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1.
Calpain对细胞骨架蛋白tau降解作用的研究(英) 总被引:4,自引:0,他引:4
Calpain是钙依赖性中性蛋白酶,根据其对钙敏感性的不同,可分为m-和μ-calpain两型.分别用不同浓度CaCl2溶液孵育Wistar大鼠脑皮质匀浆液,并用蛋白质印迹和定量图像分析技术检测不同亚型calpain对tau蛋白的降解作用.研究发现:在37℃用1 mmol/L Ca2+孵育底物15 min,可见tau蛋白明显降解,并在分子质量为29 ku处出现tau蛋白降解片段;当Ca2+浓度为5 mmol/L时,tau蛋白几乎全部被降解;这种tau蛋白降解可被calpain特异性抑制剂完全逆转.进一步的研究发现,分别用μ-calpain抑制剂(0.05 μmol/L calpastatin),m-calpain抑制剂(100 μmol/L calpain inhibitor Ⅳ)或总calpain抑制剂(552 μmol/L calpeptin)与1 mmol/L Ca2+共同孵育Wistar大鼠脑皮质匀浆液,Ca2+激活的tau蛋白降解分别被抑制8.6%,92.5%和97.8%.结果表明一定浓度的Ca2+可同时激活μ-calpain和m-calpain,这两种亚型calpain均参与降解tau蛋白,但m-calpain的作用比μ-calpain更强. 相似文献
2.
Lauren L. Patton Steven Pollack Robert B. Wellner 《In vitro cellular & developmental biology. Animal》1991,27(10):779-785
Summary Salivary electrolyte secretion is under the control of the autonomic nervous system. In this paper we report that HSY, an
epithelial cell line derived from the acinar-intercalated duct region of the human parotid gland, responds to muscarinic-cholinergic
(generation of Ca2+ signal) andβ-adrenergic (generation of cAMP signal), but not toα-adrenergic (lack of Ca2+ signal), receptor stimulation. The muscarinic response was studied in detail. Carbachol (10−4
M, muscarinic agonist) or A23187 (5 μM, calcium ionophore) stimulation of HSY cells increases both86Rb (K+) influx and efflux, resulting in no change in net equilibrium86Rb content. Atropine (10−5
M, muscarinic antagonist) blocks both the carbachol-generated Ca2+ signal and carbachol-stimulated86Rb fluxes, but has no effect on either the A23187-generated Ca2+ signal or A23187-stimulated86Rb fluxes. Carbachol- and A23187-stimulated86Rb fluxes are substantially inhibited by two K+ channel blockers, quinine (0.3 mM) and scorpion venom containing charybdotoxin (33 μg/ml). The inhibition of these stimulated fluxes by another K+ channel blocker, tetraethylammonium chloride (5 mM), is less pronounced. Protein kinase C (PKC) seems to be involved in the regulation of the86Rb fluxes as 10−7
M PMA (phorbol ester, phorbol-12-myristate-13-acetate) substantially inhibits the muscarinic-stimulated86Rb efflux and influx. Because this concentration of PMA totally inhibits the carbachol-generated Ca2+ signal and only 80% of the muscarinic-stimulated86Rb influx, it seems that a portion of the carbachol-stimulated86Rb flux (i.e. that portion not inhibited by PMA) might occur independently of the Ca2+ signal. PMA fails to inhibit the A23187-stimulated86Rb fluxes, however, suggesting that PKC regulates Ca2+-sensitive K+ channel activity by regulating the Ca2+ signal, and not steps distal to this event. 4-α-Phorbol-12,13-didecanoate, a phorbol ester which fails to activate PKC, fails to inhibit either the carbachol-stimulated
increase in intracellular free Ca2+, or carbachol-stimulated86Rb fluxes. 相似文献
3.
