蛋白质的入核运送和它在基因表达中的调节作用

蛋白质进入细胞核是由蛋白质分子内部的核定位信号（nuclear localization signal, NLS）引导的.NLS蛋白首先与NLS受体结合,然后在多种胞浆因子及核孔复合物蛋白的作用下穿过核孔、转位入核.蛋白质上存在NLS并不一定总能够引导蛋白质入核.当NLS被修饰或遮掩时,它们便不能被核转运装置所识别.因而,NLS的遮掩被解除之前,蛋白质一直被扣留在胞浆中.以调节转录因子的入核运送来控制转录因子的活性是基因表达调节的一个新概念,也是细胞生长和分化的另一水平的调节.     

Functional Coupling of G Proteins to Endothelin Receptors Is Ligand and Receptor Subtype Specific

1. The aims of the present study were (a) to determine the identity of the G proteins with which the endothelin receptor interacts and whether this interaction is subtype specific and (b) to determine whether agonist exposure can result in specific coupling between the endothelin receptor and G proteins.2. Coupling between endothelin A (ET_A) or endothelin B (ET_B) receptors and G proteins was assessed in two fibroblast cell lines, each expressing one receptor subtype. Four ligands, ET-1, ET-3, SRTXb, and SRTXc, were used for receptor stimulation. The G protein -subunit coupled to the receptor was identified by immunoprecipitation with an antibody against the endothelin receptor and immunoblotting with specific antibodies against different G protein -subunits.3. Unstimulated ET_A and ET_B receptors (ET_AR and ET_BR, respectively) were barely coupled to G_o. The unstimulated ET_AR coimmunoprecipitated with G_i3, whereas the unstimulated ET_BR was much less strongly coupled to G_i3. The coupling of ET_BR to G_i1G_i2 -subunits was much stronger than the coupling of ET_AR to these -subunits. Stimulation with the different ET agonists also resulted in differential coupling of G proteins to the receptor subtypes. All four ligands caused a strong increase in coupling of the ET_BR to G_i3, whereas coupling of the ET_AR to this subunit was not affected by ET-1 and was even decreased by SRTXc. On the other hand, all four ligands caused a much greater increase in the coupling of ET_AR to G_q/G₁₁ than in the coupling of ET_BR to these -subunits. Ligand-induced coupling between the receptors and the G_i1 and G_i2 -subunits is similar for the two receptor subtypes. The same was true for ligand-induced coupling of the receptors to G_o, except that ET-3 increased the coupling of this -subunit to ET_BR and decreased the coupling to ET_AR. Taken together, the results of this study show that coupling between ET receptors and G proteins is ligand and receptor subtype specific.4. It remains to be established whether this diversity of receptor–G protein coupling is of relevance for the various endothelin signaling pathways and/or pathological states.     

Differential Sensitivity to Cations of the Melatonin Receptors in the Rat Area Postrema and Suprachiasmatic Nuclei

We studied the effects of cations on the binding of the melatonin (MT) agonist, 125I-MT, to MT receptors in the rat area postrema (AP) and suprachiasmatic nuclei (SCN), by using quantitative autoradiography in vitro. Ca2+ promoted agonist binding in the SCN but was without effect in the AP. Na+ induced a dose-dependent loss of agonist binding in both areas. This effect was more drastic in the SCN and also in the absence of divalent cations. The presence of 0.1-4.0 mM Ca2+ or Mg2+ partially and nonselectively reversed this Na(+)-elicited inhibition. The data agree with known cationic effects on agonist binding to other G protein-coupled receptors and deepen our understanding of the mammalian MT receptor.     

