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Enhancement of germ cell apoptosis induced by ethanol in transgenic mice overexpressing Fas Ligand

Cell Research 2003年第5期13卷 361-367页

作者： JIA HUA HU, JIE JIANG, YING HUA MA, NA YANG, MAO HU ZHANG, MIN WU, JIAN FEI, LI HE GUO 1 Laboratory of Molecular Cell Biology, Institute of Biochemistry and Cell Biology, shanghai institutes for biological {3., chinese academy of {3., shanghai 20003., china 2 College of life science, Zhejiang University, Hangzhou 3.0027, china 3.shanghai Research Center for Biomodel Organism, shanghai 201203. china 1Laboratory of Molecular Cell Biology Institute of Biochemistry and Cell Biology Shanghai Institutes for Biological Sciences Chinese Academy of Sciences Shanghai 200031 China. E-mail: mhzhang@*** 2College of life science Zhejiang University Hangzhou 310027 China 1Laboratory of Molecular Cell Biology Institute of Biochemistry and Cell Biology Shanghai Institutes for Biological Sciences Chinese Academy of Sciences Shanghai 200031 China. E-mail: mhzhang@*** 1Laboratory of Molecular Cell Biology Institute of Biochemistry and Cell Biology Shanghai Institutes for Biological Sciences Chinese Academy of Sciences Shanghai 200031 China. E-mail: mhzhang@*** 1Laboratory of Molecular Cell Biology Institute of Biochemistry and Cell Biology Shanghai Institutes for Biological Sciences Chinese Academy of Sciences Shanghai 200031 China. E-mail: mhzhang@*** 2College of life science Zhejiang University Hangzhou 310027 China 3.0} Research Center for Biomodel Organism Shanghai 201203 China 1Laboratory of Molecular Cell Biology Institute of Biochemistry and Cell Biology Shanghai Institutes for Biological Sciences Chinese Academy of Sciences Shanghai 200031 China. E-mail: mhzhang@***

It was suggested that chronic ethanol exposure could result in testicular germ cell apoptosis, but the mechanism is still unclear. In the present study, we use a model of transgenic mice ubiquitously overexpressing human FasL to investigate whether Fas ligand plays a role in ethanol-induced testicular germ cell apoptosis. Both wild-type (WT) mice and transgenic (TG) mice were treated with acute ethanol (20％ v/v) by introperitoneal injection for five times. After ethanol injection, WT mice displayed up-regulation of Fas ligand in the testes, which was shown by FITC-conjugated flow cytometry and western blotting. Moreover, TG mice exhibited significantly more apoptotic germ cells than WT mice did after ethanol injection, which was demonstrated by DNA fragmentation, PI staining flow cytometry and TUNEL staining. In addition, histopathological examination revealed that degenerative changes of epithelial component of the tubules occurred in FasL overexpressing transgenic mice while testicular morphology was

关键词： Fas ligand ethanol apoptosis testes transgenic mouse.

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Tetrahydroacridine inhibits voltage-dependent Na~+ current in guinea-pig ventricular myocytes

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Acta Pharmacologica Sinica 2004年第9期 36-42页

作者： Wei WANG, Yi-ping WANG, Guo-yuan HUState Key Laboratory of Drug Research, The First Department of Pharmacology, shanghai Institute of Materia Medica, shanghai institutes for {3. sciences, chinese academy of sciences, shanghai 201203. china

AIM: To study the effects of tetrahydroacridine (tacrine) on voltage-gated Na+ channels in cardiac tissues. METHODS: Single ventricular myocytes were enzymatically dissociated from adult guinea-pig heart. Voltage-depe... 详细信息

关键词： Alzheimer disease cholinesterase inhibitors sodium channels tetrahydroacridine (tacrine) myocardium patch-clamp techniques

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Identification of a NF-кB site in the negative regulatory element (ε-NRAII) of human ε-globin gene and its binding protein NF-кB p50 in the nuclei of K562 cells

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Cell Research 2002年第1期12卷 79-82页

