水平回转对水稻幼苗叶细胞的影响

对在模拟微重力装置上回转１４ 天的水稻幼苗叶细胞进行了亚显微形态、电子探针和细胞酶化学研究。发现叶细胞质膜上Ｃａ２＋ －ＡＴＰ酶活性消失,膜内钙总量上升、膜外钙总量下降,细胞骨架变得疏松,细胞壁变薄并凹凸不平。叶绿体的基粒和线粒体的内嵴亦有部分变化。其变化机制,首先是细胞质膜上Ｃａ２＋ －ＡＴＰ酶活性消失,膜上钙泵停止工作,跨膜钙浓度差减小,膜内钙浓度上升,微管、微丝聚合受阻,细胞骨架疏松,分泌泡移动失去导向,从而导致细胞壁变薄等状态     

Modulation of human erythrocyte Ca2+-ATPase activity by glycerol: The role of calmodulin

The effect of an intracellular cryoprotectant glycerol on human erythrocyte Ca2+-ATPase activity and possible involvement of calmodulin in the regulation of Ca2+-pump under these conditions were investigated. The experiments were carried out using saponin-permeabilized cells and isolated erythrocyte membrane fractions (white ghosts). Addition of rather low concentrations of glycerol to the medium increased Ca2+-ATPase activity in the saponin-permeabilized cells; the maximal effect was observed at 10% glycerol. Subsequent increase in glycerol concentrations above 20% was accompanied by inhibition of Ca2+-ATPase activity. Lack of stimulating effect of glycerol on white ghost Ca2+-ATPase may be attributed to removal of endogenous compounds regulating activity of this ion transport system. Inhibitory analysis using R24571 revealed that activation of Ca2+-ATPase by 10% glycerol was observed only in the case of inhibitor administration after modification of cells with glycerol; in the case of inhibitor addition before erythrocyte contact with glycerol, this phenomenon disappeared. These data suggest the possibility of regulation of human erythrocyte Ca2+-ATPase by glycerol; this regulatory effect may be attributed to both glycerol-induced structural changes in the membrane and also involvement of calmodulin in modulation of catalytic activity of the Ca2+-pump.     

脱硫废弃物对碱胁迫下水稻叶片钙分布、Ca2+-ATPase活性及抗氧化特征的影响

为了探讨脱硫废弃物提高水稻抗盐碱的作用机制,采用盆栽法,研究脱硫废弃物对碱胁迫下水稻幼苗叶片总钙含量、Ca2+分布、细胞膜Ca2+-ATPase活性及活性氧含量等的变化.结果表明： 对照处理的细胞中钙颗粒零星分布于细胞壁和叶绿体中,添加脱硫废弃物和CaSO4处理的细胞质膜、细胞间隙、细胞壁和液泡中有大量的钙颗粒分布;随着脱硫废弃物和CaSO4添加量的增加,叶片总钙含量增加,质膜和液泡膜Ca2+-ATPase活性呈上升趋势,质膜透性、MDA含量和活性氧O2〖SX(B-*3〗-〖〗·〖SX)〗产生速率呈下降趋势,SOD、POD等保护酶活性升高.添加脱硫废弃物在一定程度上能够减缓碱胁迫对水稻造成的细胞伤害,起主要作用的物质可能是其主要成分CaSO4.     

Flue gas desulfurization gypsum by-products alters cytosolic Ca2 + distribution and Ca2+-ATPase activity in leaf cells of oil sunflower in alkaline soil

Soil salinization in arid zones is a major factor that resulted in the reduction in the yield and quality of many important crops in Northwestern China. In this study, the potential mechanism of flue gas desulfurization gypsum by-product (FGDB) mediated amendment of alkaline soils was investigated in an oil sunflower model by accessing the Ca²⁺ distribution and Ca²⁺-ATPase activity in leaf cells. Our results demonstrated an increased calcium concentration, as well as intact chloroplast structure with increasing calcium precipitates in the cell wall, intercellular space, and vacuole of leaf cells in the plants grown in alkaline soils supplied with FGDB or CaSO₄. Additionally, a dose-dependent Ca²⁺-ATPase activity was detected in the plasma membrane and tonoplast of leaf cells from the plants grown in FGDB or CaSO₄ supplemented soils. These results implied that the Ca²⁺-ATPase activity cause cytosolic Ca²⁺ efflux. The Ca²⁺ influx is through the Ca²⁺-channels, and increasing cytosolic Ca²⁺ concentration might benefit the stability and integrity of cell membrane and cell wall, sequentially alleviated the injury of oil sunflower against alkali stress.     

