一个拟南芥叶绿体镁离子转运蛋白AtMGT10/VAR5的功能研究

Magnesium ion (Mg2+) is one of the essential cations in plants, and it plays important roles in regulating physiological, biochemical and developmental processes in plants. Magnesium transporters participate in the balancing and transporting of Mg2+ in plant cells. Through large-scale screening of Arabidopsis leaf-color mutants, we have obtained a new reticulate mutant, designated variegated 5-1 (var5-1). Map-based cloning, complementation analysis, and protoplast transient expression indicate that VAR5 encodes AtMGT10, a putative chloroplast localized magnesium transporter. In contrast to typical magnesium deficiency phenotypes, var5-1 shows pale-green vein tissues and green intervenal tissues. Because knock-out mutations of AtMGT10 are lethal, in this proposal, we are going to use the unique material var5-1 mutants to study: 1) cellular and subcellular phenotypes of yellow reticulate vein; 2) how plants with different expression levels of AtMGT10/VAR5 respond to different treatments of Mg2+; 3) the subcellular localization and topology of AtMGT10/VAR5. This work will help us understand the functions of AtMGT10/VAR5 in regulating plant Mg2+ homeostasis and provide new insight into plant mineral metabolism and transportation.

Mg2+是植物必需金属离子之一，其平衡对于植物细胞生理、生化过程及生长发育等具有重要作用。Mg2+转运体参与到细胞内镁离子的平衡和运输。在拟南芥叶色突变体的筛选过程中，我们获得了一株具有黄色网状叶脉表型的拟南芥突变体var5-1。图位克隆、回复验证以及原生质体瞬时表达显示VAR5编码一个定位于叶绿体的可能的Mg2+转运蛋白AtMGT10。与植物典型的缺镁表型截然相反，var5-1突变体叶脉为黄绿色，叶肉细胞为正常绿色。鉴于已报道的AtMGT10/VAR5完全敲除突变体致死，本研究将利用var5-1突变体这一特殊材料，从var5-1网状叶脉表型的进一步确定、不同AtMGT10/VAR5表达量植物对Mg2+的响应和AtMGT10/VAR5的精细蛋白定位来研究镁离子转运蛋白AtMGT10/VAR5是如何参与植物和叶绿体的镁离子平衡和运输的，这对揭示植物矿质元素代谢和运输机理具有重要的指导意义。

镁离子是植物必需金属离子之一，其平衡影响植物叶绿体的发育。镁离子转运体参与了植物细胞内镁离子的平衡和运输。我们筛选得到了两个等位基因花斑突变体var5-1和var5-2。通过对两个突变等位基因突变位点的鉴定和分子互补实验，证实了VAR5编码一个名为AtMRS2-11/AtMGT10的镁离子转运体CorA家族成员。通过原生质体瞬时表达和生化实验，我们证明AtMRS2-11/AtMGT10/VAR5可能定位于叶绿体被膜。通过双向蓝绿胶非变性电泳和蛋白免疫印迹分析，我们发现AtMRS2-11/AtMGT10/VAR5在叶绿体中形成了大分子量的复合物，其大小明显超过细菌的CorA家族复合物，说明在叶绿体和细菌中CorA镁离子转运复合体的组成不同。我们的发现表明AtMRS2-11/AtMGT10/VAR5在叶绿体发育的组织特异性调控中起着重要作用。