant Science, Department of Horticulture, Washington State University, Pullman, WA 99164-6414, USA.
Plant Sci. 2012 Apr;185-186:274-80. doi: 10.1016/j.plantsci.2011.11.013. Epub 2011 Nov 28.
GT factors are a family of plant-specific transcription factors with conserved trihelix DNA-binding domains that bind GT elements. By screening a cDNA expression library with (35)S-labeled recombinant calmodulin (CaM), we identified AtGT2L, a classic member of GT-2 subfamily, as a Ca(2+)-dependent CaM-binding protein. AtGT2L specifically targets the nucleus and possesses both transcriptional activation and DNA-binding abilities, implicating its function as a nuclear transcription factor. The CaM-binding domain (CaMBD) of AtGT2L is embedded in its C-terminal DNA-binding domain, but the in vitro DNA-binding ability of AtGT2L was not affected by its interaction with CaM. Site-directed mutagenesis experiments further revealed that Met(506) and Leu(507) in the CaMBD are critical for both CaM- and DNA-binding abilities of AtGT2L. Northern blot analysis showed that AtGT2L mRNA levels are higher in old rosette leaves, cauline leaves and flowers than in other organs. In addition, the expression of AtGT2L was responsive to cold, NaCl and abscisic acid treatments. Furthermore, both basal and chilling/NaCl-induced expressions of the cold- and salt-inducible marker genes RD29A and ERD10 were higher in plants overexpressing AtGT2L than those in wild-type plants. Thus, our data indicate that AtGT2L is a Ca(2+)/CaM-binding nuclear transcription factor involved in plant responses to cold and salt stresses.
GT 因子是一类植物特异性转录因子,具有保守的三螺旋 DNA 结合结构域,能够结合 GT 元件。通过用(35)S 标记的重组钙调素(CaM)筛选 cDNA 表达文库,我们鉴定出 AtGT2L 是 GT-2 亚家族的经典成员,是一种 Ca(2+)依赖的 CaM 结合蛋白。AtGT2L 特异性靶向细胞核,具有转录激活和 DNA 结合能力,暗示其作为核转录因子的功能。AtGT2L 的 CaM 结合域(CaMBD)嵌入其 C 末端 DNA 结合域,但 CaM 与 AtGT2L 相互作用并不影响其体外 DNA 结合能力。定点突变实验进一步表明,CaMBD 中的 Met(506)和 Leu(507)对 AtGT2L 的 CaM 和 DNA 结合能力都是至关重要的。Northern blot 分析表明,AtGT2L mRNA 水平在老的莲座叶、茎生叶和花中高于其他器官。此外,AtGT2L 的表达对冷、NaCl 和脱落酸处理有反应。此外,过表达 AtGT2L 的植物中,基础水平和冷/NaCl 诱导的冷诱导和盐诱导标记基因 RD29A 和 ERD10 的表达均高于野生型植物。因此,我们的数据表明,AtGT2L 是一种参与植物对冷和盐胁迫反应的 Ca(2+)/CaM 结合核转录因子。