Metal tolerance proteins (MTPs) from the CDF (Cation Diffusion Facilitator) family are efflux transporters that play a crucial role in metal homeostasis by maintaining optimal metal concentrations in the cytoplasm. Here, a novel tobacco NtMTP2 transporter was cloned and characterized. It encodes a 512 aa protein containing all specific CDF family domains. A GFP-NtMTP2 fusion protein localizes to the tonoplast in tobacco cells. NtMTP2 expression in yeast conferred tolerance to Co and Ni, indicating that the protein mediates transport of both metals, but not Zn, Mn, Cu, Fe, or Cd. Nonetheless, the expression level was not affected by Co or Ni, except for an increase in leaves at high Co concentrations. Its expression in plant parts remained stable during development, but increased in the leaves of older plants. Analysis of tobacco expressing a promoter-GUS construct indicates that the main sites of promoter activity are the conductive tissue throughout the plant and the palisade parenchyma in leaves. Our results suggest that NtMTP2 is a tonoplast transporter mediating sequestration of Co and Ni into vacuoles and an important housekeeping protein that controls the basal availability of micronutrients and plays a role in the sequestration of metal excess, specifically in leaves.

中文翻译：

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烟草金属耐受蛋白 NtMTP2 的鉴定和表征

来自 CDF（阳离子扩散促进剂）家族的金属耐受蛋白 (MTP) 是外排转运蛋白，通过维持细胞质中的最佳金属浓度在金属稳态中发挥关键作用。在这里，一种新型烟草NtMTP2转运蛋白被克隆和表征。它编码包含所有特定 CDF 家族域的 512 个氨基酸的蛋白质。GFP-NtMTP2 融合蛋白定位于烟草细胞的液泡膜。NtMTP2在酵母中的表达赋予对 Co 和 Ni 的耐受性，表明该蛋白质介导两种金属的转运，但不介导 Zn、Mn、Cu、Fe 或 Cd 的转运。尽管如此，表达水平不受 Co 或 Ni 的影响，除了在高 Co 浓度下叶子增加。它在植物部分的表达在发育过程中保持稳定，但在较老植物的叶子中表达增加。对表达启动子-GUS 构建体的烟草的分析表明，启动子活性的主要位点是整个植物的传导组织和叶子中的栅栏薄壁组织。我们的研究结果表明，NtMTP2 是一种液泡膜转运蛋白，介导 Co 和 Ni 螯合到液泡中，并且是一种重要的管家蛋白，可控制微量营养素的基础可用性，并在金属过量的螯合中发挥作用，特别是在叶子中。