Role of Nonhistone Chomosomal Proteins in the Restriction of Mitotic Chromatin Template Activity

RNA synthesis virtually ceases during mitosis in eukaryotic cells. This phenomenon is explainable, at least in part, by the reduced template activity for RNA synthesis of mitotic chromatin. The restriction in template activity can be accounted for by the presence in the chromatin of mitotic-specific nonhistone proteins. Chromatin reconstituted by gradient dialysis from the pooled histones of S-phase and mitotic chromatin and the nonhistone proteins of mitotic chromatin had a template activity similar to that of native mitotic chromatin, and lower than native S-phase chromatin or chromatin reconstituted with the pooled histones and the S-phase and nonhistone proteins. The template activity was identical for chromatin reconstituted from the pooled nonhistone proteins and either the histones from S-phase or mitotic chromatin. These results are supported with data showing that the procedure of reconstitution produces a chromatin indistinguishable in its protein composition both qualitatively and quantitatively from its native counterpart.     

Nonhistone Chromosomal Proteins in Synchronized HeLa Cells

Chromatin was isolated from synchronized HeLa cells at different stages of the cell division cycle and fractionated into DNA, histones, and nonhistone proteins. Electrophoresis of the nonhistone proteins in sodium dodecyl sulfate-polyacrylamide gels revealed a highly reproducible pattern of 22 bands, having estimated molecular weights of 15,000-180,000, with 85% (by mass) over 40,000. The amounts of some nonhistone proteins varied during the cell cycle by as much as 50%, while others remained at a constant level. One group of nonhistone proteins (molecular weight 75,000) was greatly reduced just before the start of DNA replication (S-phase), then returned to normal levels in the mid-S phase. These results are discussed with regard to the possible role of nonhistone proteins in regulating chromosome structure and function.     

心房颤动中钙耦联蛋白表达的实验研究

采用逆转录聚合酶链式反应 (RT PCR)方法研究起搏房颤 (AF)模型心房肌钙耦联蛋白———L型钙通道α1c亚基和肌浆网Ca2 + ATP酶的基因表达情况以及药物的干预作用。选择成年杂种犬 15条 ,随机分为起搏组 (P组 )、起搏及口服维拉帕米组 (PV组 )和对照组 (C组 )。采用心房猝发刺激诱发AF。其中P组和PV组的犬在诱发AF后置入起搏脉冲发生器作心房快速 ( 4 0 0次 /分 )起搏 7天。PV组的犬于起搏第 2天起给予维拉帕米缓释剂 12 0mg/天口服。第 7天再次诱发AF后将动物处死 ,于左、右心房和房间隔部位取材进行RT PCR扩增。结果 :心房快速起搏 7天后 ,阵发性AF和持续性AF的发生率明显增加。而加用维拉帕米后AF的发生率和起搏前没有差异。以 β actin为内参照的RT -PCR结果显示起搏组心房肌的L型钙通道α1c亚基和肌浆网Ca2 + ATP酶的mRNA水平明显降低(和C组相比分别降低 4 3 %和 4 0 %,P <0 .0 1)。而PV组第 7天心房肌L型钙通道α1c亚基和Ca2 + ATP酶的mRNA水平和C组相比则没有显著差异 (P >0 .0 5 )。心房不同部位之间的mRNA水平没有差异。结论 :心房起搏AF模型中心房肌细胞钙耦联蛋白L型钙通道α1c亚基和肌浆网Ca2 + ATP酶的基因表达明显下调 ;钙通道阻断剂维拉帕米可预防心房电重构和心房肌钙耦联蛋白的下调。     

Host Control of Endogenous Murine Leukemia Virus Gene Expression: Concentrations of Viral Proteins in High and Low Leukemia Mouse Strains

Two of the major molecular components of murine leukemia virus particles, the internal protein (p30) and the envelope glycoprotein (gp69/71) have been measured in the spleens of normal, 6- to 10-week-old mice of various inbred strains and crosses. Both proteins were detected in virtually all mice. Extracts of high leukemia, high murine leukemia virus strains (AKR, C58, PL) showed high levels of both proteins; extracts of other strains usually showed lower levels. Of particular interest, however, were the exceptions to these general observations: (1) Very little gp69/71 could be detected in spleens of BALB mice, and this trait was dominant in crosses with AKR and DBA/2, both of which express a high level of gp69/71. Thymus-deficient BALB/c-nu/nu (nude) mice, in contrast, showed a higher concentration of gp69/71 typical of other low leukemia strains, suggesting that the virtual absence of the protein in normal BALB/c mice may result from immunologic suppression. (2) With C57L, C57BL/10Sn, and C57BL/6 strains the concentration of p30 was lower, in some cases much lower, than would be expected from the concentration of gp69/71. (3) DBA/2 mice showed high levels of gp69/71, 10-fold greater than that of p30, whereas congenic DBA/2-Fv-1(b) mice showed the opposite pattern. (4) Mice of 129-G(IX) (-) strain showed no detectable levels of either p30 or gp69/71 proteins, although the congenic 129 (G(IX) (+)) showed appreciable levels of both. The absence of these proteins in 129-G(IX) (-) mice is a recessive trait, as F(1) hybrids with AKR and DBA/2 showed appropriate levels of both proteins. It is concluded that expression of viral p30 and gp69/71 proteins in mice is not coordinately linked and that expression is complex, being influenced by several genes, including Gv-1, H-2, Fv-1, and probably still others.     