拟核结合蛋白与细菌基因的表达调控

拟核结合蛋白是细菌遗传物质组织和基因表达调控的关键. 细菌基因组压缩为致密的拟核必需有拟核结合蛋白的支撑. 拟核结合蛋白、DNA超螺旋和大分子簇在拟核的结构形成中起到重要作用,其中拟核结合蛋白最重要.拟核结合蛋白还影响细菌DNA的复制、重组、转录和修复等多个重要生理过程.作为全局调控因子,拟核结合蛋白是调控细菌适应环境变化所需基因表达的关键. 本文总结拟核结合蛋白的结构、功能和调控,特别是其在致病与非致病分枝杆菌中的差别,为寻找新药物靶标提供线索.     

DNA甲基化与基因表达调控研究进展

表观遗传修饰是指不改变DNA序列的、可遗传的对碱基和组蛋白的化学修饰,主要包括DNA甲基化、组蛋白修饰、染色质重塑以及非编码RNA等.表观遗传修饰是更高层次的基因表达调控手段.DNA甲基化是一种重要的表观遗传修饰,参与基因表达调控、基因印记、转座子沉默、X染色体失活以及癌症发生等重要生物学过程.近年来随着研究方法和技术的进步,全基因组DNA甲基化的研究广泛兴起,多个物种全基因组甲基化图谱被破译,全局水平对DNA甲基化的研究不仅利于在宏观层面上了解DNA甲基化的特性与规律,同时也为深入分析DNA甲基化的生物学功能与调控奠定了基础.结合最新研究进展综述DNA甲基化在基因组中的分布模式、规律以及和基因转录的关系等.     

Regulation of Brain Nicotinic Receptors by Chronic Agonist Infusion

Several studies have demonstrated that chronic treatment with nicotine elicits an increase in the number of brain nicotinic receptors. To determine whether this effect is elicited by other nicotinic agonists found in tobacco, the effects of chronic infusion with nicotine on brain nicotinic receptors were compared with those after anabasine and lobeline. C57BL/6 mice were infused with saline or equimolar doses (18.5 mumol/kg/h) of nicotine, anabasine, or lobeline for 8 days. Nicotinic receptors, quantified by the binding of [3H]nicotine and [125I]iodo-alpha-bungarotoxin (alpha-[125I]BTX), and muscarinic receptors, quantified by the binding of [3H]quinuclidinyl benzilate ([3H]QNB), were then assayed in eight brain regions. An increase in [3H]nicotine binding was observed in all regions except cerebellum following chronic infusion with nicotine and anabasine, whereas lobeline did not alter the number or affinity of these binding sites. This increase was due to changes in Bmax and not in the affinity of the receptor for the ligand (KD). A slight increase in alpha-[125I]BTX binding was observed in cortex following chronic anabasine infusion. [3H]QNB binding sites were largely unaltered following chronic infusion with any of the nicotinic analogs. The levels of the agonists in the brain were also determined after chronic treatment, and the amounts of lobeline and anabasine were found to be higher than that of nicotine. Thus, the failure of lobeline to elicit changes in nicotine binding is not due to reduced brain concentrations.     

microRNA分子表达与调控研究的最新进展

microRNA(miRNA)是一类分子长度为19~24nt的微小RNA,通常在转录后水平调控靶基因的降解或抑制翻译。miRNA分子在进化上高度保守,已经发现越来越多的miRNA分子参与真核生物的生长发育、生理活性、细胞增殖、组织分化、细胞凋亡、复杂疾病调控等功能。通过介绍miRNA的起源、合成、修饰、细胞表达特点,以及对真核细胞调控等的最新进展与研究方法,阐述miRNA在基因表达调节中的重要地位及应用前景。     

Interactions of Muscarinic Receptors with the Heterotrimeric G Proteins Gq and G12: Transduction of Proliferative Signals