作者： CHUN Hui HOU, JIAN HUANG, Ruo LAN QIAN Group of Globin Gene Expression and Regulation, State Key Labortory of Molecular Biology, Institute of Biochemistry and Cell Biology, shanghai institutes for biological {3., chinese academy of {3., shanghai Group of Globin Gene Expression and Regulation Institute of Biochemistry and Cell Biology Shanghai Institutes for Biological Sciences Chinese Academy of Sciences State Key Labortory of Molecular Biology 上海 200031 Group of Globin Gene Expression and Regulation Institute of Biochemistry and Cell Biology Shanghai Institutes for Biological Sciences Chinese Academy of Sciences State Key Labortory of Molecular Biology 上海 200031

The developmental control of the human e-globin gene expression is mediated by transcription regulatory elements in the 5’ flanking DNA of this gene. Sequence analysis has revealed a DNA motif (GGGGAATTTGCT) similar to NF-кB consensus sequence resides in the negative regulatory element (-3.28bp～ -2902bp, termed ε-NRAII) 5’ to the cap site of this gene. NRF DNA fragment (-3.10bp～ -2986bp) containing the NF-кB motif similar sequence was synthesized and used in electrophoresis mobility shift assay (EMSA) and competitive analysis. Data showed that a protein factor from nuclear extracts of K562 cells specifically interacted with NRF DNA fragment. The synthetic NF DNA fragment (containing NF-кB consensus sequence) could competed for the protein binding, but MNF DNA fragment (mutated NF-кB motif) could not, suggesting that the binding protein is a member of NF-кB/Rel family. Western blot assay demonstrated that the molecular weight of NF-кB protein in the nuclei of K562 cells is 50ku. We suggested that NF-кB p50 may play an important role in the regulation of human c-globin gene expression.

关键词： Human ε-globin gene negative regulatory element NE-кB p50.

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Maternal zinc deficiency impairs brain nestin expression in prenatal and postnatal mice

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Cell Research 2001年第2期11卷 135-141页

作者： WANG FU DI, WEI BIAN, LING WEI KONG, FA JI ZHAO, JUN SHENG GUO, NAI HE JINGI Institute of Biochemistry and Cell Biology, shanghai institutes for biological {3., chinese academy of {3., 3.0 Yue Yang Road, shanghai 20003., china Department of Publ 2Department of Public Health Second Military Medical University Shanghai 200433 China 1Institute of Biochemistry and Cell Biology Shanghai Institutes for Biological Sciences Chinese Academy of Sciences 320 Yue Yang Road Shanghai 200031 China 1Institute of Biochemistry and Cell Biology Shanghai Institutes for Biological Sciences Chinese Academy of Sciences 320 Yue Yang Road Shanghai 200031 China 2Department of Public Health Second Military Medical University Shanghai 200433 China 2Department of Public Health Second Military Medical University Shanghai 200433 China 1Institute of Biochemistry and Cell Biology Shanghai Institutes for Biological Sciences Chinese Academy of Sciences 320 Yue Yang Road Shanghai 200031 China

Effects of maternal dietary zinc deficiency on prenatal and postnatal brain development were investigated in ICR strain mice. From d 1 of pregnancy (EO) until postnatal d 20 (P20), maternal mice were fed experimental diets that contained 1 mg Zn/kg/day (severe zinc deficient, SZD), 5 mg Zn/kg/day (marginal zinc deficient, MZD), 3. mg Zn/kg/day (zinc adequately supplied, ZA) or 100 mg Zn/kg/day (zinc supplemented, ZS and pair-fed, PF). Brains of offspring from these dietary groups were examined at various developmental stages for expression of nestin, an intermediate filament protein found in neural stem cells and young neurons. Immunocytochemistry showed nestin expression in neural tube 10. 5 d post citrus (dpc) as well as in the cerebral cortex and neural tube from 10.5 dpc to postnatal d 10 (P10). Nestin immunoreactivities in both brain and neural tube of those zinc-supplemented control groups (ZA, ZS, PF) were stronger than those in zinc-deficient groups (SZD and MZD). Western blot analysis confirmed that nestin levels in -pooled brain extracts from each of the zinc-supplemented groups (ZA, ZS, PF) were much higher than those from the zinc-deficient groups (SZD and MZD) from 10.5 dpc to P10. Immunostaining and Western blots showed no detectable nestin in any of the experimental and control group brains after P20. These observations of an association between maternal zinc deficiency and decreased nestin protein levels in brains of offspring suggest that zinc deficiency suppresses development of neural stem cells, an effect which may lead to neuroanatomical and behavioral abnormalities in adults.