The calmodulin-stimulated (Ca2+ + Mg2+)-ATPase in hemoglobin S erythrocyte membranes: effects of sickling and oxidative agents

A decrease in the reactivity of erythrocyte membrane (Ca2+ + Mg2+)-ATPase to calmodulin stimulation has been observed in aging red cells and in various types of hemolytic anemias, particularly in sickle red cell membranes. Unlike the aging process, the defect in the (Ca2+ + Mg2+)-ATPase from SS red blood cells is not secondary to a decrease in calmodulin activity and is already present in the least dense SS red blood cells separated on a discontinuous density gradient. Deoxygenated AS red cells were forced to sickle by lowering the pH, raising the osmolarity of the buffer (sickling pulse). Under these conditions an inhibition of the calmodulin-stimulated enzyme was observed only if several cycles of oxygenation/deoxygenation were applied. No alteration of the enzyme could be detected after submitting AS red blood cells to other conditions or in AA red blood cells submitted to the same treatments. This suggests that oxidative processes are involved in the alterations of the (Ca2+ + Mg2+)-ATPase activity. Treatment of membranes from AA erythrocytes by thiol group reagents and malondialdehyde, a by-product of auto-oxidation of membrane unsaturated lipids and a cross-linking agent of cytoskeletal proteins, led to a partial inhibition of the calmodulin-stimulated (Ca2+ + Mg2+)-ATPase. We postulate that the hyperproduction of free radicals described in the SS red blood cells and involved in the destabilization of the membrane may be also responsible for the (Ca2+ + Mg2+)-ATPase failure.     

Two Ca2+-dependent ATPases in rat liver plasma membrane. The previously purified (Ca2+-Mg2+)-ATPase is not a Ca2+-pump but an ecto-ATPase

We have shown that the rat liver plasma membrane has at least two (Ca2+-Mg2+)-ATPases. One of them has the properties of a plasma membrane Ca2+-pump (Lin, S.-H. (1985) J. Biol. Chem. 260, 7850-7856); the other one, which we have purified (Lin, S.-H., and Fain, J.N. (1984) J. Biol. Chem. 259, 3016-3020) and characterized (Lin, S.-H. (1985) J. Biol. Chem. 260, 10976-10980) has no established function. In this study we present evidence that the purified (Ca2+-Mg2+)-ATPase is a plasma membrane ecto-ATPase. In hepatocytes in primary culture, we can detect Ca2+-ATPase and Mg2+-ATPase activities by addition of ATP to the intact cells. The external localization of the active site of the ATPase was confirmed by the observation that the Ca2+-ATPase and Mg2+-ATPase activities were the same for intact cells, saponin-treated cells, and cell homogenates. Less than 14% of total intracellular lactate dehydrogenase, a cytosolic enzyme, was released during a 30-min incubation of the hepatocytes with 2 mM ATP. This indicates that the hepatocytes maintained cytoplasmic membrane integrity during the 30-min incubation with ATP, and the Ca2+-ATPase and Mg2+-ATPase activity measured in the intact cell preparation was due to cell surface ATPase activity. The possibility that the ecto-Ca2+-ATPase and Mg2+-ATPase may be the same protein as the previously purified (Ca2+-Mg2+)-ATPase was tested by comparing the properties of the ecto-ATPase with those of (Ca2+-Mg2+)-ATPase. Both the ecto-ATPase and the (Ca2+-Mg2+)-ATPase have broad nucleotide-hydrolyzing activity, i.e. they both hydrolyze ATP, GTP, UTP, CTP, ADP, and GDP to a similar extent. The effect of Ca2+ and Mg2+ on the ecto-ATPase activity is not additive indicating that both Ca2+- and Mg2+-ATPase activities are part of the same enzyme. The ecto-ATPase activity, like the (Ca2+-Mg2+)-ATPase, is not sensitive to oligomycin, vanadate, N-ethylmaleimide and p-chloromercuribenzoate; and both the ecto-ATPase and purified (Ca2+-Mg2+)-ATPase activities are insensitive to protease treatments. These properties indicate that the previously purified (Ca2+-Mg2+)-ATPase is an ecto-ATPase and may function in regulating the effect of ATP and ADP on hepatocyte Ca2+ mobilization (Charest, R., Blackmore, P.F., and Exton, J.H. (1985) J. Biol. Chem. 260, 15789-15794).     

