The Diffusion of KCI from Micro-electrodes

The diffusion of salt from the tips of 3 M KCI filled micro-electrodesis measured. The leakage of KCI in 3 min is found to be 5.70x 10^–12–1.08 x 10^–11 mol for tips of 0.2–0.6µm diameter, and 2.70 x 10^–11 mol for tips of 0.6–1.0µm diameter. The observed leakages compare with estimatesbased on the diffusion coefficient of KCI and micro-electrodegeometry. The results confirm that the use of 3 M KCI in micro-electrodesmay lead to serious contamination of impaled cells providedthe electrode tips remain unoccluded.     

植物细胞质膜H+-ATPase的调控

综述了质膜H ATPase在转录、翻译及翻译后水平上所受调控现象及其机制的研究进展     

Characterization of a H Efflux from Suspension-cultured Plant Cells

A readily assayed H⁺ efflux from sycamore (Acer pseudoplatanus), rye (Lolium perenne), and bean (Phaseolus vulgaris cultivars Red Kidney and Small White) suspension-cultured cells has been detected and partially characterized. The H⁺ efflux has been shown to require a source of energy, to be significantly stimulated by Na⁺ and Mg²⁺ but not by K⁺ and Ca²⁺, and to have a pH optimum at 7. The study of this H⁺ efflux was undertaken because the characteristics of auxin-induced growth and of H⁺-induced growth are sufficiently similar to suggest that a H⁺ efflux may be an intermediate in the mechanism of auxin-induced growth. However, the H⁺ efflux from these suspension-cultured cells was found to be insensitive to exogenously added hormones.     

液泡膜H+ -PPase与植物耐盐性

文章介绍植物液泡膜H^＋-PPase的结构、功能及其分子生物学的研究进展,并着重阐述液泡膜H^＋-PPase在植物耐盐性中的作用。     

H+-Independent Glutamine Transport in Plant Root Tips

Background

Glutamine is one of the primary amino acids in nitrogen assimilation and often the most abundant amino acid in plant roots. To monitor this important metabolite, a novel genetically encoded fluorescent FRET-reporter was constructed and expressed in Arabidopsis thaliana. As a candidate for the glutamine fluxes, the root tip localized, putative amino acid transporter CAT8 was analyzed and heterologously expressed in yeast and oocytes.

Principal Findings

Rapid and reversible in vivo fluorescence changes were observed in reporter-expressing root tips upon exposure and removal of glutamine. FRET changes were detected at acid and neutral pH and in the presence of a protonophore, suggesting that part of the glutamine fluxes were independent of the pH. The putative amino acid transporter CAT8 transported glutamine, had a half maximal activity at ∼100 µM and the transport was independent of external pH. CAT8 localized not only to the plasma membrane, but additionally to the tonoplast, when tagged with GFP. Ultrastructural analysis confirmed this dual localization and additionally identified CAT8 in membranes of autophagosomes. Loss-of function of CAT8 did not affect growth in various conditions, but over-expressor plants had increased sensitivity to a structural substrate analog, the glutamine synthetase inhibitor L-methionine sulfoximine.

Conclusions

The combined data suggest that proton-independent glutamine facilitators exist in root tips.     

The Repertoire of Plant Cells

Cryptic (camouflaged) prey often seek out backgrounds that match their coloration, and when at rest adopt an attitude that makes their crypsis most effective. We suggest a simple method for investigating the adaptive significance of such orientation. We used flat discs of pastry as the ‘prey’, either plain white or painted with a central black stripe, and wild garden birds as the predators. In the eight main experiments the backgrounds were white wooden boards painted with black parallel stripes of the same width as the stripes on the prey. In each experiment we presented equal numbers of two (of seven) ‘treatments’ of prey. The selection resulting from the combined predation by the birds confirmed the advantages of resting on a matching background and in the ‘correct’ orientation. We suggest that the technique can be developed further to explore the adaptive significance of background matching     

PPiase-Activated ATP-Dependent H+ Transport at the Tonoplast of Mesophyll Cells of the CAM Plant Kalanchoë daigremontiana*

