Li J, Lee Y R, Assmann S M
Department of Biology, Pennsylvania State University, University Park 16802, USA.
Plant Physiol. 1998 Feb;116(2):785-95. doi: 10.1104/pp.116.2.785.
Increasing evidence suggests that changes in cytosolic Ca2+ levels and phosphorylation play important roles in the regulation of stomatal aperture and as ion transporters of guard cells. However, protein kinases responsible for Ca2+ signaling in guard cells remain to be identified. Using biochemical approaches, we have identified a Ca(2+)-dependent protein kinase with a calmodulin-like domain (CDPK) in guard cell protoplasts of Vicia faba. Both autophosphorylation and catalytic activity of CDPK are Ca2+ dependent. CDPK exhibits a Ca(2+)-induced electrophoretic mobility shift and its Ca(2+)-dependent catalytic activity can be inhibited by the calmodulin antagonists trifluoperazine and N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide. Antibodies to soybean CDPK alpha cross-react with CDPK. Micromolar Ca2+ concentrations stimulate phosphorylation of several proteins from guard cells; cyclosporin A, a specific inhibitor of the Ca(2+)-dependent protein phosphatase calcineurin enhances the Ca(2+)-dependent phosphorylation of several soluble proteins. CDPK from guard cells phosphorylates the K+ channel KAT1 protein in a Ca(2+)-dependent manner. These results suggest that CDPK may be an important component of Ca2+ signaling in guard cells.
越来越多的证据表明，胞质Ca2+水平的变化和磷酸化在气孔孔径调节以及作为保卫细胞的离子转运体中发挥重要作用。然而，负责保卫细胞中Ca2+信号传导的蛋白激酶仍有待确定。我们采用生化方法，在蚕豆保卫细胞原生质体中鉴定出一种具有类钙调蛋白结构域（CDPK）的Ca(2+)依赖性蛋白激酶。CDPK的自身磷酸化和催化活性均依赖于Ca2+。CDPK表现出Ca(2+)诱导的电泳迁移率变化，其Ca(2+)依赖性催化活性可被钙调蛋白拮抗剂三氟拉嗪和N-(6-氨基己基)-5-氯-1-萘磺酰胺抑制。针对大豆CDPKα的抗体与CDPK发生交叉反应。微摩尔浓度的Ca2+可刺激保卫细胞中几种蛋白质的磷酸化；环孢菌素A，一种Ca(2+)依赖性蛋白磷酸酶钙调神经磷酸酶的特异性抑制剂，可增强几种可溶性蛋白质的Ca(2+)依赖性磷酸化。保卫细胞中的CDPK以Ca(2+)依赖性方式使K+通道KAT1蛋白磷酸化。这些结果表明，CDPK可能是保卫细胞中Ca2+信号传导的重要组成部分。
