National Key Laboratory of Crop Biology, MOA Key Laboratory of Horticultural Crop Biology and Germplasm Innovation in Huanghuai Region, College of Horticulture Science and Engineering, Shandong Agricultural University, Tai-An, Shandong, China.
Plant Biotechnol J. 2019 Mar;17(3):625-637. doi: 10.1111/pbi.13003. Epub 2018 Oct 2.
Sugars increase with drought stress in plants and accumulate in the vacuole. However, the exact molecular mechanism underlying this process is not clear yet. In this study, protein interaction and phosphorylation experiments were conducted for sucrose transporter and CIPK kinase in apple. The specific phosphorylation site of sucrose transporter was identified with mass spectrometry. Transgenic analyses were performed to characterize their biological function. It was found that overexpression of sucrose transporter gene MdSUT2.2 in apple plants promoted sugar accumulation and drought tolerance. MdSUT2.2 protein was phosphorylated at Ser site in response to drought. A DUALmembrane system using MdSUT2.2 as bait through an apple cDNA library got a protein kinase MdCIPK22. Bimolecular fluorescence complementary (BiFC), pull-down and co-immunoprecipitation (Co-IP) assays further demonstrated that MdCIPK22 interacted with MdSUT2.2. A series of transgenic analysis showed that MdCIPK22 was required for the drought-induced phosphylation at Ser site of MdSUT2.2 protein, and that it enhanced the stability and transport activity of MdSUT2.2 protein. Finally, it was found that MdCIPK22 overexpression promoted sugar accumulation and improved drought tolerance in an MdSUT2.2-dependent manner in transgenic apple plants. MdCIPK22-MdSUT2.2 regulatory module shed light on the molecular mechanism by which plant accumulates sugars and enhances tolerance in response to drought stress.
在植物中，糖分会随着干旱胁迫而增加，并积累在液泡中。然而，这一过程的确切分子机制尚不清楚。在本研究中，对苹果中的蔗糖转运蛋白和 CIPK 激酶进行了蛋白互作和磷酸化实验。利用质谱技术鉴定了蔗糖转运蛋白的特定磷酸化位点。通过转基因分析来表征它们的生物学功能。结果发现，在苹果植株中过表达蔗糖转运蛋白基因 MdSUT2.2 可以促进糖的积累和提高耐旱性。MdSUT2.2 蛋白在干旱胁迫下在 Ser 位点发生磷酸化。利用 MdSUT2.2 作为诱饵的双分子荧光互补（BiFC）实验、pull-down 和 co-immunoprecipitation（Co-IP）实验进一步表明，MdCIPK22 与 MdSUT2.2 相互作用。一系列的转基因分析表明，MdCIPK22 是干旱诱导的 MdSUT2.2 蛋白 Ser 位点磷酸化所必需的，并且增强了 MdSUT2.2 蛋白的稳定性和转运活性。最后发现，在转基因苹果植株中，过表达 MdCIPK22 以依赖于 MdSUT2.2 的方式促进糖的积累并提高耐旱性。MdCIPK22-MdSUT2.2 调控模块阐明了植物在响应干旱胁迫时积累糖和增强耐受性的分子机制。
