保卫细胞中的微丝骨架

微丝骨架存在于多种植物的保卫细胞中,周质微丝骨架的排列和结构是动态的。越来越多的证据表明保卫细胞中的微丝骨架可作为信号调节物,对气孔的启闭运动起着重要的调控作用。本文综述了保卫细胞微丝骨架的标记方法、结构,以及其在气孔运动中的功能和作用机制的最新研究进展。     

微丝骨架在植物细胞信号转导中的作用

文章从小G蛋白、离子浓度和肌醇磷脂信号系统等方面阐述植物细胞微丝骨架与细胞信号转导的关系.     

蚕豆水孔蛋白可能参与微丝骨架对气孔运动的调节

水孔蛋白的抑制剂HgCl2可明显抑制壳梭孢菌素（FC）和微丝骨架的解聚剂细胞松弛素D（CD）对蚕豆保卫细胞原生质体膨胀的诱导作用,而对微丝骨架的稳定剂鬼笔环肽（phalloidin）的抑制作用影响不明显。这表明水孔蛋白可能介导了FC和微丝骨架对气孔运动的调节。     

GFP-mTn融合蛋白对蓝猪耳生活细胞微丝骨架的标记

用农杆菌介导法将嵌合基因GFP-mTn(mTn是微丝结合蛋白Talin的微丝结合域,可以显示活体细胞中微丝的结构)导入蓝猪耳.经激光共聚焦显微镜观察了转基因植株的各种不同组织中融合蛋白的表达和分布情况.在叶片的表皮细胞、保卫细胞、根部的皮层细胞中有融合蛋白的不同程度表达.但仅在保卫细胞中微丝标记状况良好,显示基因表达的组织特异性.经光诱导处于开放态的气孔的保卫细胞微丝呈网状结构,在细胞内无规则分布;经黑暗诱导处于关闭态的气孔保卫细胞中微丝束沿保卫细胞纵轴排列,呈卷曲状分布,并观察到螺旋和环状的微丝结构.在转基因植株的其他部位,例如茎表皮细胞、根毛细胞和花粉粒中,未检测到目的基因的表达.本研究获得的转基因植株为研究气孔运动过程中微丝动态变化提供了有用的材料.     

保卫细胞钙信号的研究进展

钙(Ca^2 )是多种信号途径的第二信使。Ca^2 成像技术的成熟和发展为显示保卫细胞胞质Ca^2 浓度([Ca^2 ]cyt)的分布及外界刺激引起[Ca^2 ]cyt的变化模式提供了很好的研究工具,关于细胞内外Ca^2 库释放Ca^2 的机制也有了较清楚的认识。拟南芥突变体的研究使Ca^2 信号上游分子及其排序更加明确,[Ca^2 ]cyt增加下游的磷酸化和去磷酸化过程也是气孔关闭必需的生理过程。     

植物微管和微丝骨架研究的进展

从以上叙述的资料中可以看出,近年来在植物微管蛋白的分离及其化学性质、微管的组织中心、微管的异质性、微丝的分布,以及微管和微丝骨架的功能及基因调节等方面的研究取得不少新的进展;特别是从植物中直接分离微管蛋白取得成功、以及微管蛋白异型、微管冷稳定性与植物抗寒性的关系及微丝分布广泛性等的发现,对植物细胞骨架的进一步研究具有重要意义。     

拟南芥保卫细胞微丝骨架的解聚可能参与了细胞外钙调素诱导的气孔关闭

细胞外钙调素(CaM)在植物的许多生理活动中都执行着重要功能, 但它对气孔运动的作用及其调控机制, 人们了解的很少. 以模式植物拟南芥为材料, 研究了细胞外CaM在保卫细胞壁上的存在及其对气孔运动的调控机制. 结果表明, 拟南芥保卫细胞壁中存在有分子量为17 kD的CaM, 并应用W7-琼脂糖和CaM抗血清初步证明了保卫细胞壁中存在的CaM可能具有促进气孔关闭和抑制气孔开放的作用. 在应用外源CaM诱导气孔关闭的实验中, 保卫细胞微丝骨架由长而呈辐射状分布的聚合态逐步解聚, 气孔开度也随着降低. 药理学实验结果表明, 保卫细胞微丝骨架的解聚能明显地促进外源CaM诱导的气孔关闭, 而微丝骨架的聚合则抑制这一过程. 研究结果还表明, 外源CaM能诱导保卫细胞[Ca²⁺]_cyt升高; 当使用Ca²⁺螯合剂EGTA时, 外源CaM诱导的[Ca²⁺]_cyt升高和气孔关闭运动均受到抑制. 为此推测细胞外CaM可能是通过诱导保卫细胞[Ca²⁺]_cyt升高, 导致微丝骨架的解聚, 进而促进气孔的关闭运动.     