关键词： Zinc deficiency nestin brain development neural stem cells.

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Substitutions of the Conserved Gly47 Affect the CF1 Inhibitor and Proton Gate Functions of the Chloroplast ATP Synthase ε Subunit

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Acta Biochimica et Biophysica Sinica 2005年第7期37卷 453-462页

作者： Hui DONG Zhang-Lin NI Jia-Mian WEI Institute of Plant Physiology & Ecology,shanghai institutes for biological {3.,chinese academy of {3.,shanghai 20003.,china Institute of Plant Physiology and Ecology Shanghai Institutes for Biological Sciences Chinese Academy of Sciences Shanghai 200032 China

The conserved residue Gly47 of the chloroplast ATP synthase ε subunit was substituted withLeu,Arg,Ala and Glu by site-directed *** process generated the mutants εG47L,εG47R,εG47A and εG47E,*** the ε variants showed lower inhibitory effects on the soluble CF1（-ε）Ca2+-ATPase compared with wild-type ε.In reduced conditions,εG47E and εG47R had a lower inhibitoryeffect on the oxidized CF1（-ε）Ca2+-ATPase compared with wild-type ε.In contrast,εG47L and εG47Aincreased the Ca2+-ATPase activity of soluble oxidized CF1（-ε）.The replacement of Gly47 significantlyimpaired the interaction between the subunit ε and γ in an in vitro binding *** study showed thatall ε variants were more effective in blocking proton leakage from the thylakoid *** enhancedATP synthesis of the chloroplast and restored ATP synthesis activity of the reconstituted membranes to alevel that was more efficient than that achieved by wild-type ε.These results indicate that the conservedGly47 residue of the ε subunit is very important for maintaining the structure and function of the ε subunitand may affect the interaction between the ε subunit,β subunit of CF1and subunit Ⅲ of CF_o,therebyregulating the ATP hydrolysis and synthesis,as well as the proton translocation role of the subunit Ⅲ of CF_o.

关键词： chloroplast ATP synthase ε subunit GST pull-down assay site-directed mutagenesis

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The spike protein of severe acute respiratory syndrome (SARS) is cleaved in virus infected Vero-E6 cells

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Cell Research 2004年第5期14卷 400-406页

作者： Xiao Dong WU 1*, Bo SHANG1*, Rui Fu YANG2*, Hao YU3., Zhi Hai MA1*, Xu SHEN3., Yong Yong JI1, Ying LIN , Ya Di WU1, Guo Mei LIN1, Lin TIAN1, Xiao Qing GAN1, Sheng YANG2, Wei Hong JIANG2, 1 Er Hei DAI4, Xiao Yi WANG4, Hua Liang JIANG3. You Hua XIE1, Xue Liang ZHU1, Gang PEI1, Lin LI1, Jia Rui WU1**, Bing SUN 1** 1Institute of Biochemistry and Cell Biology, shanghai institutes for biological {3., chinese academy of {3., 3.0 Yueyang Road, shanghai 20003., china. 2Institute of Microbiology and Epidemiology, Institute of Basic Medical {3., academy of Military Medical {3., Beijing 100071, china. 3.nstitute of Plant Physiology and Ecology, shanghai institutes for biological {3., chinese academy of {3., 3.0 Fenglin Road, shanghai 20003., china. 3.nstitute of Materia Medica, shanghai institutes for biological {3., chinese academy of {3., 3.0 Taiyuan Road, shanghai 20003., china. Institute of Biochemistry and Cell Biology Shanghai Institutes for Biological Sciences Chinese Academy of Sciences 320 Yueyang Road Shanghai 200031 China.