Zinc ions are essential, but at elevated concentrations, they also have toxic effects on mammalian cells. Zinc plays a crucial
role in cell proliferation and differentiation and it even protects cells against apoptosis caused by various reagents. On
the other hand, zinc at high concentrations causes cell death that was characterized as apoptotic by internucleosomal DNA
fragmentation, formation of apoptotic bodies, and breakdown of the mitochondrial membrane potential. In the present work,
a clone of rat C6 glioma cells that was resistant to toxic effects of ZnCl2 up to 250 μM was employed to study the effect of the ionophore A23187 on zinc-induced apoptosis. Neither 150 μM Zn2+ nor 100 nM A23187 alone caused apoptosis as measured by internucleosomal DNA fragmentation. However, combined exposure of C6 cells to
100 nM A23187 and 150 μM Zn2+ for 48 h was effective in inducing apoptosis. Because the so-called calcium ionophore A23187 is not specific for Ca2+ ions but also transports Zn2+ with high selectivity over Ca2+, we investigated whether this substance promoted the uptake of Zn2+ ions into C6 cells. Employing the zinc-specific fluorescence probe Zinquin, we observed that the very low concentration of
1.9 nM A23187 significantly and rapidly raised the intracellular mobile Zn2+ content. Analysis by atomic absorption spectroscopy revealed that incubation with 1.9 nM A23187 caused a doubling of the total intracellular zinc level within 60 min. We conclude that the apoptosis evoked by the
combined action of Zn2+ and A23187 was the result of enhanced Zn2+ influx evoked by the ionophore, resulting in higher intracellular zinc levels. 相似文献
4.
张光毅 《生物化学与生物物理进展》1991,18(4):290-293
本文报道了CaM依赖性磷酸化和脱磷酸化对牛脑63kD PDE同工酶活性的调节作用。实验结果如下:(1)在存在Ca2+和CaM时,提纯的牛脑Ca2+/CaM-PK Ⅱ能催化牛脑63kD PDE同工酶磷酸化。最大磷酸参入量是1mol/mo1亚基;(2)在Ni2+和CaM存在时,提纯的牛脑钙调神经磷酸酶能催化磷酸化型63kDPDE同工酶脱磷酸化;(3)CaH2+对磷酸化型63kD PDE同工酶的半激活浓度(AC50)高于非磷酸化型。 相似文献
5.
用单细胞阳离子测定系统研究了SeO2-3对巨噬细胞内游离Ca2+和Mg2+的影响.实验结果表明:SeO2-3高于10-4mol/L时,有显著的细胞毒性.SeO2-3对细胞的毒性作用使细胞内游离Ca2+和Mg2+的浓度升高但Ca2+浓度的升高速率比Mg2+快.还有,高于10-4mol/L的SeO2-3对红细胞膜上的Ca2+-ATP酶活性有明显抑制作用. 相似文献
6.
以猪脑为材料,经匀浆、差速离心、蔗糖密度梯度离心分离突触体. 低渗破膜得到突触体膜. Triton X-100增溶后,经钙调蛋白亲和层析可得去脂的质膜Ca2+-ATPase. 用大体积亲和柱和大体积低Ca2+淋洗液淋洗,可得产率、纯度和活性均较高的质膜Ca2+-ATPase. 与大豆磷脂保温后,去脂的Ca2+-ATPase的水解活力可恢复达3.32 μmol/(mg·min).SDS-聚丙烯酰胺凝胶电泳银染显示单一蛋白质带,分子质量约为140 ku,纯度在90%以上. 不同Ca2+浓度明显影响酶的活力. 相似文献
7.
用Fura-2双波长荧光法测定神经细胞内游离钙 总被引:9,自引:0,他引:9
采用新型Ca2+荧光指示剂Fura-2建立双波长荧光法测定分离的大鼠神经细胞内游离钙浓度([Ca2+]i).结果显示,在静息状态下,其[Ca2+]i为109±12nmol/L.30mmol/L KCl可显著增加[Ca2+]i,并且KCl的这种效应呈一定的剂量依赖关系,提示该法灵敏、可靠. 相似文献
8.
