Zhang Mei, Liu Baoxiu
Provincial Key Laboratory of Applied Botany, South China Botanical Garden, Chinese Academy of Sciences, Guangzhou, China.
University of Chinese Academy of Sciences, Beijing, China.
PLoS One. 2017 Apr 10;12(4):e0174987. doi: 10.1371/journal.pone.0174987. eCollection 2017.
Metal tolerance proteins (MTPs) are a gene family of cation efflux transporters that occur widely in plants and might serve an essential role in metal homeostasis and tolerance. Our research describes the identification, characterization, and localization of OsMTP11, a member of the MTP family from rice. OsMTP11 was expressed constitutively and universally in different tissues in rice plant. Heterologous expression in yeast showed that OsMTP11 complemented the hypersensitivity of mutant strains to Mn, and also complemented yeast mutants to other metals, including Co and Ni. Real time RT-PCR analysis demonstrated OsMTP11 expression was substantially enhanced following 4 h under Cd, Zn, Ni, and Mn treatments, suggesting possible roles of OsMTP11 involvement in heavy metal stress responses. Promoter analysis by transgenic assays with GUS as a reporter gene and mRNA in situ hybridization experiments showed that OsMTP11 was expressed specifically in conducting tissues in rice. DNA methylation assays of genomic DNA in rice treated with Cd, Zn, Ni, and Mn revealed that decreased DNA methylation levels were present in the OsMTP11 promoter region, which was consistent with OsMTP11 induced-expression patterns resulting from heavy metal stress. This result suggested that DNA methylation is one of major factors regulating expression of OsMTP11 through epigenetic mechanisms. OsMTP11 fused to green fluorescent protein (GFP) localized to the entire onion epidermal cell cytoplasm, while vacuolar membrane exhibited increased GFP signals, consistent with an OsMTP11 function in cation sequestration. Our results indicated that OsMTP11 might play vital roles in Mn and other heavy metal transportation in rice.
金属耐受性蛋白（MTPs）是一类阳离子外流转运体基因家族，广泛存在于植物中，可能在金属稳态和耐受性方面发挥重要作用。我们的研究描述了水稻MTP家族成员OsMTP11的鉴定、特性及定位。OsMTP11在水稻植株的不同组织中组成型且普遍表达。在酵母中的异源表达表明，OsMTP11弥补了突变菌株对锰的超敏性，也弥补了酵母突变体对包括钴和镍在内的其他金属的超敏性。实时逆转录聚合酶链反应（RT-PCR）分析表明，在镉、锌、镍和锰处理4小时后，OsMTP11的表达显著增强，这表明OsMTP11可能参与重金属胁迫反应。以GUS作为报告基因的转基因分析和mRNA原位杂交实验进行的启动子分析表明，OsMTP11在水稻的输导组织中特异性表达。对用镉、锌、镍和锰处理的水稻基因组DNA进行的DNA甲基化分析显示，OsMTP11启动子区域的DNA甲基化水平降低，这与重金属胁迫诱导的OsMTP11表达模式一致。该结果表明，DNA甲基化是通过表观遗传机制调节OsMTP11表达的主要因素之一。与绿色荧光蛋白（GFP）融合的OsMTP11定位于整个洋葱表皮细胞的细胞质中，而液泡膜显示出增强的GFP信号，这与OsMTP11在阳离子螯合中的功能一致。我们的结果表明，OsMTP11可能在水稻对锰和其他重金属的转运中发挥重要作用。