保卫细胞的ABA信号转导

植物激素脱落酸（ABA）调节植物体多种生理过程,尤其在一些逆境条件下,植物体中ABA大量合成,诱导气孔关闭,从而有效地调控植物体内的水分平衡,尽管人们对ABA诱导气孔关闭作用已得到共识,但有关信号转导的细节还很不清楚,该文简要介绍了研究气孔保卫细胞信号转导途径的相关技术以及与ABA信号转导直接相关的ABA受体,第二信使,蛋白质磷酸化和离子通道调节等方面的最新研究进展,并在前人研究工作的基础上,勾画出气孔保卫细胞ABA,H2O2的信号转导模式图。     

GFP-mTn融合蛋白对蓝猪耳生活细胞微丝骨架的标记

用农杆菌介导法将嵌合基因GFP-mTn(mTn是微丝结合蛋白Talin的微丝结合域,可以显示活体细胞中微丝的结构)导入蓝猪耳。经激光共聚焦显微镜观察了转基因植株的各种不同组织中融合蛋白的表达和分布情况。在叶片的表皮细胞、保卫细胞、根部的皮层细胞中有融合蛋白的不同程度表达。但仅在保卫细胞中微丝标记状况良好,显示基因表达的组织特异性。经光诱导处于开放态的气孔的保卫细胞微丝呈网状结构,在细胞内无规则分布;经黑暗诱导处于关闭态的气孔保卫细胞中微丝束沿保卫细胞纵轴排列,呈卷曲状分布,并观察到螺旋和环状的微丝结构。在转基因植株的其他部位,例如茎表皮细胞、根毛细胞和花粉粒中,未检测到目的基因的表达。本研究获得的转基因植株为研究气孔运动过程中微丝动态变化提供了有用的材料。     

Hormone interactions in stomatal function

Research in recent years on the biology of guard cells has shown that these specialized cells integrate both extra- and intra-cellular signals in the control of stomatal apertures. Among the phytohormones, abscisic acid (ABA) is one of the key players regulating stomatal function. In addition, auxin, cytokinin, ethylene, brassinosteroids, jasmonates, and salicylic acid also contribute to stomatal aperture regulation. The interaction of multiple hormones can serve to determine the size of stomatal apertures in a condition-specific manner. Here, we discuss the roles of different phytohormones and the effects of their interactions on guard cell physiology and function.     

Early ABA Signaling Events in Guard Cells

The plant hormone abscisic acid (ABA) regulates a wide variety of plant physiological and developmental processes, particularly responses to environmental stress, such as drought. In response to water deficiency, plants redistribute foliar ABA and/or upregulate ABA synthesis in roots, leading to roughly a 30-fold increase in ABA concentration in the apoplast of stomatal guard cells. The elevated ABA triggers a chain of events in guard cells, causing stomatal closure and thus preventing water loss. Although the molecular nature of ABA receptor(s) remains unknown, considerable progress in the identification and characterization of its downstream signaling elements has been made by using combined physiological, biochemical, biophysical, molecular, and genetic approaches. The measurable events associated with ABA-induced stomatal closure in guard cells include, sequentially, the production of reactive oxygen species (ROS), increases in cytosolic free Ca²⁺ levels ([Ca²⁺]_i), activation of anion channels, membrane potential depolarization, cytosolic alkalinization, inhibition of K⁺ influx channels, and promotion of K⁺ efflux channels. This review provides an overview of the cellular and molecular mechanisms underlying these ABA-evoked signaling events, with particular emphasis on how ABA triggers an “electronic circuitry” involving these ionic components.     

细胞迁移中微丝微管的变化及其信号转导通路研究进展

细胞迁移是一个多步骤协调的过程。在此过程中,细胞骨架蛋白微丝和微管的动态变化提供了细胞运动的主要动力。而迁移的过程又被多种信号分子组成的复杂的网络所调控。本文主要综述了细胞迁移中微丝和微管的变化以及调控此种变化的分子机制。     

拟南芥保卫细胞中特异性表达基因启动子片段基序分析

将16个在拟南芥保卫细胞中进行特异性表达和8个非特异表达基因的转录起始密码子ATG上游-500bp的启动子序列在PLACE上进行分析,认为AAAAG基序和TAAAG基序控制某些基因在保卫细胞中进行特异性表达,但分析结果显示有的保卫细胞特异表达启动子的序列中却不含有这两个基序,说明还要其它类型的未知基序控制着保卫细胞中的特异表达,这一理论分析为进一步的实验研究提供了理论依据。     

Intermediate Filaments Play a Pivotal Role in Regulating Cell Architecture and Function

Intermediate filaments (IFs) are composed of one or more members of a large family of cytoskeletal proteins, whose expression is cell- and tissue type-specific. Their importance in regulating the physiological properties of cells is becoming widely recognized in functions ranging from cell motility to signal transduction. IF proteins assemble into nanoscale biopolymers with unique strain-hardening properties that are related to their roles in regulating the mechanical integrity of cells. Furthermore, mutations in the genes encoding IF proteins cause a wide range of human diseases. Due to the number of different types of IF proteins, we have limited this short review to cover structure and function topics mainly related to the simpler homopolymeric IF networks composed of vimentin, and specifically for diseases, the related muscle-specific desmin IF networks.     