Abstract: The proliferative and transforming properties of m2 and m5 muscarinic acetylcholine receptors and a series of wild-type, chimeric, and mutant G proteins were measured alone or in combination in NIH 3T3 cells to determine which G proteins mediate these signals and to what extent these signals can be influenced by changing the stoichiometry of receptors and G proteins. Responses were measured using the focus-forming assay and a novel assay called R-SAT (R eceptor S election and A mplification T echnology) in which proliferative responses are monitored using a reporter gene. Individually, GTPase-deficient mutants (^?) of Gαq and Gα12, wild-type Gαq, and m5 were active in R-SAT. Gα12^? and m5 also induced focus formation. m2 was inactive in both assays. The ability of m5 to induce foci was significantly reduced by coexpression of Gαq^?. Synergistic effects of receptor/G protein combinations were not observed in focus-forming assays but were readily detected by R-SAT. Coexpression of Gαq with m5 induced constitutive activity in R-SAT and increased the potency of agonists at m5 by 90-fold. Gαq also evoked agonist-dependent responses from m2 but not constitutive activity. Agonist potency was increased 10-fold at m2 and decreased 15-fold at m5 when these receptors were coexpressed with Gαqi5, a chimeric G protein containing the five C-terminal residues of Gαi2, compared with coexpression with Gαq. Both Gαq and Gαqi5 had biphasic effects on the proliferative responses to m5 and m2, respectively, inhibiting responses at high agonist concentrations. Coexpression of Gα12 or Gα12i5 had no effect on the concentration-response relationships of m5, but both elicited weak responses from m2. We conclude that although Gα12 is a more potent oncogene, Gαq transduces m5-driven cellular responses. The demonstrations that proliferative responses can be elicited from a nonmitogenic receptor by altering the type and concentration of available G proteins and that constitutive responses can be induced by G proteins imply that both the magnitude and type of receptor-initiated signal can be regulated at the level of G proteins in vivo.     

Cannabinoid Receptors and Their Endogenous Agonist,Anandamide

Cannabinoids are a class of compound found in marijuana which have been known for their therapeutic and psychoactive properties for at least 4000 years. Isolation of the active principle in marijuana, 9-THC, provided the lead structure in the development of highly potent congeners which were used to probe for the mechanism of marijuana action. Cannabinoids were shown to bind to selective binding sites in brain tissue thereby regulating second messenger formation. Such studies led to the cloning of three cannabinoid receptor subtypes, CB1, CB2, and CB1A all of which belong to the superfamily of G protein-coupled plasma membrane receptors. Analogous to the discovery of endogenous opiates, isolation of cannabinoid receptors provided the appropriate tool to isolate an endogenous cannabimimetic eicosanoid, anandamide, from porcine brain. Recent studies indicate that anandamide is a member of a family of fatty acid ethanolamides that may represent a novel class of lipid neurotransmitters. This review discusses recent progress in cannabinoid research with a focus on the receptors for 9-THC, their coupling to second messenger responses, and the endogenous lipid cannabimimetic, anandamide.     

家鸡G蛋白偶联受体85基因的结构、序列与组织表达分析

G蛋白偶联受体(GPCR)超家族是细胞膜上广泛存在的一类受体,是细胞跨膜信号转导的一类重要受体分子,参与许多生理过程调节。它们中仍有很多至今尚未找到内源性配体,这类受体被称为孤儿型受体。G蛋白偶联受体85(GPR85)是GPCR超家族中孤儿型受体的一员。目前,在非哺乳类脊椎动物中,针对GPR85的研究极少。本研究以家鸡Gallus gallus domesticus为模型,通过反转录PCR和RACE-PCR等方法从脑中克隆到GPR85基因的cDNA全长序列,揭示其基因结构,并用实时荧光定量PCR(qPCR)方法探究了该基因在家鸡各组织中的表达情况。结果显示:家鸡GPR85基因位于1号染色体上,由2个外显子组成,其编码区位于第2个外显子上,长为1 113 bp,可编码1个370个氨基酸的7次跨膜受体蛋白。家鸡GPR85与其他脊椎动物(人Homo sapiens、小鼠Mus musculus、大鼠Rattus norvegicus、热带爪蟾Xenopus tropicalis和斑马鱼Danio rerio)的GPR85具有高度的氨基酸序列一致性(>93%)。qPCR分析发现,GPR85基因mRNA在家鸡全脑、垂体、肾上腺、精巢中有较高表达,而在所检测的其他外周组织中表达极低。本研究首次揭示了家鸡GPR85基因的结构与表达特征,为后续探究GPR85基因在家鸡等非哺乳类中的生理功能奠定基础。     