Spike protein is one of the major structural proteins of severe acute respiratory syndrome-coronavirus. It is essential for the interaction of the virons with host cell receptors and subsequent fusion of the viral envelop with host cell membrane to allow infection. Some spike proteins of coronavirus, such as MHV, HCoV-OC43. AIBV and BcoV, are proteolytically cleaved into two subunits, S1 and S2. In contrast, TGV, FIPV and HCoV-229E are not. Many studies have shown that the cleavage of spike protein seriously affects its function. In order to investigate the maturation and proteolytic processing of the S protein of SARS CoV, we generated S1 and S2 subunit specific antibodies (Abs) as well as N, E and 3.L protein-specific Abs. Our results showed that the antibodies could efficiently and specifically bind to their corresponding proteins from *** expressed or lysate of SARS-CoV infected Vero-E6 cells by Western blot analysis. Furthermore, the anti-S1 and S2 Abs were proved to be capable of binding to SARS CoV under electron microscope observation. When S2 Ab was used to perform immune precipitation with lysate of SARS-CoV infected cells, a cleaved S2 fragment was detected with S2-specific mAb by Western blot analysis. The data demonstrated that the cleavage of S protein was observed in the lysate, indicating that proteolytic processing of S protein is present in host cells.

关键词： SARS-CoV spike protein nucleocapsid protein envelope protein 3.L protease polyclonal antibody.

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Thansgenic peanut plants obtained by particle bombardment via somatic embryogenesis regeneration system

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Cell Research 2001年第2期 156-160页

作者： DENG XIANG YANG, ZHI MING WEI, HAI LONG AN Institute of Plant Physiology and Ecology, shanghai institutes for biological {3., chinese academy {3., shanghai 20003., china 中科院上海植物生理生态所上海 200032

After pre-culture and treatment of osmosis, cotyledons of immature peanut (Arachis hypogaea L.) zygotic embryos were transformed via particle bombardment with a plasmid containing a chimeric hph gene conferring resistance to hygromycin and a chimeric intron-gus gene. Selection for hygromycin resistant calluses and somatic embryos was initiated at 10th d post-bombardment on medium containing 10-25 mg/L hygromycin. Under continuous selection, hygromycin resistant plantlets were regenerated from somatic embryos and were recovered from nearly 1.6% of the bombarded cotyledons. The presence and integration of foreign DNA in regenerated hygromycin resistant plants was confirmed by PCR (polymerase chain reaction) for the intron-gus gene and by Southern hybridization of the hph gene. GUS enzyme activity was detected in leaflets from transgenic plants but not from control, non-transformed plants. The production of transgenic plants are mainly based on a newly improved somatic embryogenesis regeneration system developed by us.

关键词： Peanut (Arachis hypogaea) somatic embryo hygromycin resistance particle bombardment.

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Effect of osmotic stress on spontaneous calcium sparks in rat ventricular myocytes

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Acta Pharmacologica Sinica 2006年第7期27卷 877-887页

作者： Hong XIE Pei-hong ZHU Unit of Cell Signal Transduction,shanghai institutes for biological {3.,chinese academy of {3.,shanghai 20003.,china UnitofCellSignalTransduction ShanghaiInstitutesforBiologicalSciencesChineseAcademyofSciencesShanghai200031China

