王存等《PNAS》2022年

论文题目：Ca2+-dependent phosphorylation of NRAMP1 by CPK21 andCPK23 facilitates manganese uptake and homeostasis inArabidopsis

论文作者：Dali Fu#, Zhenqian Zhang#, Lukas Wallrad#, Zhangqing Wang, Stefanie Höller, ChuanFeng Ju, Ina Schmitz-Thom, Panpan Huang,Lei Wang, Edgar Peiter,Jörg Kudla*, and Cun Wang*

论文摘要：Homeostasis of the essential micronutrient manganese (Mn) is crucially determinedthrough availability and uptake efficiency in all organisms. Mn deficiency of plantsespecially occurs in alkaline and calcareous soils, seriously restricting crop yield. How-ever, the mechanisms underlying the sensing and signaling of Mn availability and conferring regulation of Mn uptake await elucidation. Here, we uncover that Mn depletiontriggers spatiotemporally defined long-lasting Ca2+oscillations inArabidopsisroots.These Ca2+signals initiate in individual cells, expand, and intensify intercellularlyto transform into higher-order multicellular oscillations. Furthermore, through an inter-action screen we identified the Ca2+-dependent protein kinases CPK21 and CPK23 asCa2+signal-decoding components that bring about translation of these signals into regulation of uptake activity of the high-affinity Mn transporter natural resistance associated macrophage proteins 1 (NRAMP1). Accordingly, acpk21/23double mutantdisplays impaired growth and root development under Mn-limiting conditions, whilekinase overexpression confers enhanced tolerance to low Mn supply to plants. In addition, we define Thr498 phosphorylation within NRAMP1 as a pivot mechanisticallydetermining NRAMP1 activity, as revealed by biochemical assays and complementationof yeast Mn uptake andArabidopsis nramp1mutants. Collectively, thesefindings delineate the Ca2+-CPK21/23-NRAMP1 axis as key for mounting plant Mn homeostasis.

锰是植物生长发育的必需微量元素之一。在pH值较高的碱性土壤和钙质土壤，植物可吸收利用的锰减少，导致植物缺锰。目前关于锰转运的研究多集中于转运体的发掘及功能验证，对上游的调控机制却知之甚少。该研究最新发现，低锰胁迫可以触发一种长时间的Ca2+信号振荡，而在根的伸长区（EZ），这种Ca2+信号振荡的响应最为强烈。通过蛋白互作筛选的手段，该研究发现很可能是钙依赖性蛋白激酶CPK21和CPK23与定位在质膜上的锰转运体NRAMP1互作来响应这种长时间的Ca2+信号振荡。同时，通过CRISPR/Cas9技术构建的cpk21/23双突变体在低锰胁迫下表现出敏感表型，而CPK23的过量表达则增强了植物对低锰胁迫的耐受性。通过磷酸化实验，发现CPK21/23可以磷酸化修饰锰转运体NRAMP1蛋白的Ser20和Thr498位点。最后，通过生物化学分析、低锰敏感酵母转运分析和转基因株系的表型分析证明了Thr498对于NRAMP1的锰转运活性至关重要。该研究发现了Ca2+-CPK21/23-NRAMP1调控途径是植物响应低锰胁迫的重要机制（图1）。该研究加深了对Ca2+信号调控植物锰稳态的理解，并为今后碱性土壤上的低锰逆境应答研究和农作物遗传改良奠定了坚实的基础。

图1Ca2+信号调控低锰胁迫的模式图

西北农林科技大学必赢唯一官方网站博士生傅大利，副教授张振乾以及德国明斯特大学的Lukas Wallrad为本文共同第一作者。西北农林科技大学必赢唯一官方网站王存教授为论文通讯作者，德国明斯特大学的Jörg Kudla为共同通讯作者。德国哈勒大学的EdgarPeiter和中国农业科学院生物技术研究所王磊研究员参与了此项研究的指导。该论文得到了国家自然科学基金、中国博士后科学基金和陕西省自然科学基金等项目的资助。

文章链接：https://doi.org/10.1073/pnas.2204574119