College of Horticulture, China Agricultural University, Beijing, China.
Department of resources and environment, Beijing University of Agriculture, Beijing, China.
Plant Biotechnol J. 2018 Mar;16(3):737-748. doi: 10.1111/pbi.12824. Epub 2017 Oct 12.
Potassium (K ), an abundant cation in plant cells, is important in fruit development and plant resistance. However, how cellular K is directed by potassium channels in fruit development and quality formation of strawberry (Fragaria × ananassa) is not yet fully clear. Here, a two-pore K (TPK) channel gene in strawberry, FaTPK1, was cloned using reverse transcription-PCR. A green fluorescent protein subcellular localization analysis showed that FaTPK1 localized in the vacuole membrane. A transcription analysis indicated that the mRNA expression level of FaTPK1 increased rapidly and was maintained at a high level in ripened fruit, which was coupled with the fruit's red colour development, suggesting that FaTPK1 is related to fruit quality formation. The down- and up-regulation of the FaTPK1 mRNA expression levels using RNA interference and overexpression, respectively, inhibited and promoted fruit ripening, respectively, as demonstrated by consistent changes in firmness and the contents of soluble sugars, anthocyanin and abscisic acid, as well as the transcript levels of ripening-regulated genes PG1 (polygalacturonase), GAL6 (beta-galactosidase), XYL2 (D-xylulose reductase), SUT1 (sucrose transporter), CHS (chalcone synthase) and CHI (chalcone flavanone isomerase). Additionally, the regulatory changes influenced fruit resistance to Botrytis cinerea. An isothermal calorimetry analysis showed that the Escherichia coli-expressed FaTPK1 recombinant protein could bind K with a binding constant of 2.1 × 10 m and a dissociation constant of 476 μm. Thus, the strawberry TPK1 is a ubiquitously expressed, tonoplast-localized two-pore potassium channel that plays important roles in fruit ripening and quality formation.
钾（K）是植物细胞中丰富的阳离子，对果实发育和植物抗性很重要。然而，钾通道如何在草莓（Fragaria × ananassa）果实发育和品质形成中指导细胞内的 K 尚不完全清楚。在这里，使用反转录-PCR 克隆了草莓中的一个双孔钾（TPK）通道基因 FaTPK1。绿色荧光蛋白亚细胞定位分析表明 FaTPK1 定位于液泡膜上。转录分析表明，FaTPK1 的 mRNA 表达水平在成熟果实中迅速增加，并保持在高水平，与果实的红色发育相耦合，表明 FaTPK1 与果实品质形成有关。使用 RNA 干扰和过表达分别下调和上调 FaTPK1 mRNA 表达水平，分别抑制和促进果实成熟，通过硬度和可溶性糖、花青素和脱落酸的含量以及成熟调控基因 PG1（多聚半乳糖醛酸酶）、GAL6（β-半乳糖苷酶）、XYL2（D-木酮糖还原酶）、SUT1（蔗糖转运蛋白）、CHS（查尔酮合酶）和 CHI（查尔酮黄酮异构酶）的转录水平的一致变化来证明。此外，调控变化影响了果实对灰葡萄孢的抗性。等温量热法分析表明，大肠杆菌表达的 FaTPK1 重组蛋白可以与 K 结合，结合常数为 2.1×10 -6 M，解离常数为 476 μm。因此，草莓 TPK1 是一种普遍表达的、液泡定位的双孔钾通道，在果实成熟和品质形成中发挥重要作用。