Age-Associated Changes in Pineal Adrenergic Receptors and Melatonin Synthesizing Enzymes in the Wistar Rat

The nocturnal stimulation of pineal melatonin synthesis and elevation of serum melatonin is known to be reduced in old age in several species. In Wistar rats the capacity of the beta-adrenoceptor to develop supersensitivity (increase in Bmax) during the light period of the diurnal light/dark cycle is lost during maturation (3-6 months) rather than old age. Further, the present study shows that neither the alpha 1- nor beta-adrenoceptor density of the pineal declines as rats age. Pineal hydroxyindole-O-methyltransferase activity does fall (17-55%) in rats after 18 months of age, but nocturnal pineal arylalkylamine N-acetyltransferase activity is not significantly altered. Thus, from examination of these parameters across the life span of the rat, it seems likely that the reported reduction in serum melatonin in old animals is related to a reduced capacity of the pineal to synthesize melatonin, rather than an altered responsiveness of the gland to neural stimulation.     

水稻种子储藏蛋白及其基因表达

作为人类氮素营养的一个重要来源,水稻种子储藏蛋白的组成、结构及其合成过程一直是为研究者所关注。随着研究的深入,对于谷蛋白基因的结构特点,表达方式以及与谷蛋白基因表达相关的转录因子也都为人们所逐渐了解。这些知识对于人们改善水稻籽粒的品质以及利用水稻籽粒来生产外源蛋白都具有十分重要的意义。本文对这些方面做一简要概述。 As an important nitrogen source of human being. The composition, structure and synthesis of rice storage protein were concerned by scientists. Now people know a lot about the structure ,expression patterns of rice glutelin genes. On the basis of these knowledge, we may improve the quality of rice grain and use it to produce foreign proteins. In this paper we summarize the knowledge about rice storage proteins that we have got in these years.     

定点诱变G6PD基因的表达及突变酶的生物学功能研究

首次将9种人工定点诱变的G6PD基因转化至G6PD缺陷的大肠杆菌HB351(DE3)中表达,并对突变酶的生物学功能进行研究。初步证实G6PD基因m1376 G to T(Arg 459keu),1388 G to A(Arg 463 His)突变可降低酶活性并引起酶动力学改变。这可能与取代氨基酸的化学结构、所带电荷的性质及极性有关。这两个部位的精氨酸在酶与NADP~ 的结合过程中亦起到重要作用。赖氨酸取代精氨酸对酶与NADP~ 的结合影响不大。引入无义突变,证实G6PD第459位以后的氨基酸对酶活性有重要影响。     

囊胚形成的基因表达与调控(英文)

囊胚形成是胚胎早期发育过程中一个重要阶段 ,涉及几个重要的生理事件 ,即细胞融合 (compaction ,亦称致密化作用 )、囊胚腔出现、囊胚腔扩张及滋养层和内细胞团的分化。在细胞间连接蛋白的作用下 ,各种细胞间连接方式逐步建立起来 ,在合子型基因组表达调控下 ,促进了最终囊胚的形成。细胞间连接蛋白和细胞粘附相关蛋白参与组建各种细胞间连接 ,参与细胞融合、囊胚腔形成、滋养层分化和囊胚扩张等过程。通过顶部的紧密连接、侧部的缝隙连接和桥粒 ,建立起细胞的连接复合体。在人胚胎 8 细胞之前 ,卵裂球细胞界限明显 ,可能以中间连接方式相互作用 ;8 细胞期发生致密化作用 ,通过紧密连接将细胞分成顶部和基部 ,使得胚胎处于半封闭状态 ,促进胚胎内部积液 ,形成囊胚腔。细胞融合的同时也产生缝隙连接。桥粒最初出现在人胚胎达到 3 2 细胞阶段 ,桥粒连接参与囊胚腔形成以及在囊胚扩张时维持滋养层的稳定性。桥粒由一些跨膜粘蛋白组成 ,包括参与细胞内粘附的桥粒子和桥粒球以及一些细胞质内蛋白 (如desmoplakins,plakoglobin ,plakophilin) ,由细胞内蛋白质形成空斑结构并介导细胞角蛋白丝固定。对植入前牛胚胎的研究表明 ,只有DcII,DcIII和plako三种桥粒蛋白参与桥粒组建。在鼠囊胚中DcII的表达部位位于     

