复合胁迫下茭白体内镉、铅的亚细胞分布和植物络合素的合成

以茭白（Zizania latifoliaTurcz.）的单季茭品种‘蒋墅茭’和双季茭品种‘葑红早’为试材,进行Cd2＋、Pb2＋的单一及复合胁迫处理,测定了茭白根系和叶片中的非蛋白巯基（NPT）、谷胱甘肽（GSH）、植物络合素（PCs）的含量,同时测定了茭白植株各亚细胞组分中Cd2＋、Pb2＋的积累量,以探讨茭白对重金属镉、铅胁迫的耐性机理。结果表明：Cd2＋、Pb2＋的单一及其复合胁迫均能促进两茭白品种根系和叶片中NPT、GSH、PCs的含量及茭白各亚细胞组分中Cd2＋、Pb2＋积累量的显著增加;复合胁迫时两茭白品种的NPT、GSH、PCs含量及各亚细胞组分中Cd2＋、Pb2＋的积累量均高于单一胁迫,茭白的不同部位间,以根系中的NPT、GSH、PCs含量显著高于叶片;茭白各亚细胞组分中Cd2＋、Pb2＋的积累量表现为：细胞壁高于原生质体,而可溶性部分高于细胞器。     

根系抗坏血酸在小麦幼苗铝耐性中的作用

以3个铝耐性不同的小麦品种为材料,研究了Al胁迫下小麦幼苗根系质外体和共质体抗坏血酸含量以及抗坏血酸氧化酶和过氧化物酶活性的变化。结果显示,Al耐性品种‘Atlas 66’质外体中抗坏血酸总含量随着处理 Al浓度的增加显著升高,而在Al敏感品种‘Scout 66’和‘扬麦9号’中显著降低。同时,‘Atlas 66’质外体中还原型抗坏血酸含量在高浓度Al处理下显著升高,2个敏感品种则在低浓度Al处理下还原型抗坏血酸含量略有升高。耐性品种‘Atlas 66’根系共质体还原型抗坏血酸和抗坏血酸总量在5～40μmol·L-1AlCl3处理下无显著变化,而在 2个敏感品种中则随处理Al浓度的增加显著下降。80μmol·L-1AlCl3处理下‘Atlas 66’根系质外体和共质体抗坏血酸氧化酶以及抗坏血酸过氧化物酶活性与对照相比均无显著变化,而在‘Scout 66’和‘扬麦9号’中则均显著降低。因此Al胁迫下‘Atlas 66’根系质外体抗坏血酸含量的升高和共质体抗坏血酸含量的维持以及Al毒害下抗坏血酸利用率较高可能是其Al耐性的一个重要机制。     