Aim:To study whether the volume of cardiomyocytes and their functions wouldchange under severe pathological conditions or osmotic *** clarify the roleof ryanodine receptors/calcium release channels(RyRs)in the functional change,the effect of osmotic stress on spontaneous Ca2+sparks in rat ventricular myocyteswas ***:A laser scanning confocal microscope was used todetect spontaneous Ca2+sparks of intact or saponin permeabilized myocytes loadedwith *** and low tonicity was obtained by adding sucrose and reducingNaCl concentration in the external medium,***:In intactmyocytes the frequency of Ca2+sparks was increased and decreased byhyperosmotic(1.5 T)and hyposmotic(0.6 T)exposure,*** addition,hyperosmotic exposure increased the temporal parameters and decreased the spa-tial parameter of Ca2+sparks,while opposite changes occurred with *** spatio-temporal properties of Ca2+sparks were slightly affected byaltering[K~+]i(50-200 mmol/L)in saponin permeabilized myocytes in the presenceof 8% *** was observed that the spatio-temporal parameters of the Ca2+sparks in permeabilized myocytes were dose-dependently altered by *** velocity of Ca2+waves in intact and permeabilized myocyte was alsoaffected by osmotic pressure or ***:The effect of osmotic stresson the frequency of spontaneous Ca2+sparks might be ascribed to the change ofmyoplasmic Ca2+and Ca2+content in the sarcoplasmic reticulum,while the effecton the spatio-temporal properties is caused by the alteration of Ca2+diffusionmainly resulting from the morphological change of the myocytes.

关键词： osmotic stress calcium sparks rat ventricular myocytes

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Transgenic mice overexpressingγ-aminobutyric acid transporter subtypeⅠ develop obesity

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Cell Research 2000年第4期10卷 303-310页

作者： MA YING HUA JIA HUA HU XIAO GANG ZHOU RUO WANG ZENG ZHEN TONG MEI JIAN FEI LI HE GUO Institute of Biochemistry and Cell Biology, shanghai institutes for biological {3., chinese academy of Science, shanghai 20003., china shanghai Institute of harmacy.ankara.edu.tr

Transgenic mice ubiquitously overexpressing murine γ aminobutyric acid transporter subtype Ⅰ were created. Unexpectedly, these mice markedly exhibited heritable obesity, which features significantly increased body weight and fat deposition. Behavioral examination revealed that transgeinc mice have slightly reduced spontaneous locomotive capacity and altered feeding pattern. Tills preliminary finding indicates that the inappropriate level of γ-aminobutyric acid transporters may be directly or indirectly involved in the pathogenic mechanism underlying certain types of obesity.

关键词： γ-aminobutyric acid transporter subtype Ⅰ transgenic mouse obesit€

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Conformational flexibility of β-secretase: molecular dynamics simulation and essential dynamics analysis

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Acta Pharmacologica Sinica 2004年第6期25卷 3-11页

作者： Bing XIONG, Xiao-qin HUANG, Ling-ling SHEN, Jian-hua SHEN 2,Xiao-min LUO, Xu SHEN, Hua-liang JIANG2, Kai-xian CHENDrug Discovery and Design Center, State Key Laboratory of Drug Research, shanghai Institute of Materia Medica,shanghai institutes for {3. sciences, chinese academy of sciences, shanghai 201203. china Drug Discovery and Design Center State Key Laboratory of Drug Research Shanghai Institute of Materia Medica Shanghai Institutes for Biological Sciences Chinese Academy of Sciences Shanghai 201203 China

AIM: Based on the structural analysis to reveal the mechanism of ligand binding to b-secretase and the specificityof each binding sub-site. METHODS: Molecular dynamics was used to simulate on the ligand free b-secretase andligand bound b-secretase. The trajectories were analyzed using the essential dynamics, and the significant confor-mational change was illustrated employing the DynDom program. RESULTS: The essential dynamics and DynDomanalyses clearly showed that the b-secretase experienced a large conformational change upon the substrate orinhibitor binding. The flap structure adopted a swing motion, gradually covering the active site to facilitate theligand binding process. Residues Ser86 and Ile87 served as the hinge point. Inhibitor-enzyme interaction analysisrevealed that residues at P2, P1, and P1 positions of the inhibitor were very important for the binding, and residuesat P2 and P3.positions may be modified to improve the binding specificity. S3.subsite of the enzyme still hadspace to modify the inhibitors in increasing the binding affinity. CONCLUSION: The information presented hereis valuable and could be used to identify small molecular inhibitors of b-secretase.

关键词： molecular dynamics secretase essential dynamics computer simulation Alzheimer disease

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