Comparison of Properties of the Plasma Membrane Ca2+ -ATPase from Wheat Root and Leaf

The properties of plasma membrane Ca2 + -ATPases from wheat ( Triticum aestivum L. cv. Lengchun No. 13) root and leaf were compared, and their different properties were analyzed in association with the differentia of the functions of these two organs and their relevant environments. Root plasma membrane Ca2 + -ATPase showed a high activity in a broad range of pH and an optimum reaction temperature of 45 ℃, while the leaf enzyme activated in a narrow range of pH and an optimum reaction temperature of 50 ℃. Hill coefficient of root plasma membrane Ca2 + -ATPase for ATP was 1.6, revealing an obvious positive cooperativity. In contrast, that of leaf plasma membrane Ca2 +-ATPase was 1.0, being in keeping with Michaelis-Menten dynamics. For Ca2 + activation, Hill coefficient of plasma membrane Ca2 + -ATPases from both organs were less than 1, suggesting that both had negative cooperativity. The enzymes were activated by calmodulin and inhibited by Mg2+.     

Protein kinase C and calmodulin effects on the plasma membrane Ca2+-ATPase from excitable and nonexcitable cells

We have purified Ca2+-ATPase from synaptosomal membranes (SM)1 from ratcerebellum by calmodulin affinity chromatography. The enzyme was identifiedas plasma membrane Ca2+-ATPase by its interaction with calmodulin andmonoclonal antibodies produced against red blood cell (RBC) Ca2+-ATPase, andby thapsigargin insensitivity. The purpose of the study was to establishwhether two regulators of the RBC Ca2+-ATPase, calmodulin and protein kinaseC (PKC), affect the Ca2+-ATPase isolated from excitable cells and whethertheir effects are comparable to those on the RBC Ca2+-ATPase. We found thatcalmodulin and PKC activated both enzymes. There were significantquantitative differences in the phosphorylation and activation of the SMversus RBC Ca2+-ATPase. The steady-state Ca2+-ATPase activity of SMCa2+-ATPase was approximately 3 fold lower and significantly less stimulatedby calmodulin. The initial rate of PKC catalyzed phosphorylation (in thepresence of 12-myristate 13-acetate phorbol) was approximately two timesslower for SM enzyme. While phosphorylation of RBC Ca2+-ATPase approachedmaximum level at around 5 min, comparable level of phosphorylation of SMCa2+-ATPase was observed only after 30 min. The PKC-catalyzedphosphorylation resulted in a statistically significant increase inCa2+-ATPase activity of up to 20-40%, higher in the SM Ca2+-ATPase.The differences may be associated with diversities in Ca2+-ATPase functionin erythrocytes and neuronal cells and different isoforms composition.     

脱硫废弃物对碱胁迫下水稻叶片钙分布、Ca2+-ATPase活性及抗氧化特征的影响

为了探讨脱硫废弃物提高水稻抗盐碱的作用机制,采用盆栽法,研究脱硫废弃物对碱胁迫下水稻幼苗叶片总钙含量、Ca2+分布、细胞膜Ca2+-ATPase活性及活性氧含量等的变化.结果表明:对照处理的细胞中钙颗粒零星分布于细胞壁和叶绿体中,添加脱硫废弃物和CaSO4处理的细胞质膜、细胞间隙、细胞壁和液泡中有大量的钙颗粒分布;随着脱硫废弃物和CaSO4添加量的增加,叶片总钙含量增加,质膜和液泡膜Ca2+-ATPase活性呈上升趋势,质膜透性、MDA含量和活性氧O2-产生速率呈下降趋势,SOD、POD等保护酶活性升高.添加脱硫废弃物在一定程度上能够减缓碱胁迫对水稻造成的细胞伤害,起主要作用的物质可能是其主要成分CaSO4.     

