INO80参与拟南芥气孔数量调控的分子机制

气孔是植物表皮特有的结构,参与到植物的呼吸、蒸腾作用等多种生理活动.植物通过气孔与大气进行气体交换.INO80是一类保守的染色质重塑因子,可以与组蛋白变异体H2A.Z结合.与野生型相比,拟南芥ino80缺失突变体对于渗透压的敏感度及其失水率都有明显提高.与此相应的是,ino80缺失突变体植物表现出气孔明显增多的表型.bHLH转录因子家族中的SPCH和MUTE对于气孔的发育起到正调控的作用.RT-PCR检测显示,在ino80突变体的背景下,气孔正调控基因SPCH和MUTE的表达量都显著上调.ChIP-PCR实验显示,相对于野生型,H2A.Z在SPCH和MUTE上染色质的分布在ino80缺失突变体内下调.我们的工作表明,在拟南芥中,INO80可能通过调控气孔正调控基因染色质区域组蛋白变体H2A.Z的分布影响靶基因的转录水平,进而改变气孔数量以及植物的相应生理指标.     

Sphingolipid signalling in Arabidopsis guard cells involves heterotrimeric G proteins

In animals, the sphingolipid metabolite sphingosine-1-phosphate (S1P) functions as both an intracellular messenger and an extracellular ligand for G-protein-coupled receptors of the S1P receptor family, regulating diverse biological processes ranging from cell proliferation to apoptosis. Recently, it was discovered in plants that S1P is a signalling molecule involved in abscisic acid (ABA) regulation of guard cell turgor. Here we report that the enzyme responsible for S1P production, sphingosine kinase (SphK), is activated by ABA in Arabidopsis thaliana, and is involved in both ABA inhibition of stomatal opening and promotion of stomatal closure. Consistent with this observation, inhibition of SphK attenuates ABA regulation of guard cell inward K(+) channels and slow anion channels, which are involved in the regulation of stomatal pore size. Surprisingly, S1P regulates stomatal apertures and guard cell ion channel activities in wild-type plants, but not in knockout lines of the sole prototypical heterotrimeric G-protein alpha-subunit gene, GPA1 (refs 5, 6, 7-8). Our results implicate heterotrimeric G proteins as downstream elements in the S1P signalling pathway that mediates ABA regulation of stomatal function, and suggest that the interplay between S1P and heterotrimeric G proteins represents an evolutionarily conserved signalling mechanism.     

Guard cell abscisic acid signalling and engineering drought hardiness in plants

Guard cells are located in the epidermis of plant leaves, and in pairs surround stomatal pores. These control both the influx of CO2 as a raw material for photosynthesis and water loss from plants through transpiration to the atmosphere. Guard cells have become a highly developed system for dissecting early signal transduction mechanisms in plants. In response to drought, plants synthesize the hormone abscisic acid, which triggers closing of stomata, thus reducing water loss. Recently, central regulators of guard cell abscisic acid signalling have been discovered. The molecular understanding of the guard cell signal transduction network opens possibilities for engineering stomatal responses to control CO2 intake and plant water loss.     

Nitric oxide involved in signal transduction of Jasmonic acid-induced stomatal closure of Vicia faba L.

Nitric oxide (NO) and Jasmonic acid (JA) are two key signaling molecules involved in many and diverse biological pathways in plants. Growing evidence suggested that NO signaling interacts with JA signaling. In this work, Our experiment showed that NO exists in guard cell of Vicia faba L., and NO is involved in signal transduction of JAinduced stomata closuring: ( i ) JA enhances NO synthesis in guard cell; ( ii ) both JA and NO induced stomatal closure, and had dose response to their effects; ( iU ) there are synergetic correlation between JA and lower NO concentration in regulation of stomatal movement; (iV) JA-induced stomatal closure was largely prevented by 2-phenyl-4,4,5,5-tetramethylimidazoline-l-oxyl-3-oxide (PTIO), a specific NO scavenger. An inhibitor of NO synthase (NOS) in mammalian cells, N^G-nitro-L-Arg-methyl eater (L-NAME) also inhibits plant NOS, repressing JA-induced NO generation and JA-induced stomatal closure. We presumed that NO mainly comes from NOS after JA treatment.     