Stringent Control of Ribosomal Protein Gene Expression in Escherichia coli

The regulation of the expression of ribosomal protein genes was examined during partial inhibition of tRNA aminoacylation in isogenic rel(+) and rel(-) strains of E. coli B carrying a temperature-sensitive mutation in valyl-tRNA synthetase (EC 6.1.1.9) gene. Measurements of the differential synthesis rate of ribosomal protein, alpha(r) (ribosomal protein synthesis rate/total protein synthesis rate), indicate that ribosomal protein synthesis is regulated directly or indirectly by availability of charged tRNA, and that the synthesis of ribosomal protein, like the synthesis of rRNA, is subject to the influence of the rel gene control system.     

Gene Expression of Atrial Calcium-Handling Proteins in Patients with Rheumatic Heart Disease and Atrial Fibrillation

Objectives To investigate the gene expression of calcium - handling proteins in patients with rheumatic heart disease (RHD) and atrial fibrillation (AF) . Methods A total of 50 patients with rheumatic mitral valve disease were included. According to cardiac rhythm and duration of episode of AF, patients were divided into four groups: sinus rhythm group, paroxysmal AF group, persistent AF for less than 6 months group and persistent AF for more than 6 months group. Atrial tissue was obtained from the right atrial appendage, the right atrial free wall and the left atrial appendage respectively during open heart surgery. Total RNA was isolated and reversly transcribed into cDNA. In a semi - quantitative polymerase chain reaction the cDNA of interest and of glyceralde-hyde3 - phosphate dehydrogenase (GAPDH) were amplified and separated by ethidium bromide - stained gel electrophoresis. Multiple liner regress was used for correlation between the mRNA amount and age, sex, right atrial diameter (RAd) and left a     

酵母系统表达疾病相关蛋白的研究

随着分子生物学技术的发展 ,利用生物工程技术表达药用蛋白、生物活性蛋白已成为一种有潜在发展前景的生产方式 ,如全世界用于治疗糖尿病的胰岛素 ,大约有一半的用量是由酿酒酵母表达出来的。ThomasKjeldsen等用毕赤巴斯德酵母表达出了有活性的与人类天然胰岛素结构相同的胰岛素[1] 。与传统的原核表达系统相比 ,酵母真核表达系统具有易培养、繁殖快及真核细胞对翻译后蛋白的加工及修饰过程 ,如二硫键的正确形成 ;前体蛋白的水解加工 ;糖基化作用等 ,并且能将表达的蛋白产物分泌到细胞外的培养基中 ,简化了表达产物的回收和纯化 ,有利于蛋…     

Some Binding Parameters of Chromatin Acidic Proteins with High Affinity for Deoxyribonucleic Acid

A fraction of chromatin acidic protein from rat liver, predominantly comprised of low-molecular-weight species, showed highest in vitro affinity for DNA when compared with other fractions of nuclear acidic protein. Maximum binding of this protein fraction was observed at physiological ionic strength of 0.14 M NaCl and DNA/protein ratio > 1. Comparison of their binding to homologous and heterologous DNAs showed partial species specificity. All molecular species of this protein fraction appear to bind to DNA.     

十二指肠胃反流模型大鼠胃黏膜组织Cyclin D1、p16蛋白表达

目的探讨十二指肠胃反流(DGR)引起大鼠胃黏膜损伤及癌变的发病机制.方法健康成年雄性SD大鼠75只随机分为两组①DGR模型组55只.根据手术造模方式及反流量大小分为全反流组与部分反流组.②假手术对照组20只.采用pH监测仪测定胃液pH,用酶法测定胃液胆汁酸(TBA),确定DGR模型成功.进行为期3个月和9个月胃黏膜损伤的动态观察.采用免疫组化方法(S-P)分析不同病变胃黏膜组织Cycli nD1、p16蛋白的表达.结果DGR模型大鼠胃液pH及TBA明显升高(P＜0.01),证明DGR模型成功.模型大鼠病理政变出现浅表性胃炎→萎缩性胃炎→异型增生的动态演变过程.萎缩性胃炎组及异型增生时Cyclin D1蛋白表达显著高于假手术对照组及浅表性胃炎组(P＜0.05),癌基因Cyclin D1为高表达.在萎缩性胃炎与异型增生之间Cyclin D1表达差异无显著性(P＞0.05).而p16蛋白表达则相反,在假手术对照组最高,在异型增生组最低,两组之间差异有显著性(P＜0.01),抑癌基因p16在异型增生时为低表达.Cyclin D1与p16表达呈负相关(r=-0.53).结论Cyclin D1、p16蛋白表达异常是DGR胃黏膜损伤及癌变的分子机制之一.     