Effects of calcium and soluble cytoplasmic activator protein (calmodulin) on various states of (Ca2+ + Mg2+)-ATPase activity in isolated membranes of human red cells

(Ca2+ + Mg2+)-ATPase activity of red cells and their isolated membranes was investigated in the presence of various Ca2+ concentrations and cytoplasmic activator protein. Red cell ATPase activity was high at low Ca2+ concentrations, and low at moderate and high concentrations of Ca2+. In the case of isolated membranes, both low and moderate ca2+ concentrations produced higher (Ca2+ + Mg2+)-ATPase activity than high Ca2+ concentration. Membrane-free hemolysate containing soluble activator of (Ca2+ + Mg2+)-ATPase produced a significant increase in (Ca2+ + Mg2+)-ATPase activity only at low ca2+ concentration. Regardless of Ca2+ and activator concentrations, the enzyme activity in the membrane was lower than lysed red cells. The low level of (Ca2+ + Mg2+)-ATPase activity seen at high Ca2+ concentration can be augmented by lowering the Ca2+ concentration of EGTA in the assay medium. However, once the membrane was exposed to a high Ca2+ concentration, the activator could no longer exert it maximum stimulation at the low Ca2+ concentration brought about by addition of EGTA. This loss of activation was not attributable to the Ca2+-induced denaturation of activator protein but rather related to the alteration of (Ca2+ + Mg2+)-ATPase states in the membrane. On the basis of these data, it is suggested that only a small portion of (Ca2+ + Mg2+)-ATPase activity of isolated membranes can be stimulated by the soluble activator and that (ca2+ + Mg2+)ATPase most likely exists in various states depending upon ca2+ concentration and the presence of activator. The enzyme state exhibiting the high degree of stimulation by activator may undergo irreversible damage in the presence of high Ca2+ concentrations.     

Rabbit myocardial membrane Ca2+-adenosine triphosphatase activity: stimulation in vitro by thyroid hormone

The in vitro stimulation of human and rabbit erythrocyte membrane Ca2+-ATPase activity by physiological concentrations of thyroid hormone has recently been described. To extend these observations to a nucleated cell model, Ca2+-ATPase activity in a membrane preparation obtained from rabbit myocardium has been studied. Activity of 5'-nucleotidase in the preparation was increased 26-fold over that of myocardial homogenate, consistent with enrichment by sarcolemma. Mean basal enzyme activity in membranes from nine animals was 20.8 +/- 3.3 mumol Pi mg membrane protein-1 90 min-1, approximately 20-fold the activity described in rabbit red cell membranes. Exposure of heart membranes in vitro to L-thyroxine (T4) (10(-10)M) increased Ca2+-ATPase activity to 29.2 +/- 3.8 mumol Pi (P less than 0.001). Dose-response studies conducted with T4 showed that maximal stimulatory response was obtained at 10(-10) M). Hormonal stimulation was comparable for L-T4 and triiodo-L-thyronine (T3) (10(-10) M). Tetraiodothyroacetic acid was without biological activity, whereas triiodothyroacetic acid and D-T4, each at 10(-10) M, significantly decreased enzyme activity compared to control (basal) levels. The action of L-T4 on myocardial membrane Ca2+-ATPase activity was inhibited by trifluoperazine (100 microM) and the naphthalenesulfonamide W-7 (50-100 microM), compounds that block actions of calmodulin, the protein activator of membrane-associated Ca2+-ATPase. Radioimmunoassay revealed the presence of calmodulin (1.4 micrograms/mg membrane protein-1) in the myocardial membrane fraction and 0.35 micrograms/mg-1 in cytosol. Myocardial Ca2+-ATPase activity, apparently of sarcolemmal origin, is thus thyroid hormone stimulable. The hormonal responsiveness of this calcium pump-associated enzyme requires calmodulin.     