Strict Regulation of Gene Expression Utilizing a Dual-control Gene Expression Vector System

可严格调控性是体现原核表达载体优越性的重要指标。构建了一种双控双调节原核表达载体系统,用双载体控制调节目的基因的表达,即SP6启动子（promoter）＋乳糖（lac）调节基因表达系统和araB启动子（promoter）＋ara C调节基因表达系统,分别由乳糖类似物IPTG和阿拉伯糖（L-arabinose）诱导目的基因的表达。该系统由2个表达载体共同完成目的基因的表达。pE SP-1为主表达载体,即目的基因克隆到此表达载体上,由SP6启动子（promoter）＋乳糖（lac）调节基因调控;pA RA-SP6为辅助表达载体,该载体通过SP6 RNA聚合酶的表达来控制调节主表达载体的启动子（SP6）,由araB启动子（promoter）＋ara C调节基因调控。实验结果显示该双控双调节表达载体系统控制严格,并且表达蛋白的量具有可调控性。     

Anion Regulation of Agonist and Inverse Agonist Binding to Benzodiazepine Receptors

Binding of the benzodiazepine inverse agonist [3H]methyl-6,7-dimethoxy-4-ethyl-beta-carboline-3-carboxylate [( 3H]DMCM) and the agonist [3H]flunitrazepam [( 3H]FNZ) was compared in rat cortical membranes. Halide ions enhanced [3H]DMCM binding three- to fourfold, increasing both the apparent affinity and the number of binding sites for this radioligand. The effect was present at both 0 and 37 degrees C. In contrast, the magnitude of halide stimulation of [3H]FNZ binding was much smaller, resulting solely from an increase in the apparent affinity for this radioligand, and was not observed at 37 degrees C. The potencies but not the efficacies of a series of anions to stimulate both [3H]DMCM and [3H]FNZ binding to benzodiazepine receptors were highly correlated with their relative permeabilities through gamma-aminobutyric acid (GABA)-gated chloride channels. Two stress paradigms (10 min of immobilization or ambient-temperature swim stress), previously shown to increase significantly the magnitude of halide-stimulated [3H]FNZ binding, did not significantly affect [3H]DMCM binding. Phospholipase A2 treatment of cortical membrane preparations was equipotent in preventing the stimulatory effect of chloride on both [3H]DMCM and [3H]FNZ binding. These data strongly suggest that anions modify the binding of [3H]DMCM and [3H]FNZ by acting at a common anion binding site that is an integral component of the GABA/benzodiazepine receptor chloride channel complex.     

植物酸性转化酶基因及其表达调控

酸性转化酶是蔗糖代谢的关键酶,在植物体中具有重要的生理作用.近十几年来,许多植物酸性转化酶基因已经克隆,其基因表达调控的研究也取得了很大的进展.本文综述了植物酸性转化酶基因及其蛋白结构、基因表达的器官和发育特异性以及糖、受伤、病原、胁迫和激素对基因表达的调节和蛋白抑制因子对酶活性的影响,并讨论了当前在该研究领域存在的问题.