Mitochondria from Trigonella foenum-graecum seedlings grown independently in the presence of either selenium (0.75 ppm) or mimosine (0.1 mM) exhibited respiration-stimulated energy-dependent uptake of Ca2+. Uptake studies were carried out independently at a series of Ca2+ concentrations at two different levels: (1) 1–20 μM and (2) 25–1500 μM. Levels of uptake were 50–100% higher in the mitochondria of seedlings of both the Se and mimosine groups. Detailed kinetic
analyses revealed negative cooperative effects operative during uptake of Ca2+ at 25–1500 μM given in the medium. Hill coefficients for Ca2+ uptake by the mitochondria of different groups remained unchanged (nH, 0.75). Biphasic Scatchard plots were concave upward,
suggestive of two classes of binding sites. High-affinity binding sites were estimated to be 16 nmol/mg protein with dissociation
constant (K
Ca) of 2.5×109 L/mol. In contrast, graphical analyses of the uptake of Ca2+ in the range 1–20 μM in the medium revealed cooperative effects of positive nature. The present study demonstrates mixed
cooperative effects during Ca2+ uptake by mitochondria from seedlings of T. foenum-graecum 相似文献
9.
钙调素(CaM)拮抗剂三氟拉嗪(TFP)在10~100 μmol/L浓度范围内对豚鼠精子获能和顶体反应具有极显著的促进作用,且不依赖于外源Ca2+的存在.Forskolin,dbcAMP和咖啡因对非同步培养系统获能早期豚鼠精子的顶体反应有显著的促进作用,但对同步培养系统获能早期精子的顶体反应无显著影响.若在同步培养系统获能培养较长时间(15h)使精子获能,则这3种药物对顶体反应均具有显著的促进作用,且不依赖外源Ca2+的存在.蛋白激酶C(PKC)激活剂PMA和PDB对获能前期豚鼠精子的顶体反应无显著影响,但对获能精子的顶体反应有显著的促进作用,且这种促进作用也不依赖于外源Ca2+的存在.相反,PKC抑制剂stau-rosporine对顶体反应有显著的抑制作用. 相似文献
10.
硒对NO诱导的内皮细胞内游离钙离子浓度变化的影响 总被引:2,自引:0,他引:2
用Fura-2显微荧光测钙技术,研究了用外源性一氧化氮(NO)供体S-亚硝基谷胱甘肽(GSNO)诱导的,人脐静脉内皮细胞系ECV-304细胞胞内游离钙离子浓度([Ca2+]i )升高以及硒的抑制效应.结果表明,GSNO作用于ECV-304细胞,短时间内即可导致其胞内游离钙离子浓度升高.胞外液换为无钙液或向胞外液中加入CdCl2(1 mmol/L)对GSNO引起的[Ca2+]i升高无影响.提示,GSNO刺激主要引起胞内钙库释放.而且,一氧化氮清除剂血红蛋白(Hb)对这一过程有抑制作用,说明GSNO引起的胞内钙库释放由NO介导.经亚硒酸钠(1 μmol/L)处理的细胞,其NO引起的[Ca2+]i升高幅度明显被抑制,说明NO的这种作用可能与细胞的氧化还原状态有关. 相似文献
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Recent advances in the study of Kaposi's sarcoma-associated herpesvirus replication and pathogenesis
It has now been over twenty years since a novel herpesviral genome was identified in Kaposi's sarcoma biopsies. Since then, the cumulative research effort by molecular biologists, virologists, clinicians, and epidemiologists alike has led to the extensive characterization of this tumor virus, Kaposi's sarcoma-associated herpesvirus(KSHV; also known as human herpesvirus 8(HHV-8)), and its associated diseases. Here we review the current knowledge of KSHV biology and pathogenesis, with a particular emphasis on new and exciting advances in the field of epigenetics. We also discuss the development and practicality of various cell culture and animal model systems to study KSHV replication and pathogenesis. 相似文献
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RICHARD E. NORRIS 《Botanical journal of the Linnean Society. Linnean Society of London》1991,106(1):1-40
Comprises species occurring mostly in subtidal habitats in tropical, subtropical and warm-temperate areas of the world. An analysis of the type species, V. spiralis (Sonder) Lamouroux ex J. Agardh, a species from Australia, establishes basic characters for distinguishing species in the genus. These characters are (1) branching patterns of thalli, (2) flat blades that may be spiralled on their axis, (3) width of the blade, (4) primary or secondary derivation of sterile and fertile branchlets and (5) position of sterile and fertile branchlets on the thalli. Application of the latter two characters provides an important basic method for separation of species into three major groups. Osmundaria , a genus known only in southern Australia, was studied in relation to Vidalia , and its separation from the Vidalia assemblage is not accepted. Species of Vidalia therefore are transferred to the older genus name, Osmundaria. Two new species, Osmundaria papenfussii and Osmundaria oliveae are described from Natal. Confusion in the usage of the epithet, Vidalia fimbriala Brown ex Turner has been clarified, and Vidalia gregaria Falkenberg, described as an epiphyte on Osmundaria pro/ifera Lamouroux, is revealed to be young branches of the host, Osmundaria prolifera. 相似文献
18.