Asymmetric cell divisions promote Notch-dependent epidermal differentiation

Stem and progenitor cells use asymmetric cell divisions to balance proliferation and differentiation. Evidence from invertebrates shows that this process is regulated by proteins asymmetrically distributed at the cell cortex during mitosis: Par3-Par6-aPKC, which confer polarity, and Gα(i)-LGN/AGS3-NuMA-dynein/dynactin, which govern spindle positioning. Here we focus on developing mouse skin, where progenitor cells execute a switch from symmetric to predominantly asymmetric divisions concomitant with stratification. Using in vivo skin-specific lentiviral RNA interference, we investigate spindle orientation regulation and provide direct evidence that LGN (also called Gpsm2), NuMA and dynactin (Dctn1) are involved. In compromising asymmetric cell divisions, we uncover profound defects in stratification, differentiation and barrier formation, and implicate Notch signalling as an important effector. Our study demonstrates the efficacy of applying RNA interference in vivo to mammalian systems, and the ease of uncovering complex genetic interactions, here to gain insights into how changes in spindle orientation are coupled to establishing proper tissue architecture during skin development.     

cdc2 links the Drosophila cell cycle and asymmetric division machineries

Asymmetric cell divisions can be mediated by the preferential segregation of cell-fate determinants into one of two sibling daughters. In Drosophila neural progenitors, Inscuteable, Partner of Inscuteable and Bazooka localize as an apical cortical complex at interphase, which directs the apical-basal orientation of the mitotic spindle as well as the basal/cortical localization of the cell-fate determinants Numb and/or Prospero during mitosis. Although localization of these proteins shows dependence on the cell cycle, the involvement of cell-cycle components in asymmetric divisions has not been demonstrated. Here we show that neural progenitor asymmetric divisions require the cell-cycle regulator cdc2. By attenuating Drosophila cdc2 function without blocking mitosis, normally asymmetric progenitor divisions become defective, failing to correctly localize asymmetric components during mitosis and/or to resolve distinct sibling fates. cdc2 is not necessary for initiating apical complex formation during interphase; however, maintaining the asymmetric localization of the apical components during mitosis requires Cdc2/B-type cyclin complexes. Our findings link cdc2 with asymmetric divisions, and explain why the asymmetric localization of molecules like Inscuteable show cell-cycle dependence.     

pH induced elastic modulus of guard cell wall in stomatal movement

Guard cell wall properties are important in stomatal movement. Previous research focused on the structure and anatomy of guard cell walls, but little is known about the physical changes that take place within the walls during stomatal opening and closure. In this work, we investigate the volumetric elastic modulus (ε) of the guard cell wall at different pH values during stomatal opening in Vicia faba epidermal strips using a cell pressure probe. The volumetric elastic modulus of the guard cell wall decrease...     

A defined range of guard cell calcium oscillation parameters encodes stomatal movements.

Oscillations in cytosolic calcium concentration ([Ca2+]cyt) are central regulators of signal transduction cascades, although the roles of individual [Ca2+]cyt oscillation parameters in regulating downstream physiological responses remain largely unknown. In plants, guard cells integrate environmental and endogenous signals to regulate the aperture of stomatal pores and [Ca2+]cyt oscillations are a fundamental component of stomatal closure. Here we systematically vary [Ca2+]cyt oscillation parameters in Arabidopsis guard cells using a 'calcium clamp' and show that [Ca2+]cyt controls stomatal closure by two mechanisms. Short-term 'calcium-reactive' closure occurred rapidly when [Ca2+]cyt was elevated, whereas the degree of long-term steady-state closure was 'calcium programmed' by [Ca2+]cyt oscillations within a defined range of frequency, transient number, duration and amplitude. Furthermore, in guard cells of the gca2 mutant, [Ca2+]cyt oscillations induced by abscisic acid and extracellular calcium had increased frequencies and reduced transient duration, and steady-state stomatal closure was abolished. Experimentally imposing [Ca2+]cyt oscillations with parameters that elicited closure in the wild type restored long-term closure in gca2 stomata. These data show that a defined window of guard cell [Ca2+]cyt oscillation parameters programs changes in steady-state stomatal aperture.     

Reversible inactivation of K+ channels of Vicia stomatal guard cells following the photolysis of caged inositol 1,4,5-trisphosphate

RECENT investigations suggest that cytoplasmic D-myo-inositol 1,4,5-trisphosphate (InsP3) functions as a second messenger in plants, as in animals, coupling environmental and other stimuli to intracellular Ca2+ release. Cytoplasmic levels of InsP3 and the turnover of several probable precursors in plants are affected by physiological stimuli--including light, osmotic stress and the phytohormone indoleacetic acid--and InsP3 activates Ca2+ channels and Ca2+ flux across plant vacuolar and microsomal membranes. Complementary data also link changes in cytoplasmic free Ca2+ to several physiological responses, notably in guard cells which regulate gas exchange through the stomatal pores of higher plant leaves. Recent evidence indicates that guard cell K+ channels and, hence, K+ flux for stomatal movements may be controlled by cytoplasmic Ca2+. So far, however, direct evidence of a role for InsP3 in signalling in plants has remained elusive. Here we report that InsP3 released from an inactive, photolabile precursor, the P5-1-(2-nitrophenyl)ethyl ester of InsP3 (caged InsP3) reversibly inactivates K+ channels thought to mediate K+ uptake by guard cells from Vicia faba L. while simultaneously activating an apparently time-independent, inward current to depolarize the membrane potential and promote K+ efflux through a second class of K+ channels. The data are consistent with a transient rise in cytoplasmic free Ca2+ and demonstrate that intact guard cells are competent to use InsP3 in signal cascades controlling ion flux through K+ channels.     