Cd-K双重处理对水稻幼苗生长和Cd吸收转运的影响及相关转运通道分析

以水稻( Oryza sativa Linn.)高 Cd 积累品种‘T 优705’(‘T You 705’)和低 Cd 积累品种‘湘早籼24’(‘Xiangzaoxian 24’)为实验材料,采用水培法对不同浓度Cd(0.0和2.7μmol·L-1 Cd)和K(0、30和60 mmol·L-1 K)处理条件下2个品种幼苗的相对生长量、根系和地上部的Cd含量及其亚细胞分布特征进行了比较,并分析了添加离子通道活性抑制剂TEA和LaCl3后幼苗根系和地上部的Cd和K含量;在此基础上,比较了NSCCs(非选择性阳离子通道)和K专性通道对2个品种幼苗根系和地上部Cd和K吸收贡献率的影响。结果表明：与Cd单一处理组(2.7μmol·L-1 Cd)相比, Cd-K双重处理组(2.7μmol·L-1 Cd-30 mmol·L-1 K和2.7μmol·L-1 Cd-60 mmol·L-1 K)2个品种幼苗的相对生长量显著提高,而幼苗根系和地上部的Cd含量显著下降;随K浓度的提高,2个品种幼苗根系细胞壁和细胞液中的Cd含量显著下降,但细胞壁中Cd含量的分配比例增大而细胞液中Cd含量的分配比例则减小。在含2.7μmol·L-1 Cd和30 mmol·L-1 K的培养液中分别添加5 mmol·L-1 TEA或0.2 mmol·L-1 LaCl3后,2个品种幼苗根系和地上部的Cd和K含量均显著下降,其中,LaCl3处理组的根系Cd含量降幅高于TEA处理组,但LaCl3处理组的根系K含量降幅则低于TEA组。 NSCCs对品种‘T优705’幼苗根系和地上部Cd吸收的贡献率显著低于品种‘湘早籼24’幼苗,而K专性通道对品种‘T优705’幼苗根系K吸收和地上部Cd吸收的贡献率则显著低于品种‘湘早籼24’幼苗。研究结果显示：添加外源K可缓解Cd对水稻幼苗生长的抑制作用,并通过提高细胞壁与Cd的结合能力来降低细胞液中Cd的积累,以此减弱幼苗对Cd的吸收和转运能力;幼苗体内的K和Cd均可通过K专性通道和NSCCs转运,其中,K吸收和转运主要通过K专性通道完成,而Cd吸收和转运主要通过NSCCs完成。此外,品种‘T优705’可能具有多种离子通道参与Cd的吸收和转运,而品种‘湘早籼24’主要依赖NSCCs参与Cd的吸收和转运,且后者对K的吸收和积累强于前者。     

铬污染人工湿地薏米对铬的积累和分布

通过桶栽构筑微型垂直流人工湿地,在生活污水中添加不同浓度的Cr~(6+)(0、20、40 mg·L~(-1),用K_2Cr_2O_7配置),研究了在铬(Cr)污染条件下薏米不同器官和亚细胞水平对铬的积累及分布。结果表明:薏米根茎叶铬含量均随铬处理浓度的提高和处理时间的延长而显著增加,根部铬含量最高,其次是叶片,茎部铬含量最低。随着铬处理浓度的增加和时间的延长,细胞壁和液泡中的铬含量大幅上升,但细胞器中铬含量上升幅度较低,根部细胞壁、细胞器、可溶部分及液泡铬含量均显著大于茎、叶相关细胞组分,根、茎、叶亚细胞水平铬含量分布均依次为细胞壁液泡细胞器。其中,细胞壁铬含量极显著大于液泡以及其他细胞器,薏米根部把大部分铬隔离在细胞壁中,从而降低了地上部分对铬的吸收,导致地上部分干重、鲜重的受抑制程度均低于根系,表明根部细胞壁是降低铬在薏米体内积累的重要屏障,也是薏米在低浓度铬胁迫下能够存活的根本保证。     

外源亚精胺对盐碱胁迫下番茄幼苗根系线粒体功能的影响

为探讨外源亚精胺(Spd)对盐碱胁迫下番茄根系线粒体功能的影响,采用水培法,以耐性不同的两个番茄品种‘金棚朝冠’(耐盐型)和‘中杂9号’(敏感型)为试材,通过模拟盐碱生态条件(Na Cl∶Na_2SO_4∶Na HCO3∶Na2CO3=1∶9∶9∶1),结合叶面喷施外源0.25mmol·L~(-1)Spd,研究盐碱胁迫8 d后Spd对番茄幼苗根系形态和根系线粒体功能的影响.结果表明:盐碱胁迫下,两个品种番茄根系线粒体内H_2O_2和丙二醛(MDA)含量增加,线粒体膜通透性明显增大,流动性降低,膜电位、线粒体内细胞色素c/a(Cyt c/a)吸光度比值、膜H~+-ATPase活性显著下降,使线粒体受到不同程度的损伤,从而抑制根系生长,且‘金棚朝冠’的上述指标变化幅度均小于‘中杂9号’.盐碱胁迫下,喷施外源Spd处理的两个品种根系线粒体H_2O_2和MDA含量显著降低,膜通透性减小、流动性增加,膜电位、线粒体内Cyt c/a吸光度比值、膜H~+-ATPase活性显著提高,可有效缓解盐碱胁迫对番茄幼苗根系线粒体的伤害作用,且这种缓解作用在‘中杂9号’上的表现效果更佳.     