Cytochemical localization and quantification of plasma membrane Ca2+-ATPase activity in mollusc digestive gland cells

A cytochemical method allowing the localization and quantification of plasma membrane Ca2+-ATPase (PMCA) in frozen sections obtained from digestive gland cells of Mytilus galloprovincialis, Tapes tapes and Chamelea gallina, is presented. The method utilizes lead as a trapping agent of PO4(2-) ions released by Ca2+-ATPase activity. The amount of lead sulphide precipitate proportionally related to PMCA activity was quantified by a light microscopy digital imaging analysis system. The optimal assay conditions of Ca2+-ATPase activity evaluated at pH 7.4 were: 200 microM free Ca2+, 200 mM KCl, 2 mM ATP, and under such analysis conditions the enzyme showed a linear trend up to 60 min (at 20 degrees C). The PMCA activity was substrate specific: ADP was utilized only at a low rate (24% with respect to an equimolar ATP concentration), while glucose-6-phosphate and beta-glycerophosphate were poorly hydrolyzed. The enzyme activity was strongly inhibited by sodium ortho-vanadate. Our detection of a Ca2-ATPase activity at nanomolar concentrations of free Ca2+ suggests that we have identified a plasma membrane Ca2-ATPase involved in Ca2+ homeostasis. The Ca2+-ATPase was found to be localized in the basal part of the plasma membrane in the digestive gland cells of Mytilus galloprovincialis and Tapes tapes, but in the apical plasma membrane of Chamelea gallina. The possible implications of the different cellular distributions of PMCA activity is discussed.     

Subcellular distribution of ATP-dependent calcium transport in rat duodenal epithelium

Subcellular fractionation studies were performed to delineate plasma membrane and intracellular membrane populations which might be involved in intracellular Ca2+-homeostasis of rat small intestinal epithelial cells. After a low-speed supernatant fraction had been suspended in 5% sorbitol and subjected to equilibrium centrifugation in a zonal rotor, the Golgi and endoplasmic reticulum markers, galactosyltransferase and NADPH-cytochrome -c reductase, were concentrated in a density region designated Window II. The basal-lateral membrane marker (Na+-K+)-ATPase was concentrated in a higher-density region designated Window III. ATP-dependent Ca2+ transport was equally distributed between the two windows. Several membrane populations could be resolved from each window with good recovery of Ca2+-transport activity by a second density gradient centrifugation step. Second density gradient fractions were subjected to counter-current partitioning in an aqueous polymer two-phase system. Basal-lateral membranes, characterized by an 11-fold enrichment of (Na+-K+)-ATPase, contained ATP-dependent Ca2+-transport activity with Vmax = 3.7 nmol/mg per min and Km = 0.5 microM. A major Golgi-derived population exhibited Ca2+-transport activity with Vmax and Km values similar to those of the basal-lateral membranes. One membrane population, presumed to have been derived from the endoplasmic reticulum, contained Ca2+-transport activity with Vmax = 4 nmol/mg per min and Km = 0.5 microM. In addition to demonstrating that ATP-dependent Ca2+-transport activity has a complex distribution within enterocytes, this study raises the possibility that the basolateral plasma membranes might account for a relatively minor portion of the cell's Ca2+-pumping ability.     

Detection and localization of a Ca2+-ATPase activity in Toxoplasma gondii

Toxoplasma gondii, the agent causing toxoplasmosis, is an obligate intracellular protozoan parasite. A calcium signal appears to be essential for intracellular transduction during the active process of host cell invasion. We have looked for a Ca2+-transport ATPase in tachyzoites and found Ca2+-ATPase activity (11-22 nmol Pi liberated/mg protein/min) in the tachyzoite membrane fraction. This ATP-dependent activity was stimulated by Ca2+ and Mg2+ ions and by calmodulin, and was inhibited by pump inhibitors (sodium orthovanadate or thapsigargin). We used cytochemistry and X-ray microanalysis of cerium phosphate precipitates and immunolabelling to find the Ca2+, Mg2+-ATPase. It was located mainly in the membrane complex, the conoid, nucleus, secretory organelles (rhoptries, dense granules) and in vesicles with a high calcium concentration. Thus, Toxoplasma gondii possesses Ca2+-pump ATPase (Ca2+, Mg2+-ATPase) as do eukaryotic cells.     