G蛋白在杨树气孔运动中的作用

群众杨叶片下表皮经过不同浓度的G蛋白激活剂霍乱毒素(CTX)处理后,在扫描电镜下观察了气孔开度的变化,并用透射电镜结合X-射线能谱显微分析技术,对保卫细胞内的K^ 、Cl^-含量进行了研究。结果表明：CTX能促进气孔关闭,作用强度随CTX浓度的增加而增强。伴随着气孔关闭,保卫细胞液泡和细胞质中的K^ 、Cl^-含量都明显下降,而细胞壁中的K^ 、Cl^-含量增加。这提示G蛋白可能通过对保卫细胞内K^ 、Cl^-的调节作用而参与了气孔运动过程。     

人参果试管苗与移栽成活苗气孔行为的比较

通过对人参果（Solanum muricatum Ait.)试管苗与移栽成活苗气孔大小、气孔开度日变化及保卫细胞对碘-硫酸反应的观察比较,研究了试管苗与移栽成活苗气孔行为的差异,结果表明,试管苗与移栽成活苗的气孔大小日变化幅度不大,但两者气孔开度的日变化明显不同;试管苗日变化曲线平稳,无明显波动;移栽成活苗日变化曲线呈明显波峰状,在13：00达到最大值,碘-硫酸反应表明,滴加硫酸后,试管苗与移栽成活苗气孔开度都显著增大,与硫酸作用前相比,移栽成活苗的增加幅度约6倍于试管苗,这可能与两者保卫细胞壁刚性不同有关。     

SLAC1 is required for plant guard cell S-type anion channel function in stomatal signalling

Stomatal pores, formed by two surrounding guard cells in the epidermis of plant leaves, allow influx of atmospheric carbon dioxide in exchange for transpirational water loss. Stomata also restrict the entry of ozone--an important air pollutant that has an increasingly negative impact on crop yields, and thus global carbon fixation and climate change. The aperture of stomatal pores is regulated by the transport of osmotically active ions and metabolites across guard cell membranes. Despite the vital role of guard cells in controlling plant water loss, ozone sensitivity and CO2 supply, the genes encoding some of the main regulators of stomatal movements remain unknown. It has been proposed that guard cell anion channels function as important regulators of stomatal closure and are essential in mediating stomatal responses to physiological and stress stimuli. However, the genes encoding membrane proteins that mediate guard cell anion efflux have not yet been identified. Here we report the mapping and characterization of an ozone-sensitive Arabidopsis thaliana mutant, slac1. We show that SLAC1 (SLOW ANION CHANNEL-ASSOCIATED 1) is preferentially expressed in guard cells and encodes a distant homologue of fungal and bacterial dicarboxylate/malic acid transport proteins. The plasma membrane protein SLAC1 is essential for stomatal closure in response to CO2, abscisic acid, ozone, light/dark transitions, humidity change, calcium ions, hydrogen peroxide and nitric oxide. Mutations in SLAC1 impair slow (S-type) anion channel currents that are activated by cytosolic Ca2+ and abscisic acid, but do not affect rapid (R-type) anion channel currents or Ca2+ channel function. A low homology of SLAC1 to bacterial and fungal organic acid transport proteins, and the permeability of S-type anion channels to malate suggest a vital role for SLAC1 in the function of S-type anion channels.     

沙打旺单细胞产生的愈伤组织分化及植株形成

将由单细胞培养产生的愈伤组织,作为一个整体,分析它内部分化的栓化组织、拟分生组织、维管组织结节、薄壁组织.栓化组织是不连续的,由愈伤组织周缘的愈伤形成层产生,位于愈伤组织的表层,具周皮的作用.它的形成是愈伤组织产生分化的标志.拟分生组织,由愈伤组织中的薄壁细胞脱分化形成,位于愈伤组织的各个位置,相当于愈伤组织中的分生组织.其作用有三点:(1)形成器官原基;(2)拟分生组织分化的拟形成层形成维管组织结节;(3)产生薄壁细胞补充到愈伤组织中.维管组织结节是由拟形成层产生.位于愈伤组织的深层,由木质部、形成层、韧皮部三部份构成.可能具有输导作用.维管组织的分化,有一个“向心性”,可能与淀粉的积累有关.在维管组织结节部位,淀粉粒在木质部积累较少;在韧皮部侧积累较多;形成层中极少有淀粉粒.再生植株的芽是外起源的,与维管组织结节无联系.根为内起源,与维管组织结节有联系.     

