Yu Xiang-Chun, Li Mei-Jun, Gao Gui-Feng, Feng Hai-Zhong, Geng Xue-Qing, Peng Chang-Cao, Zhu Sai-Yong, Wang Xiao-Jing, Shen Yuan-Yue, Zhang Da-Peng
China State Key Laboratory of Plant Physiology and Biochemistry, China Agricultural University, 100094 Beijing, China.
Plant Physiol. 2006 Feb;140(2):558-79. doi: 10.1104/pp.105.074971. Epub 2006 Jan 11.
It has been demonstrated that calcium plays a central role in mediating abscisic acid (ABA) signaling, but many of the Ca2+-binding sensory proteins as the components of the ABA-signaling pathway remain to be elucidated. Here we identified, characterized, and purified a 58-kD ABA-stimulated calcium-dependent protein kinase from the mesocarp of grape berries (Vitis vinifera x Vitis labrusca), designated ACPK1 (for ABA-stimulated calcium-dependent protein kinase1). ABA stimulates ACPK1 in a dose-dependent manner, and the ACPK1 expression and enzyme activities alter accordantly with the endogenous ABA concentrations during fruit development. The ABA-induced ACPK1 stimulation appears to be transient with a rapid effect in 15 min but also with a slow and steady state of induction after 60 min. ABA acts on ACPK1 indirectly and dependently on in vivo state of the tissues. Two inactive ABA isomers, (-)-2-cis, 4-trans-ABA and 2-trans, 4-trans-(+/-)-ABA, are ineffective for inducing ACPK1 stimulation, revealing that the ABA-induced effect is stereo specific to physiological active (+)-2-cis, 4-trans-ABA. The other phytohormones such as auxin indoleacetic acid, gibberellic acid, synthetic cytokinin N-benzyl-6-aminopurine, and brassinolide are also ineffective in this ACPK1 stimulation. Based on sequencing of the two-dimensional electrophoresis-purified ACPK1, we cloned the ACPK1 gene. The ACPK1 is expressed specifically in grape berry covering a fleshy portion and seeds, and in a developmental stage-dependent manner. We further showed that ACPK1 is localized in both plasma membranes and chloroplasts/plastids and positively regulates plasma membrane H+-ATPase in vitro, suggesting that ACPK1 may be involved in the ABA-signaling pathway.
已有研究表明，钙在介导脱落酸（ABA）信号传导中起核心作用，但作为ABA信号通路组成部分的许多Ca2+结合传感蛋白仍有待阐明。在此，我们从葡萄浆果（Vitis vinifera x Vitis labrusca）的中果皮中鉴定、表征并纯化了一种58-kD的ABA刺激的钙依赖性蛋白激酶，命名为ACPK1（ABA刺激的钙依赖性蛋白激酶1）。ABA以剂量依赖性方式刺激ACPK1，并且ACPK1的表达和酶活性在果实发育过程中与内源ABA浓度一致变化。ABA诱导的ACPK1刺激似乎是短暂的，在15分钟内有快速效应，但在60分钟后也有缓慢且稳定的诱导状态。ABA间接作用于ACPK1，且依赖于组织的体内状态。两种无活性的ABA异构体，（-）-2-顺式，4-反式-ABA和2-反式，4-反式-（+/-）-ABA，对诱导ACPK1刺激无效，这表明ABA诱导的效应对于生理活性（+）-2-顺式，4-反式-ABA具有立体特异性。其他植物激素，如生长素吲哚乙酸、赤霉素、合成细胞分裂素N-苄基-6-氨基嘌呤和油菜素内酯，在这种ACPK1刺激中也无效。基于二维电泳纯化的ACPK1的测序，我们克隆了ACPK1基因。ACPK1在葡萄浆果中特异性表达，覆盖肉质部分和种子，并以发育阶段依赖性方式表达。我们进一步表明，ACPK1定位于质膜和叶绿体/质体中，并在体外正向调节质膜H+-ATP酶，这表明ACPK1可能参与ABA信号通路。