外源水杨酸对NaCl胁迫下棉花幼苗生长生理特性的影响

以‘中棉所41号’(耐盐品种)和‘中棉所49号’(中等耐盐品种)为试验材料,采用水杨酸(SA)浸种+叶面喷施复合处理,通过温室盆栽试验探究了0.60%NaCl胁迫下外源SA对棉花幼苗生长、渗透调节能力和抗氧化能力的影响。结果表明:(1)NaCl胁迫显著抑制了棉花幼苗的生长,外源SA处理下棉花幼苗的株高、叶面积、干物质质量、根系活力、根冠比均有升高,叶片中丙二醛(MDA)、可溶性糖(SS)、可溶性蛋白(SP)和脯氨酸(Pro)含量增加,叶片和根系中超氧化物歧化酶(SOD)、过氧化物酶(POD)和过氧化氢酶(CAT)活性显著提高,且棉花幼苗根系的SOD、POD和CAT活性小于叶片,SA处理对棉花幼苗根系酶活力的提高幅度大于叶片。(2)SA浸种+叶面喷施复合处理对0.60%NaCl胁迫的缓解作用优于单独SA浸种处理,尤以0.05mmol·L~(-1)浸种+0.2mmol·L~(-1)叶面喷施处理最好。(3)两品种比较,SA对‘中棉所49号’的盐胁迫缓解作用大于‘中棉所41号’,且以棉花幼苗根系生长发育的表现最为显著;外源SA使‘中棉所41号’的根系活力提高10.58%~57.56%,‘中棉所49号’提高15.08%~80.48%。研究发现,外源SA能通过调控棉花幼苗的渗透调节和抗氧化能力来减轻细胞膜损伤,提高其耐盐性,但缓解效应在不同耐性品种和处理方式间存在明显差异。     

干旱胁迫对不同苦荞品种苗期生长和根系生理特征的影响

采用盆栽人工控水试验,研究了不同水分处理(正常供水、中度干旱和重度干旱)对耐旱型(‘迪庆苦荞’、‘西农9909’)和不耐旱型(‘西荞1号’和‘黑丰1号’)苦荞品种苗期生理、形态指标的影响,并通过隶属函数法与主成分分析对品种抗旱性进行综合评价,以揭示苦荞苗期的抗旱生理机制。结果表明:(1)与正常供水相比,除‘迪庆苦荞’和‘西农9909’在重度干旱胁迫下主根长呈升高趋势外,其余苦荞品种在2个干旱条件下的株高、茎粗、叶面积、地上部干重、地下部干重、根系体积、根系表面积均呈下降趋势,且耐旱品种降幅小于不耐旱品种;重度干旱胁迫使得‘迪庆苦荞’的根冠比升高,而其余品种根冠比在干旱胁迫下均无显著变化。(2)干旱胁迫使苦荞叶片的叶绿素含量、相对含水量、最大荧光产量(Fm)、最大光化学效率(Fv/Fm)、根系活力和可溶性蛋白含量显著降低,而根系超氧化物歧化酶(SOD)活性、过氧化物酶(POD)活性、丙二醛(MDA)、可溶性糖及游离脯氨酸含量呈升高趋势;不同抗旱性品种间的升降幅度存在差异。(3)各苦荞品种耐旱能力综合评价值(D)表现为‘迪庆苦荞’‘西农9909’‘黑丰1号’‘西荞1号’;幼苗株高、地下部干重及根系SOD活性和蛋白质含量与D值呈显著正相关关系,而根系脯氨酸含量和可溶性糖含量与D值呈极显著正相关关系。研究发现,在中度与重度干旱逆境下,苦荞品种‘迪庆苦荞’和‘西农9909’综合表现较好,具有更强的耐旱能力,而品种‘西荞1号’和‘黑丰1号’综合表现较差,其抗旱性较弱;苗期株高、地下部干重以及根系SOD活性、蛋白质含量、脯氨酸含量和可溶性糖含量可作为苦荞抗旱性快速鉴定的指标。     