Evidences for Regulation of the Inward K+ channels by CDPK in Vicia faba Guard Cells

Regulation of the inward K+ -channels in the guard cell plasma membranes plays impotant roles in regulation of stomatal movement in responses to exogenous and endogenous signals. It is well-known that elevation of cytosolic Ca2+ in guard cells inactivates these inward K + channels, and consequently inhibits stomatal opening or induces stomatal closing, yet the downstream molecular mechanism for the Ca2 + -mediated inhibition of the inward K+ channels remains unknown. The calmodulin-like domain protein kinases (CDPKs) have been identified as an unique group of protein kinases in higher plant cells. As a downstream regulator, CDPK may play roles in mediating Ca2+ regulation on the inward K+ -channels in stomatal guard cells. The authors have applied the patchclamp technique to investigate if CDPK be involved in the regulation of the inward K+ -channels in Vicia faba guard cells by cytosolic Ca2+ . The presence of the 1.5 μmol/L intracellular Ca2 + result-ed in inhibition of the inward K+ channel activity by 60%, while the addition of purified CDPK from the cytoplasmic side resulted in greater inhibition than Ca2+ alone. Histone Ⅲ-S and protamine, which is the substrate and substrate competitive inhibitor of CDPKs respectively, completely reversed the Ca2+ -induced inhibition of the inward K+ channel activities. These results are the first reported evidences for that CDPKs are involved in the Ca2+ -mediated inward K+ -channel regulation in guard cells.     

保卫细胞碳代谢与气孔运动

作为气孔运动渗透调节的代谢基础 ,气孔保卫细胞的碳代谢有特殊的调控机理。本文介绍了气孔保卫细胞中参与碳代谢的主要酶的特性及调控特点 ,特别是保卫细胞叶绿体中催化苹果酸形成的PEP羧化酶 ,其磷酸化和去磷酸化参与了保卫细胞信号传递。保卫细胞碳代谢调控在气孔运动调节中的作用 ,并讨论了保卫细胞碳代谢与能量代谢的关系     

Extensive Distribution of Acetylcholinesterase in Guard Cells of Vicia faba

Acetylcholinesterase, an enzyme responsible for hydrolyzing of acetylcholine to choline and acetic acid residues, is detected in the guard cell protoplasts. Extensive acetylcholinesterase activity has been found in the guard cell protoplasts as compared with the mesophyll cell protoplasts. Moreover, light could stimulate the enzyme activity. Localization of acetylcholinesterase in the stomata of Vicia faba L. was undertaken using Karnovsky and Roots cytochemical method. It was found that in the stomata of this plant products of acetylcholinesterase enzymatic reaction mainly appeared in the outer side of the guard cell ventral wall and inner wall. When the staining time was prolonged, products of acetylcholinesterase enzymatic reaction could also be found in the ventral and inner wall of the guard cells. In addition, more extensive product of enzymatic reaction was observed in the opened stomata than in the closed stomata. It was assumed that acetylcholineaterase may participate in the regulation of stomatal movement by hydrolyzing acetylcholine around the stomata.     

External Protons Enhance the Activity of the Hyperpolarization-activated K Channels in Guard Cell Protoplasts of Vicia faba

Hyperpolarization-activated K channels (K _H channels) in the plasmalemma of guard cells operate at apoplastic pH range of 5 to over 7. Using patch clamp in a whole-cell mode, we characterized the effect of varying the external pH between 4.4–8.1 on the activity of the K _H channels in isolated guard cell protoplasts from Vicia faba leaves. Acidification from pH 5.5 to 4.4 increased the macroscopic conductance of the K _H channels by 30–150% while alkalinization from pH 5.5 to 8.1 decreased it only by roughly 15%. The voltage-independent maximum cell conductance, increased by ∼60% between pH 8.1 and 4.4 with an apparent pK _a of 5.3, most likely owing to the increased availability of channels. Voltage-dependent gating was affected only between pH 5.5 and 4.4. Acidification in this range shifted the voltage-dependent open probability by over 10 mV. We interpret this shift as an increase of the electrical field sensed by the gating subunits caused by the protonation of external negative surface charges. Within the framework of a surface charge model the mean spacing of these charges was ∼30 ? and their apparent dissociation constant was 10^−4.6. The overall voltage sensitivity of gating was not altered by pH changes. In a subgroup of protoplasts analyzed within the framework of a Closed-Closed-Open model, the effect of protons on gating was limited to shifting of the voltage-dependence of all four transition rate constants. Received: 26 April 1996/Revised: 29 June 1996