The co-ordinated action of the two proton-transporting enzymes at the tonoplast of the CAM plants. daigremontiana, viz. the ATPase and the PP_iase, was studied by measuring fluorescent dye quenching. The initial rates of ATP and PP_i-dependent H⁺ transport into tonoplast vesicles were additive, i.e. the sum of the rates obtained with each substrate alone was in the range obtained with both substrates added together at the same time. Conversely, the activities of the two H⁺ pumps were non-additive in establishing the steady-state level, indicating that the final steady state was under thermodynamic control of a maximal attainable proton gradient. The initial rates of ATP-dependent H⁺ transport were stimulated enormously if ATP was added a few minutes after pre-energization of the vesicles with PP_i. This stimulation was observed only when the PP_iase was active. A similar effect was not found for PP_i-dependent H⁺ transport after pre-energization with ATP. Hence, a PP_iase-activated ATP-dependent H⁺ transport can be distinguished from the basic ATP- and the basic PP_i-dependent H⁺ transport. In parallel a PP_i-dependent stimulation of ATP hydrolysis in the absence of ionophores was measured, which can only be attributed to the activity of the PP_iase. PP_iase-activated ATP-dependent H⁺ transport depends on the presence of permeant anions. It shows properties of both H⁺ transport activities, i.e. the chloride and malate stimulation and the DCCD inhibition of the ATP-dependent H⁺ transport activity, the nitrate stimulation and the KF inhibition of the PP_i-dependent H⁺ transport activity. Only MgPP_i and MgATP were effective as the respective substrates. The PP_iase-activated ATP-dependent H⁺ transport had a half life of about 5–9 minutes. It is concluded that the PP_iase may play an important role in kinetic regulation of the ATPase, and implications for CAM metabolism are discussed.     

Respiration-Dependent H+ Efflux from Intact Cells of Cyanidium caldarium

An active H⁺ efflux depending on respiration was found in anacidophilic unicellular alga, Cyanidium caldarium. Alkalizationof the medium due to passive H⁺ transport into the cells wasobserved when the respiratory activity was inhibited by addingrespiratory poisons, such as rotenone or antimycin A, or byintroducing pure nitrogen into the cell suspension. The extentof the H⁺ influx increased as the pH of the medium was loweredto 2.9, indicating that H⁺ leaks into the cells according tothe pH gradient across the plasma membrane. The medium pH whichhad increased under anaerobic condition returned to the originallevel with aeration of the cell suspension. This suggests thatan active H⁺ transport, related to respiration, pumps out theexcess H⁺ accumulated in the cells during anaerobic preincubation.The pH changes in the cell suspension were related to the intracellularATP level. From these results it was concluded that active H⁺efflux dependent upon oxidative phosphorylation functions inthe dark to maintain a constant intracellular pH against passiveH⁺ leakage through the plasma membrane. The light-induced H⁺ efflux and the respiration-dependent H⁺efflux were also compared in relation to the physiological roleof the active H⁺ efflux, especially with respect to the intracellularpH regulation in this alga. ¹The data in this paper are included in the Ph. D. dissertationsubmitted by M. Kura-Hotta to Tokyo Metropolitan University. (Received February 3, 1984; Accepted June 14, 1984)     

Light-induced H+ Efflux from Intact Cells of Cyanidium caldarium

Light-induced pH changes in suspensions of an acidophilic unicellularalga, Cyanidium caldarium Geitler, were studied as a functionof the pH of the medium. In the neutral pH region, alkalizationof the medium due to photosynthetic CO₂ uptake was observed.In the acidic pH region, illumination caused a significant decreasein the pH of the medium, indicating the efflux of H⁺ from thecells. Both the rate and extent of the pH decrease increasedas the pH of the medium was lowered to 3.0. The H⁺ efflux wasnot affected by 3-(3',4'-dichlorophenyl)-l,l-dimethylurea, butwas inhibited by phenylmercuric acetate. The fastest H⁺ effluxoccurred at 45°C, whereas its extent was almost constantfrom 25 to 50°C. The activity decreased at temperaturesabove 50°C and was inactivated completely at 60°C. Itsaction spectrum corresponded the spectrum for chlorophyll aabsorption. Results indicate that the light-induced H⁺ effluxis driven by photosystem I and is important in the maintenanceof the intracellular pH at the functional neutral region againsta steep pH gradient across the cell membrane. (Received May 6, 1981; Accepted August 8, 1981)     

The Cost of Maintenance Processes in Plant Cells

The most important maintenance processes in plants are proteinturnover and active transport processes to maintain certainion concentrations in the cells. In this paper an attempt ismade to calculate the total energy cost of these processes fromwhat is known about their specific costs and what has been observedabout their rates. Because of insufficient reliable data aboutrates of individual maintenance processes, only approximatevalues can be obtained. The average turnover rate of leaf proteins may be about 100mg protein per g proteins per day at normal temperature in leavesassimilating at moderate light intensities. This process consumes28–53 mg glucose per g protein per day, which equals 7–13mg glucose per g dry weight per day in leaves. It is likelythat the rates of protein turnover and of CO₂-assimilation arerelated. The cost of maintaining ion concentrations is estimatedto be about 6–10 mg glucose per g dry weight per day inleaves. The sum of these figures is lower than is indicatedby measurements of maintenance respiration. One reason for theunderestimation may be that the protein turnover rates usedin the calculations apply to plants with lower photosyntheticrates than the plants in which the maintenance respiration wasmeasured. Effects of water stress and salinity, temperatureand other environmental factors on the rate of maintenance processesare discussed. The consumption of assimilates for maintenance of plant cellsis a significant, negative factor in plant productivity. A betterunderstanding of the maintenance processes may give a clue howto manipulate plant characteristics or the environment to reducethe amount of assimilates consumed in these processes. It issuggested that reduction in protein turnover rates may be onesuch manipulation.     