水稻和小麦根尖细胞壁多糖的铝积累能力比较

采用水培法比较4种禾本科植物水稻(Oryza sativa L.)、玉米(Zea mays L.)、高粱(Sorghum bicolor(L.) Moench)和小麦(Triticum aestivum L.) 8个基因型的抗铝(Al)能力,并对他们在Al积累后细胞壁的多糖组分进行分析。结果显示,在5～200μmol/L Al处理下,水稻抗Al能力较强,而小麦抗Al能力较弱。在50μmol/L Al处理下,小麦根尖的果胶和半纤维素1含量的增幅明显高于水稻。水稻基因型‘日本晴’与‘浙辐802’的细胞壁Al含量分别占根尖总Al含量的78. 7%和91. 6%;小麦基因型‘扬麦18’与‘扬麦16’Al含量分别占根尖总Al含量的64.9%和72.1%。Al吸附-解吸实验结果显示,小麦根尖细胞壁上Al的吸附量高于水稻。研究结果表明,细胞壁是Al积累的主要部位,对Al敏感的水稻和小麦基因型细胞壁中的Al主要分布在果胶中;而对Al耐性较强的水稻和小麦基因型细胞壁中的Al主要分布在半纤维素1中。     

外源Ca~(2+)连续喷施时间对辣椒淹水胁迫的缓解效应研究

以‘博辣红牛’辣椒为材料,研究外源Ca~(2+)连续喷施不同天数对淹水胁迫下辣椒幼苗农艺性状和生理指标的影响,探讨Ca~(2+)对辣椒淹水胁迫伤害的缓解作用和适宜的喷施处理天数。结果显示:(1)辣椒幼苗生物量、壮苗指数、叶绿素、根系活力、脯氨酸、可溶性糖以及CAT和SOD活性随施Ca~(2+)天数的增加呈先升高后下降的趋势,MDA含量随施Ca~(2+)天数的增加呈先下降后上升的趋势。(2)施Ca~(2+)1d(T1d)处理对辣椒淹水胁迫伤害无明显缓解作用,连续施Ca~(2+)3d(T3d)和6d(T6d)处理的缓解效果不断增强,至连续施Ca~(2+)9d(T9d)时缓解效果达到最佳,随后连续施Ca~(2+)12d(T12d)和20d(T20d)处理的缓解效果又逐渐减弱,但仍显著优于T1d处理。研究表明,外源Ca~(2+)可以诱导增加淹水胁迫下辣椒幼苗渗透调节物质含量,上调抗氧化酶活性,降低叶绿素的降解,大幅提高根系活力,从而缓解淹水胁迫所造成的各种伤害,增强其忍耐淹水胁迫能力,并以连续施钙9d对淹水胁迫的缓解效果最佳。     

铜胁迫对狭叶香蒲生长及生理特性的影响

采用水培法,研究不同浓度Cu~(2+)胁迫对狭叶香蒲生长及生理特性的影响。结果表明:狭叶香蒲叶宽、株高的增长量及整株干物质累积量在较低浓度Cu~(2+)处理时均未受影响,此后随Cu~(2+)浓度升高显著下降。Cu~(2+)浓度为o～30 mg·L~(-1)时,叶绿素含量显著上升,此后随Cu~(2+)浓度增加其含量显著下降。在Cu~(2+)浓度0～5 mg·L~(-1)范围内根系活力显著上升,此后随Cu~(2+)浓度升高则大幅下降。根系和叶片SOD、POD、CAT、活性随Cu~(2+)浓度增加均呈先显著升高后显著降低的趋势,根系SOD、POD活性在30 mg·L~(-1)时出现最大值,叶片SOD、POD活性在55 mg·L~(-1)时出现最大值,根系和叶片CAT活性均在80 mg·L~(-1)时出现最大值;同一Cu~(2+)浓度下,根系SOD、POD、CAT活性明显高于叶片,说明根系比叶片对Cu~(2+)胁迫反应更敏感。从根系和叶片SOD/POD、SOD/CAT的比值变化上看出,在Cu~(2+)为1 mg·L~(-1)和5 mg·L~(-1)时起主要保护作用的酶是SOD、POD,后来CAT起到主要作用。MDA含量自Cu~(2+)浓度为30 mg·L~(-1)时开始持续上升。说明在Cu~(2+)为30～55 mg·L~(-1)时狭叶香蒲表现为积极的生理响应。     