TFPI基因转染对血管平滑肌细胞中细胞凋亡抑制蛋白的调控

目的研究组织因子途径抑制物(TFPI)基因转染对大鼠血管平滑肌细胞中细胞凋亡抑制蛋白(IAP)表达的影响,探讨TFPI诱导细胞凋亡的可能机制。方法将含有人TFPI基因的重组腺病毒或含β-半乳糖苷酶(LacZ)基因的重组腺病毒或DMEM在体外分别转染大鼠血管平滑肌细胞,用ELISA方法测定转染后细胞中TFPI蛋白的表达水平,用RT-PCR方法测定基因转染后不同时间点细胞中c-IAP1mRNA的表达,用Western-blot方法检测基因转染后不同时间点细胞中survivin的表达。结果基因转染后1天在血管平滑肌细胞中即可检测到TFPI蛋白表达,峰值出现在转染后第3天;基因转染后3天和7天,TFPI组c-IAP1 mRNA的表达与对照组相比明显减少(P0.05);基因转染后3、5、7天,TFPI组survivin的表达与对照组相比明显减少(P0.05),且具有明显的时间依赖性。结论 TFPI可能通过抑制IAP的表达来发挥诱导平滑肌细胞凋亡的作用,从而抑制冠状动脉介入治疗后再狭窄发生。     

Autoregulation: A Role for a Biosynthetic Enzyme in the Control of Gene Expression

It was previously proposed, primarily on the basis of evidence in vitro, that L-threonine deaminase, the ilvA gene product, is required for repression of its own synthesis and for repression of the other genes in the ilv-ADE operon. In this communication, evidence in vivo is presented that supports this autoregulatory model. Further evidence is presented that suggests that L-threonine deaminase is also required for induction of the ilvC gene product. The autoregulatory model is presented in an expanded form to include recent evidence that L-threonine deaminase (EC 4.2.1.16) is a central element for repression of the ilvADE and ilvB operons, and for induction of the ilvC operon.     

Ethanol and Gene Expression in Brain

This article represents the proceedings of a symposium at the 2000 ISBRA Meeting in Yokohama, Japan. The chairs were Izuru Matusmoto and Peter A. Wilce. The presentations were (1) GABA receptor subunit expression in the human alcoholic brain, by Tracey Buckley and Peter Dodd; (2) NMDAR gene expression during ethanol addiction, by Jorg Puzke, Rainer Spanagel, Walther Zieglgansberger, and Gerald Wolf; (3) Differentially expressed gene in the nucleus accumbens from ethanol-administered rat, by Shuangying Leng; (4) Expression of a novel gene in the alcoholic brain, by Peter A. Wilce; and (5) Investigations of haplotypes of the dopamine D2-receptor gene in alcoholics, by Hans Rommelspacher, Ulrich Finckh, and Lutz G. Schmidt.     

Reconstituted HDL in Acute Coronary Syndromes

Objectives: The strong inverse relationship between plasma high‐density lipoprotein (HDL)‐cholesterol and atherosclerotic cardiovascular disease provides the epidemiological basis that HDL is atheroprotective. Since HDL enhances cholesterol efflux and exhibits potent antiinflammatory properties, the aim of the present study was to investigate whether infusion of reconstituted HDL (rHDL) impacts on vascular function, a well‐established surrogate of atherosclerotic vascular disease, as well as markers of inflammation and oxidative stress in patients with acute coronary syndromes (ACS). Methods: Twenty‐nine patients with ACS were randomized to double‐blind treatment with rHDL or albumin. Endothelium‐dependent and independent vasodilatation to intraarterial acetylcholine and sodium nitroprusside were measured by forearm venous occlusion plethysmography. In addition, oxidized LDL and high‐sensitivity C‐reactive protein were determined as markers of oxidative stress and vascular inflammation. Results: rHDL infusion increased plasma HDL (P < 0.0001) and decreased LDL (P < 0.0001). Oxidized LDL (P= 0.11), high‐sensitivity C‐reactive protein (P= 0.12) and the response to endothelium‐dependent and ‐independent vasodilatators remained unchanged after rHDL compared to albumin infusion (14.9 ± 9.2 versus 14.5 ± 12.4, P= 0.93 and 12.8 ± 7.1 versus 13.2 ± 9.6, P= 0.27, respectively). Conclusions: An increase of HDL and a reduction of LDL notwithstanding, human rHDL did not improve vascular function in patients with ACS thus further challenging the clinical benefit of interventions, which rapidly raise HDL in ACS, particularly with the infusion of reconstituted HDL.