JOAN VALUÈS MONTSERRAT TORRELL NÚRIA GARCIA JACAS 《Botanical journal of the Linnean Society. Linnean Society of London》2001,137(4):399-407
Fifteen chromosome counts of six Artemisia taxa and one species of each of the genera Brachanthemum, Hippolytia, Kaschgaria, Lepidolopsis and Turaniphytum are reported from Kazakhstan. Three of them are new reports, two are not consistent with previous counts and the remainder are confirmations of very scarce (one to four) earlier records. All the populations studied have the same basic chromosome number, x = 9, with ploidy levels ranging from 2x to 6x. Some correlations between ploidy level, morphological characters and distribution are noted. 相似文献
19.
肝癌中HBV和HCV基因和抗原的分布及意义 总被引:1,自引:0,他引:1
采用原位分子杂交方法检测HCV
RNA及HBV X基因;采用免疫组织化学方法研究HCV核心抗原,非结构区C33c抗原及HBxAg在肝细胞肝癌中的定位及分布.结果表明(1)HCV
RNA、HBV X基因在肝细胞肝癌组织检出率分别为40%(55/136)和82%(112/136).HCV
RNA定位于癌细胞的胞浆内,阳性细胞呈散在、灶状及弥漫分布三种形式;HBV
X基因在肝癌细胞中的分布呈胞浆型、核型及核浆型,阳性细胞也呈上述三种分布形式;(2)HCV
C33c抗原、核心抗原在肝细胞肝癌中的阳性率为81%(133/164)及86%(141/164).C33c抗原定位于癌细胞及肝细胞的胞浆内;核心抗原既定位于癌细胞核中,又可定位于胞浆中.C33c抗原阳性细胞以灶状分布为主;而核心抗原阳性细 相似文献
20.
H.-R. GREGORIUS 《Biological journal of the Linnean Society. Linnean Society of London》1984,23(2-3):157-165
For a plant selection model with frequency-independent viabilities, fertilities and selfing rates, it is shown that apart from global fixation, for certain parameter combinations a protected polymorphism and facultative fixation (either allele may become fixed according to initial frequencies) may both occur. Facultative fixation requires different selling rates for the dominant and recessive type. Protection of the polymorphism requires resource allocation for male and female function. In this connection the problem of purely genetically caused population extinction is discussed.
For general frequency dependence and regular segregation, the chances for establishment of a completely recessive gene are compared to those of a completely dominant gene. It is proven that the process of establishment of the recessive gene, despite a fitness advantage, may be considerably endangered by drift effects if random mating prevails. The recessive gene may reach the same effectivity in establishment as a dominant gene, only if the recessive homozygote mates exclusively with its own type during the period of establishment. 相似文献
For general frequency dependence and regular segregation, the chances for establishment of a completely recessive gene are compared to those of a completely dominant gene. It is proven that the process of establishment of the recessive gene, despite a fitness advantage, may be considerably endangered by drift effects if random mating prevails. The recessive gene may reach the same effectivity in establishment as a dominant gene, only if the recessive homozygote mates exclusively with its own type during the period of establishment. 相似文献