(Na + K)-ATPase activity of crude homogenates of rat skeletal muscle as estimated from their K-dependent 3-O-methylfluorescein phosphatase activity

A highly sensitive fluorimetric assay using 3-O-methylfluorescein phosphate as substrate was used in the determination of K⁺-dependent phosphatase activity in preparations of rat skeletal muscle. The gastrocnemius muscle was chosen because of mixed fibre composition. Crude, detergent treated homogenate was used so as to avoid loss of activity during purification. K⁺-dependent phosphatase activities in the range 0.19–0.37 μmol · (g wet weight)⁻¹ · min⁻¹ were obtained, the value decreasing with age and K⁺-deficiency. Complete inhibition of the K⁺-dependent phosphatase was obtained with 10⁻³ M ouabain. Using a KSCN-extracted muscle enzyme the intimate relation between K⁺-dependent phosphatase activity and (Na⁺ + K⁺)-activated ATP hydrolysis could be demonstrated. A molecular activity of 620 min⁻¹ was estimated from simultaneous determination of K⁺-dependent phosphatase activity and [³H]ouabain binding capacity using the partially purified enzyme preparation. The corresponding enzyme concentration in the crude homogenates was calculated and corresponded well with the number of [³H]ouabain binding sites measured in intact muscles or biopsies hereof.     

Fluorescent labeling of (Na + K)-ATPase by 5-iodoacetamidofluorescein

5-Iodoacetamidofluorescein (5-IAF) covalently labels dog kidney (Na⁺ + K⁺)-ATPase with approximately 2 moles incorporated per mole of enzyme. ATPase and K⁺-phosphatase activities are fully retained after reaction, and the kinetic parameters for Na⁺, K⁺, Mg²⁺, ATP and p-nitrophenyl phosphate are likewise not significantly affected. The fluorescence of the bound 5-IAF is increased by ATP, Na⁺, and Mg²⁺, and decreased by K⁺. These fluorescence changes likely reflect ligand-induced stabilization of the E₁ or E₂ states of the enzyme.     

Annular and non-annular binding sites on the (Ca + Mg)-ATPase

Quenching of the fluorescence of the (Ca²⁺ + Mg²⁺)-ATPase purified from muscle sarcoplasmic reticulum can be used to measure relative binding constants of hydrophobic compounds to the phospholipid-protein interface. We show that the binding constant for cholesterol is considerably less than that for phosphatidylcholine, so that cholesterol is effectively excluded from the phospholipid annulus around the ATPase. However, dibromocholestan-3β-ol causes quenching of the fluorescence of the ATPase, and so has access to other, non-annular sites. We suggest that these non-annular sites could be at protein/protein interfaces in ATPase oligomers. Oleic acid can bind at the phospholipid/protein interface, although its binding constant is less than that for a phosphatidylcholine, and it can also bind at the postulated non-annular sites. The effects of these compounds on the activity of the ATPase depend on the structure of the phospholipid present in the systems.     

The phospholipid requirement of the (Ca + Mg)-ATPase from human platelets

The phospholipid requirement of the (Ca²⁺ + Mg²⁺)-ATPase present in a membrane fraction from human platelets was studied using various purified phospholipases. Only those phospholipases, which hydrolyse the negatively charged phospholipids, inhibited the (Ca²⁺ + Mg²⁺)-ATPase activity. The ATPase activity could be restored by adding mixed micelles of Triton X-100 and phosphatidylserine or phosphatidylinositol. Micelles with phosphatidic acid, phosphatidylcholine, phosphatidylethanolamine or sphingomyelin could not be used for reconstitution and inhibited the activity of the native enzyme.     

