Graduate School of Agriculture, Osaka Metropolitan University, Sakai, Osaka, 599-8531, Japan.
Division of Evolutionary Biology, National Institute for Basic Biology, Okazaki, Aichi, 444-8585, Japan.
J Plant Res. 2024 Sep;137(5):939-950. doi: 10.1007/s10265-024-01559-8. Epub 2024 Jul 28.
Zinc (Zn) is an essential element for plants. Numerous proteins in different cellular compartments require Zn for their structure and function. Zn can be toxic when it accumulates in high levels in the cytoplasm. Therefore, Zn homeostasis at tissue, cell, and organelle levels is vital for plant growth. A part of the metal tolerance protein (MTP) / Cation Diffusion Facilitator (CDF) transporters functions as Zn transporters, exporting Zn from the cytosol to various membrane compartments. In Arabidopsis thaliana, MTP1, MTP2, MTP3, MTP4, MTP5, and MTP12 are classified as Zn transporters (Zn-CDF). In this study, we systematically analyzed the localization of GFP-fused Zn-CDFs in the leaf epidermal cells of Nicotiana benthamiana. As previously reported, MTP1 and MTP3 were localized to tonoplast, MTP2 to endoplasmic reticulum, and MTP5 to Golgi. In addition, we identified the localization of MTP4 to trans-Golgi Network (TGN). Since MTP4 is specifically expressed in pollen, we analyzed the localization of MTP4-GFP in the Arabidopsis pollen tubes and confirmed that it is in the TGN. We also showed the Zn transport capability of MTP4 in yeast cells. We then analyzed the phenotype of an mtp4 T-DNA insertion mutant under both limited and excess Zn conditions. We found that their growth and fertility were not largely different from the wild-type. Our study has paved the way for investigating the possible roles of MTP4 in metallating proteins in the secretory pathway or in exporting excess Zn through exocytosis. In addition, our system of GFP-fused MTPs will help study the mechanisms for targeting transporters to specific membrane compartments.
锌（Zn）是植物必需的元素。不同细胞区室中的大量蛋白质需要 Zn 来维持其结构和功能。当细胞质中 Zn 积累到高水平时，Zn 可能会有毒。因此，组织、细胞和细胞器水平的 Zn 稳态对植物生长至关重要。一部分金属耐受蛋白（MTP）/阳离子扩散促进剂（CDF）转运蛋白作为 Zn 转运体，将 Zn 从细胞质输出到各种膜区室。在拟南芥中，MTP1、MTP2、MTP3、MTP4、MTP5 和 MTP12 被归类为 Zn 转运体（Zn-CDF）。在这项研究中，我们系统地分析了 GFP 融合的 Zn-CDF 在烟草原生质体叶片表皮细胞中的定位。如前所述，MTP1 和 MTP3 定位于液泡膜，MTP2 定位于内质网，MTP5 定位于高尔基体。此外，我们鉴定了 MTP4 位于反式高尔基网络（TGN）。由于 MTP4 特异性表达于花粉中，我们分析了 MTP4-GFP 在拟南芥花粉管中的定位，并证实它位于 TGN 中。我们还展示了 MTP4 在酵母细胞中的 Zn 转运能力。然后，我们在有限和过量 Zn 条件下分析了 mtp4 T-DNA 插入突变体的表型。我们发现它们的生长和育性与野生型没有太大区别。我们的研究为研究 MTP4 在分泌途径中金属化蛋白或通过胞吐作用输出过量 Zn 的可能作用铺平了道路。此外，我们的 GFP 融合 MTP 系统将有助于研究将转运蛋白靶向特定膜区室的机制。