植物细胞内钙信号的特异性

文章对植物细胞内钙信号特异性的形式、特异性钙信号的产生以及解读等方面的研究结果进行了评述。     

The Plasma Membrane Ca2+-Pump of Plant Cells: A Radiation Inactivation Study

The functional molecular weight of the plasma membrane Ca²⁺-ATPase of radish (Raphanus sativus L.) seeds was determined by measuring the Ca²⁺-dependent ATPase activity and the MgATP-dependent Ca²⁺ transport activity of membrane samples irradiated, in the lyophilized state, with γ rays from [⁶⁰Co] source. The results gave a target size of about 270,000 dalton for both the measured activities, thus confirming (i) that both activities are catalyzed by the same enzyme and (ii) the similarity between the plasma membrane Ca²⁺-ATPase of higher plants and that of the erythrocytes.     

标尺模板在Branemark种植修复中的制作和应用

目的:制作并观察标尺模板在指导Branemark种植手术,提高手术安全性方面的作用。方法:选择15例患者52颗种植体,铸造制作标尺模板后戴入口腔拍摄曲面体层片。根据影象所显示的适宜种植区域与颏孔、上颌窦等颌骨重要解剖结构的位置,结合上部修复设计选择Branemark种植体数量与植入点,并借助标尺模板的定位网格予以记录。手术时将标尺模板戴入口腔,根据记录结果在颌骨表面显示种植体入点与颌骨重要解剖结构的位置,指导手术进行。术后戴入标尺模板拍摄对照片,判断种植体与预定位置的吻合性。结果:52颗种植体中,除1颗种植体因术中临时改变种植位置,其余51颗种植入位置均与术前计划一致。结论:在Branemark种植修复中制作并应用标尺模板,能方便直观地帮助术者将种植体植入预定位置,避免损伤颏孔等颌骨重要解剖结构,提高了手术的正确性与安全性。     

The Use of Papain in Clearing Plant Tissues for Whole Mounts

Materials killed and fixed in FAA (formalin-acetic acid-alcohol) and similar fixatives frequently are difficult to clear for whole mounts because the denatured proteins will not become soluble in NaOH and other clearing agents. If tissues are washed for 3 days in running water, then incubated at 40 C for 5-7 days in 2% papain buffered to pH 7.2 and activated with 15 ml of .02 M Na₂S, cell contents are partly digested. Normal clearing with 5-10% NaOH followed by chloral hydrate (sat. aq.) can then effect complete solublility of cell contents and their removal. Permanent slides can be made after staining (1% safranin O in 50% alcohol for 12 hr is successful), by dehydration through alcohols, clearing in xylene, and mounting in resin.     

The Influence of Ca2+ and K+ on H14CO3-Influx in Internodal Cells of Chara corallina

The influences of Ca²⁺-free solutions and increasing K⁺ concentrationson the H¹⁴CO^–₃ influx capacity of Chara corallina wereinvestigated. It was found that contact with Ca^2–freesolutions resulted in a gradual reduction in the H¹⁴CO^–₃influx capacity of these cells. Recovery of this influx capacity,following the return of Ca²⁺ to the experimental solution, followeda ‘mirror-image’ of the time course of decay. Potassium concentrations above a certain critical value (2 mM)induced a rapid reduction in H¹⁴CO^–₃ influx capacity.Normal activity was recovered within 60–90 min followingthe return to 0.2 mMK⁺ solutions. It was also shown that 10mM K⁺ can be used to determine the relative contribution of¹⁴C supplied by diffusion of ¹⁴CO₂ and transport of H¹⁴CO^–₃.The Ca²⁺ and K⁺ results are discussed in relation to the effectsof these treatments on the electrical properties of the plasmalemma.     

植物细胞中的前纤维蛋白

肌动蛋白组成的微丝骨架是真核细胞中的重要结构,在体内处于高度动态变化之中,受多种肌动蛋白结合蛋白（actin-binding proteins）的调节。前纤维蛋白（profilin）是一种单体肌动蛋白结合蛋白,存在于所有的真核细胞中,在植物细胞中也得到较多的研究。前纤维蛋白除可以结合单体肌动蛋白之外,还可以与磷脂酰肌醇及富含多聚脯氨酸的蛋白质等多种分子结合,在细胞信号转导中行使着重要的功能。本文结合本实验室的研究结果,概述了前纤维蛋白的最新研究进展。     

植物细胞中的膜联蛋白（annexin）

膜联蛋白（annexin）是存在于动植物细胞中的钙依赖性磷脂结合蛋白,广泛参与受钙离子调控的生命活动,如囊泡运输、膜融合、信号转导、钙离子通道的形成、细胞分化和细胞骨架蛋白间的相互作用等。本文就植物细胞中annexin的研究进展进行了介绍。