Synthesis of N-monomethylagmatine, a decarboxylation product of N-monomethyl- -arginine

N^G-Monomethylagmatine, a decarboxylation product of N^G-monomethyl- -arginine, has been synthesized by reacting putrescine with N,S-dimethylthiopseudouronium iodide. The structural identity of the product was confirmed by proton NMR and mass spectroscopy, and its properties were determined on thin-layer and electrophoretic chromatography.     

The efficiency of (Na + K)-ATPase in tumorigenic cells

The efficiency of (Na⁺ + K⁺)-ATPase (i.e. the amount of K⁺ pumped per ATP hydrolyzed) in intact tumorigenic cells was estimated in this study. This was accomplished by simultaneously measuring the rate of ouabain-sensitive K⁺ uptake and oxygen consumption in tumorigenic cell suspensions during the reintroduction of K⁺ to K⁺-depleted cells. The ATP turnover was then estimated by assuming 5.6–6 ATP/O₂ as the stoichiometry of NADH-linked respiration in these cells. In the three cell lines tested (hamster and chick embryo cells transformed with Rous sarcoma virus and Ehrlich ascites cells), the K⁺/ATP ratio was approximately 2, the same value as that found in normal tissues. Furthermore, only 20% of the total ATP production of these cells was used by (Na⁺ + K⁺)-ATPase.     

Mathematical modelling of ATP, K and Na interactions with (Na + K)-ATPase occurring under equilibrium conditions

The controlling effect of ATP, K⁺ and Na⁺ on the rate of (Na⁺ + K⁺)-ATPase inactivation by 7-chloro-4-nitrobenzo-2-oxa-1,3-diazole (NBD-Cl) is used for the mathematical modelling of the interaction of the effectors with the enzyme under equilibrium conditions.

1. 1. Of a series of conceivable interaction models, designed without conceptual restrictions to describe the effector control of inactivation kinetics, only one fits the experimental data described in a preceding paper.

2. 2. The model is characterized by the coexistence of two binding sites for ATP and the coexistence of two separate binding sites for K⁺ and Na⁺ on the enzyme-ATP complex. On the basis of this model, the effector parameters fitting the experimental data most closely are estimated by means of nonlinear least-squares fits.

3. 3. The apparent dissociation constants for ATP of the enzyme-ATP complex or of the enzyme-(ATP)₂ complex are computed to lie near 0.0024 mM and 0.34 mM, respectively, irrespective of whether K⁺ and Na⁺ were absent or K⁺ and K⁺ plus Na⁺, respectively, were present in the experiments.

4. 4. The origin of the high and the low affinity site for binding of ATP to the (Na⁺ + K⁺)-ATPase molecule is traced back to the coexistence of two catalytic centres which, although primarily equivalent as to the reactivity of their thiol groups with NBD-Cl, are induced into anticooperative communication by ATP binding and thus show an induced geometric asymmetry.

Molecular analysis of two genes between let-653 and let-56 in the unc 22(IV) region of Caenorhabditis elegans

Summary A previous study of genomic organization described the identification of nine potential coding regions in 150 kb of genomic DNA from the unc-22(IV) region of Caenorhabditis elegans. In this study, we focus on the genomic organization of a small interval of 0.1 map unit bordered on the right by unc-22 and on the left by the left-hand breakpoints of the deficiencies sDf9, sDf19 and sDf65. This small interval at present contains a single mutagenically defined locus, the essential gene let-56. The cosmid C11F2 has previously been used to rescue let-56. Therefore, at least some of C11F2 must reside in the interval. In this paper, we report the characterization of two coding elements that reside on C11F2. Analysis of nucleotide sequence data obtained from cDNAs and cosmid subclones revealed that one of the coding elements closely resembles aromatic amino acid decarboxylases from several species. The other of these coding elements was found to closely resemble a human growth factor activatable Na⁺/H⁺ antiporter. Pairs of oligonucleotide primers, predicted from both coding elements, have been used in PCR experiments to position these coding elements between the left breakpoint of sDf19 and the left breakpoint of sDf65, between the essential genes let-653 and let-56.