肾蕨叶表皮结构和气孔器发育的研究

在光学和电子显微镜下对肾蕨(Nephrolepis auriculata (L.)Trimen)叶表皮结构和气孔器发育进行了观察.肾蕨成熟叶表皮细胞呈不规则形,含有叶绿体.气孔器仅存在于下表皮,每个气孔器有2个保卫细胞,但副卫细胞3～6个不等.仅有2个副卫细胞和保卫细胞同源.保卫细胞和副卫细胞都含有叶绿体.气孔器的发育过程可分为3个主要时期:1)保卫细胞母细胞形成期;2)气孔器幼期;3)气孔器成熟期.在扫描电镜下观察,覆盖在表皮基本细胞和气孔器细胞外壁的角质膜呈波纹状.     

ATHK1 acts downstream of hydrogen peroxide to mediate ABA signaling through regulation of calcium channel activity in Arabidopsis guard cells

Plants gradually develop their ability to tolerate environmental water deficit as part of the evolutionary process.Abscisic acid(ABA) plays a critical role during drought and osmotic stress.Several histidine protein kinases are regarded as osmotic sensors or regulators in the adaptive response of plants to water deficit.In this study,we report that ATHK1,which was previously shown to function as an osmotic regulator,is involved in ABA-induced stomatal signaling in Arabidopsis.Mutants null for ATHK1 expression were unable to transmit normal ABA responses in guard cells,including inducing stomatal closure,producing hydrogen peroxide and activating calcium influx.Moreover,patch clamp and confocal analysis demonstrated that ATHK1 may function downstream of hydrogen peroxide in ABA-induced stomatal closure,by regulating calcium channel activity and calcium oscillation in Arabidopsis guard cells.     

Phot1 and phot2 mediate blue light regulation of stomatal opening.

The stomatal pores of higher plants allow for gaseous exchange into and out of leaves. Situated in the epidermis, they are surrounded by a pair of guard cells which control their opening in response to many environmental stimuli, including blue light. Opening of the pores is mediated by K(+) accumulation in guard cells through a K(+) channel and driven by an inside-negative electrical potential. Blue light causes phosphorylation and activation of the plasma membrane H(+)-ATPase that creates this potential. Thus far, no blue light receptor mediating stomatal opening has been identified, although the carotenoid, zeaxanthin, has been proposed. Arabidopsis mutants deficient in specific blue-light-mediated responses have identified four blue light receptors, cryptochrome 1 (cry1), cryptochrome 2 (cry2), phot1 and phot2. Here we show that in a double mutant of phot1 and phot2 stomata do not respond to blue light although single mutants are phenotypically normal. These results demonstrate that phot1 and phot2 act redundantly as blue light receptors mediating stomatal opening.     

蓝光调节气孔运动的信号途径及其与脱落酸相互关系研究

气孔运动与植物水分代谢密切相关.保卫细胞可有效感知和整合多种环境信号,通过控制离子进出调节其膨压,影响气孔开与闭.诸多研究表明,蓝光信号诱导气孔开放和逆境信号脱落酸(ABA)促进气孔关闭构成了气孔运动的两大研究领域.该文就保卫细胞中蓝光信号传递及与ABA信号交叉控制气孔开闭的研究进展进行综述,以了解气孔对蓝光和ABA反应的最新进展,为发展耐旱与提高作物水分利用效率生物技术的改进提供理论支持.     

Effects of the disaggregation of high—polymerized particles in guard cell vacuoles on osmoregulation of stomatal aperture（stomata opening）

Observation under an electron microscope reveals that in closed and open stomata of V.faba,the average volume of particles in guard cell vacuoles(GCV) reduces about 3 orders in magnitude,while the distribution density of the particles increases about 2 orders of magnitude.By using the method of the ration of fluorescent emissions with laser scanning confocal microscopy,the monitoring to stomata opening shows that during 10 to 30 s before the first distinguishable aperture of stomata,there is a change of pH in GCV about-0.5 units.A quick stomatal opening immediately follows the changes of pH in GCV to reach a steady aperture about 12μm in 100-200s.This work proposes a model for the osmoregulation in GCV for stomatal opening.The proposed osmoregulation is related to the disaggregation of some polymerized particles inside GCV,which is probably induced by a-ΔpH in the vacuole.This model describes a process of osmoregulation that avoids the massive energy consuming transportation across cell membranes,which is a foundation of the current chemiosmotic hypothesis.This model is a supplement to the multiple controlling hypothesis for the stomatal movement,which widens research principle ideas for other quick movements in plants.