毛竹"韧皮部结"发育过程中Ca2+-ATP酶的超微定位

用电镜和细胞化学技术对毛竹[Phyllostachys edulis（Carr.）H.De Lehaie]节部“韧皮部结”发育过程中Ca^2＋-ATP酶进行了超微细胞化学定位研究.结果显示：在“韧皮部结”形成期,仅细胞质膜和细胞核上具有很高的Ca^2＋-ATP酶活性;随着“韧皮部结”的发育,发育期细胞质膜上的Ca^2＋-ATP酶活性较形成期有所降低,而细胞核上仍保持较高的Ca^2＋-ATP酶活性,胞间连丝、运输小泡膜上都具有Ca^2＋-ATP酶活性;发育后期,液泡膜及内质网上也开始出现Ca^2＋-ATP酶沉积物;成熟期的“韧皮部结”细胞质膜上的Ca^2＋-ATP酶活性较发育期有所升高,并且在“韧皮部结”成熟的过程中,细胞核、内质网、胞间连丝、质体膜和细胞质降解物上始终都有较高的Ca^2＋-ATP酶活性.实验结果表明“韧皮部结”细胞具有活跃的生理代谢以及频繁的共质体运输和信息交流.     

Studies on the bivalent-cation-activated ATPase activities of highly purified human platelet surface and intracellular membranes.

Membrane-bound Ca2+-ATPases are responsible for the energy-dependent transport of Ca2+ across membrane barriers against concentration gradients. Such enzymes have been identified in sarcoplasmic reticulum of muscle tissues and in non-muscle cells in both surface membranes and endoplasmic-reticulum-like intracellular membrane complexes. In a previous study using membrane fractionation by density-gradient and free-flow electrophoresis, we reported that the intracellular membranes of human blood platelets were a major storage site for Ca2+ and involved in maintaining low cytosol [Ca2+] in the unactivated cell. In the present report we demonstrated that the intracellular membranes also exhibit a high-affinity Ca2+-ATPase which appears to be kinetically associated with the Ca2+-sequestering process. We found that both the surface membrane and the intracellular membrane exhibited a basal Mg2+-ATPase activity, but Ca2+ activation of this enzyme was confined only to the intracellular membrane. Use of Ca2+-EGTA buffers to control the extravesicle [Ca2+] allowed a direct comparison of the Ca2+-ATPase and the Ca2+-uptake process over a Ca2+ range of 0.01 microM to 1.0 mM, and it was found that both properties were maximally expressed in the range of external [Ca2+] 1-50 microM, with concentrations greater than 100 microM showing substantial inhibition. Double-reciprocal plots for the Ca2+-ATPase activity and Ca2+ uptake gave apparent Km values for Ca2+ of 0.15 and 0.13 microM respectively. However, similar plots for ATP with the enzyme revealed a discontinuity (two affinity sites, with Km 20 and 145 microM), whereas plots for the Ca2+ uptake gave a single Km value for Ca2+, 1.1 microM. Phosphorylation studies during Ca2+ uptake using [gamma-32P]ATP revealed two components of 90 and 95 kDa phosphorylated at extravesicle [Ca2+] of 3 microM. The Ca2+-ATPase activity, Ca2+ uptake and phosphorylation were all almost completely inhibited in the presence of 500 microM-Ca2+. Similar studies using mixed membranes revealed four other phosphoproteins (50, 40, 20 and 18 kDa) formed in addition to the 90 and 95 kDa components. The findings are discussed in the context of platelet Ca2+ mobilization for function and the mechanisms whereby Ca2+ homoeostasis is controlled in the unactivated cell.     

The leaf tonoplast V-H(+)-Atpase activity of a C3 halophyte Suaeda salsa is enhanced by salt stress in a Ca-dependent mode

Suaeda salsa seedlings grown in Hoagland nutrient solution were treated with different concentrations of NaCl combined with two levels of Ca2+ (0 and 20 mmol/L) to study the effect of Ca2+ nutrition on the growth and activity of leaf tonoplast V-H(+)-ATPase. Increase of Ca2+ concentration in the solution markedly increased the relative growth quantity of S. salsa seedlings and Ca2+ and K+ concentration in the leaf cell sap under NaCl stress. The leaf V-H(+)-ATPase activity was significantly increased with increasing NaCl concentration under high Ca2+ application (20 mmol/L), but little changed under Ca2+ starvation (0 mmol/L). Western blot analysis showed that the leaf V-H(+)-ATPase of S. salsa was at least composed of A, B, D and c subunits, and their protein amounts were not affected by NaCl treatments under Ca2+ starvation (0 mmol/ L) with an exception of 100 mmol/L NaCl, but increased under high Ca2+ application (20 mmol/L). There was a positive correlation between activity of V-H(+)-ATPase and the protein amounts of the subunits. The results suggest that Ca2+ nutrition played an important role in the salt tolerance of S. salsa, and that enhancement of V-H(+)-ATPase activity under salt stress was Ca2(+)-dependent.     

