Ca2+-dependent successive phosphorylation of vacuolar transporter MTP8 by CBL2/3-CIPK3/9/26 and CPK5 is critical for manganese homeostasis in Arabidopsis
Ca2+-dependent successive phosphorylation of vacuolar transporter MTP8 by CBL2/3-CIPK3/9/26 and CPK5 is critical for manganese homeostasis in Arabidopsis
复制标题
CBL2/3-CIPK3/9/26 和 CPK5 对液泡转运蛋白 MTP8 的 Ca~(2 ) 依赖性连续磷酸化对于拟南芥中的锰稳态至关重要
DOI：
10.1016/j.molp.2021.11.012
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发表时间：
2022-03-07
期刊：
影响因子：
27.5
通讯作者：
Wang, Cun
中科院分区：
文献类型：
--
作者：
Ju, Chuanfeng;Zhang, Zhenqian;Wang, Cun
锰（Mn）是所有生物体必需的微量营养素。然而，在酸性或渍水土壤中可能出现的过量锰供应会对植物生理和发育产生毒害作用。尽管多种锰转运蛋白家族已被鉴定，但我们对这些转运蛋白如何被调控以维持和调节植物体内的锰稳态仅有初步了解。在此，我们证明了两种类钙调神经磷酸酶B蛋白（CBL2/3）及其相互作用的激酶（CIPK3/9/26）是植物锰稳态的关键调节因子。缺乏CBL2和CBL3或其相互作用的蛋白激酶CIPK3/9/26的拟南芥突变体表现出极高的锰耐受性。有趣的是，CIPK3/9/26与液泡膜定位的锰和铁（Fe）转运蛋白MTP8相互作用并主要在丝氨酸35位点将其磷酸化，该位点在不同物种的MTP8蛋白中是保守的。酵母中的锰转运互补实验结合多种生理实验表明，CBL - CIPK介导的MTP8磷酸化负向调节其从细胞质到液泡的转运活性。此外，我们发现MTP8的顺序磷酸化，首先由钙依赖性蛋白激酶CPK5在丝氨酸31/32位点磷酸化，随后由CIPK26在丝氨酸35位点磷酸化，为锰转运的差异调节提供了一种激活/失活的微调机制。总之，我们的研究结果确定了一种在锰供应波动条件下动态调节锰稳态的两级钙控制机制。
Manganese (Mn) is an essential micronutrient for all living organisms. However, excess Mn supply that can occur in acid or waterlogged soils has toxic effects on plant physiology and development. Although a variety of Mn transporter families have been characterized, we have only a rudimentary understanding of how these transporters are regulated to uphold and adjust Mn homeostasis in plants. Here, we demonstrate that two calcineurin-B-like proteins, CBL2/3, and their interacting kinases, CIPK3/9/26, are key regulators of plant Mn homeostasis. Arabidopsis mutants lacking CBL2 and 3 or their interacting protein kinases CIPK3/9/26 exhibit remarkably high Mn tolerance. Intriguingly, CIPK3/9/26 interact with and phosphorylate the tonoplast-localized Mn and iron (Fe) transporter MTP8 primarily at Ser35, which is conserved among MTP8 proteins from various species. Mn transport complementation assays in yeast combined with multiple physiological assays indicate that CBL-CIPK-mediated phosphorylation of MTP8 negatively regulates its transport activity from the cytoplasm to the vacuole. Moreover, we show that sequential phosphorylation of MTP8, initially at Ser31/32 by the calcium-dependent protein kinase CPK5 and subsequently at Ser35 by CIPK26, provides an activation/deactivation fine-tuning mechanism for differential regulation of Mn transport. Collectively, our findings define a two-tiered calcium-controlled mechanism for dynamic regulation of Mn homeostasis under conditions of fluctuating Mn supply.