Armstrong F, Leung J, Grabov A, Brearley J, Giraudat J, Blatt M R
University of London, Wye College, Kent, England.
Proc Natl Acad Sci U S A. 1995 Oct 10;92(21):9520-4. doi: 10.1073/pnas.92.21.9520.
Abscisic acid (ABA) modulates the activities of three major classes of ion channels--inward- and outward-rectifying K+ channels (IK,in and IK,out, respectively) and anion channels--at the guard-cell plasma membrane to achieve a net efflux of osmotica and stomatal closure. Disruption of ABA sensitivity in wilty abi1-1 mutants of Arabidopsis and evidence that this gene encodes a protein phosphatase suggest that protein (de)-phosphorylation contributes to guard-cell transport control by ABA. To pinpoint the role of ABI1, the abi1-1 dominant mutant allele was stably transformed into Nicotiana benthamiana and its influence on IK,in, IK,out, and the anion channels was monitored in guard cells under voltage clamp. Compared with guard cells from wild-type and vector-transformed control plants, expression of the abi1-1 gene was associated with 2- to 6-fold reductions in IK,out and an insensitivity of both IK,in and IK,out to 20 microM ABA. In contrast, no differences between control and abi1-1 transgenic plants were observed in the anion current or its response to ABA. Parallel measurements of intracellular pH (pHi) using the fluorescent dye 2',7'-bis(2-carboxyethyl)-5-(and -6)-carboxyfluorescein (BCECF) in every case showed a 0.15- to 0.2-pH-unit alkalinization in ABA, demonstrating that the transgene was without effect on the pHi signal that mediates in ABA-evoked K+ channel control. In guard cells from the abi1-1 transformants, normal sensitivity of both K+ channels to and stomatal closure in ABA was recovered in the presence of 100 microM H7 and 0.5 microM staurosporine, both broad-range protein kinase antagonists. These results demonstrate an aberrant K+ channel behavior--including channel insensitivity to ABA-dependent alkalinization of pHi--as a major consequence of abi1-1 action and implicate AB11 as part of a phosphatase/kinase pathway that modulates the sensitivity of guard-cell K+ channels to ABA-evoked signal cascades.
脱落酸（ABA）调节保卫细胞质膜上三类主要离子通道的活性，即内向整流和外向整流钾通道（分别为IK,in和IK,out）以及阴离子通道，以实现渗透物的净外流和气孔关闭。拟南芥萎蔫abi1 - 1突变体中ABA敏感性的破坏以及该基因编码一种蛋白磷酸酶的证据表明，蛋白质（去）磷酸化参与了ABA对保卫细胞运输的控制。为了明确ABI1的作用，将abi1 - 1显性突变等位基因稳定转化到本氏烟草中，并在电压钳制下监测其对保卫细胞中IK,in、IK,out和阴离子通道的影响。与野生型和载体转化对照植物的保卫细胞相比，abi1 - 1基因的表达与IK,out降低2至6倍以及IK,in和IK,out对20微摩尔ABA不敏感有关。相反，对照植物和abi1 - 1转基因植物在阴离子电流或其对ABA的反应方面未观察到差异。在每种情况下，使用荧光染料2',7'-双（2 - 羧乙基）-5-（和-6）-羧基荧光素（BCECF）对细胞内pH（pHi）进行平行测量，结果显示ABA处理后pHi有0.15至0.2个pH单位的碱化，这表明转基因对介导ABA诱导的钾通道控制的pHi信号没有影响。在abi1 - 1转化体的保卫细胞中，在存在100微摩尔H7和0.5微摩尔星形孢菌素（两种广谱蛋白激酶拮抗剂）的情况下，两种钾通道对ABA的正常敏感性和气孔关闭得以恢复。这些结果表明，异常的钾通道行为，包括通道对ABA诱导的pHi碱化不敏感，是abi1 - 1作用的主要后果，并暗示AB11是磷酸酶/激酶途径的一部分，该途径调节保卫细胞钾通道对ABA诱导信号级联反应的敏感性。
