植物螯合肽合酶基因AtPCS2的表达调控

Regulation of the Phytochelatin Synthase Gene AtPCS2 in Arabidopsis thaliana

植物螯合肽合酶(PCS)受重金属离子激活,并以还原型谷胱甘肽为底物合成植物螯合肽(PCs),在植物和真菌的重金属解毒机制中起重要作用.拟南芥基因组中有两个编码PCS的基因AtPCS1和AtPCS2,但AtPCS1单基因功能缺失即可导致相应的突变体cad1-3对镉高度敏感,其体内也检测不到PCs;而体外表达分析表明,AtPCS2具有完全的PCs合酶活性,预示植物体内可能存在AtPCS2的负向调控机制.基于该推测,构建了CaMV 35S启动子驱动的AtPCS2基因编码区与c-Myc抗原标签融合的过表达载体.结果表明,在cad1-3的CaMV 35S/AtPCS2:cMyc的异位表达株系中,AtPCS2的mRNA和蛋白都保持较高的表达量.不仅如此,AtPCS2具有植物螯合肽合成能力,并完全互补了cad1-3突变体的镉敏感性状.AtPCS2和EYFP的融合蛋白在细胞质有明显表达,在细胞核也检测到一定信号.以上结果表明,AtPCS2在植物体内可能主要受转录水平调控,而且可能具有调节PCs合成以外的其他生化功能.

Phytochelatin synthase （PCS） plays an essential role in heavy metal detoxification in plants and fungi, via the catalyzation of phytochelatins （PCs） biosynthesis from the substrate of glutathione （GSH）. Two PCS genes AtPCS1 and AtPCS2 are present in Arabidopsis genome. However, loss of function of only the AtPCS1 gene resulted in undetectable PCs and the consequent metal hypersensitivity in the corresponding mutant eadl-3, while in vitro assay indicated that AtPCS2 is capable to efficiently catalyze PCs biosynthesis. These results suggest that repression mechanisms exist in Arabidopsis to tightly control AtPCS2 function in vivo. To test and decipher this hypothesis, we generated the construct 35S/AtPCS2：：cMyc and transformed it into the PCs-deficient mutant cadl-3. Our results showed that both the AtPCS2 mRNA and protein levels were steadily high in the transgenic plants. Consistently, PCs biosynthesis and metal tolerance were also observed when AtPCS2 was introduced into cadl-3. Furthermore, in addition to the cytosol, AtPCS2 was also localized to the nuclei. Taken together, all these results indicate that AtPCS2 is under tight control mainly at the transcription level in Arabidopsis, and it may function beyond catalyzing PCs biosynthesis.