Abnormal response to calmodulin in vitro of dystrophic chicken muscle membrane Ca2+-ATPase activity

A skeletal muscle membrane fraction enriched in sarcoplasmic reticulum (SR) contained Ca2+-ATPase activity which was stimulated in vitro in normal chickens (line 412) by 6 nM purified bovine calmodulin (33% increase over control, P less than 0.001). In contrast, striated muscle from chickens (line 413) affected with an inherited form of muscular dystrophy, but otherwise genetically similar to line 412, contained SR-enriched Ca2+-ATPase activity which was resistant to stimulation in vitro by calmodulin. Basal levels of Ca2+-ATPase activity (no added calmodulin) were comparable in muscles of unaffected and affected animals, and the Ca2+ optima of the enzymes in normal and dystrophic muscle were identical. Purified SR vesicles, obtained by calcium phosphate loading and sucrose density gradient centrifugation, showed the same resistance of dystrophic Ca2+-ATPase to exogenous calmodulin as the SR-enriched muscle membrane fraction. Dystrophic muscle had increased Ca2+ content compared to that of normal animals (P less than 0.04) and has been previously shown to contain increased levels of immuno- and bioactive calmodulin and of calmodulin mRNA. The calmodulin resistance of the Ca2+-ATPase in dystrophic muscle reflects a defect in regulation of cell Ca2+ metabolism associated with elevated cellular Ca2+ and calmodulin concentrations.     

毛竹茎秆纤维细胞发育过程中的细胞程序性死亡

利用TUNEL检测、细胞学及细胞化学方法,对毛竹茎秆纤维细胞发育过程中的细胞程序性死亡进行了研究。在次生壁形成的早期,纤维细胞出现染色质凝聚、细胞器膨胀、液泡膜解体和细胞质泡状化等典型的细胞程序性死亡形态学特征;TUNEL检测反应呈阳性,显示此时的纤维细胞核DNA发生了片段化。此时,在纤维细胞裂解的液泡膜、降解的细胞质和凝聚的染色质上具有ATPase活性。纤维细胞质的Ca^2＋水平会随着次生壁的形成而逐渐升高,随后Ca^2＋聚集成块状。在初生壁形成后期,纤维细胞染色质上的酸性磷酸酶（APase）活性增强。随着纤维次生壁的持续增厚,ATPase、酸性磷酸酶和Ca^2＋将在裂解的细胞质和凝聚的染色质上持续存在多年。结果表明,毛竹茎秆纤维细胞的次生壁形成过程是一个主动自溶的细胞程序性死亡过程。初生壁形成后期染色质上酸性磷酸酶活性增强及次生壁形成期胞质Ca^2＋的聚集,与纤维细胞的程序性死亡密切相关。ATPase,Ca^2＋和APase参与了纤维细胞程序性死亡过程中原生质体的降解。     

Observations on the (Ca2+ plus Mg2+)-ATPase activator found in various mammalian erythrocytes.

1. A soluble activator of membrane (Ca2+ plus Mg2)-ATPase is present in hemolysates of the newborn calf and cow, the new born and adult pig as well as human erythrocytes. 2. The activator is also found in reticulocytes of the adult pig. 3. The activator obtained from any of the above species is capable of stimulating the membrane (Ca2+ plus Mg2+)-ATPases of the other species, regardless of the age of the animals. 4. The results obtained from density fractionation of human erythrocytes revealed that the soluble factor has little simulatory effect on membranes of young erythrocytes from which it is derived but caused a marked stimulation on (Ca2+ plus Mg2+)-ATPase activity of the intermediate aged and old erythrocyte membranes. 5. The above observations support the following conclusions: (a) the extremely low levels of (Ca2+ plus Mg2+)-ATPase in cow erythrocytes is not due to the lack of a (Ca2+ plus Mg2+)-ATPase activator; (b) the distribution of (Ca2+ plus Mg2+)-atpase activator is not species specific and the differences in the level of membrane (Ca2+ plus Mg2+)-ATPase activity in various species of cells is an inherent property of that particular membrane (c) the (Ca2+ plus Mg2+)-ATPase activator is present at least from the time of reticulocyte formation and remain during tthe life span of the erythrocyte.