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A Viral Protein Suppresses siRNA-directed Interference in Tobacco Mosaic Virus Infection

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Acta Biochimica et Biophysica Sinica 2005年第4期37卷 248-253页

作者： Ming-Min ZHAO De-Rong AN Guang-Hua HUANG~1 Zu-Hua HE~2 Jiang-Ye CHEN~1 College of Plant Protection,North West Science and Technology University of Agriculture and Forestry,Shaanxi 712100,china ~1State key laboratory of Molecular {3.,Institute of Biochemistry and Cell {3.,shanghai institutes for biological sciences. chinese Academy of sciences,shanghai 20003.,china ~2State key laboratory of Plant Molecular Genetics.Institute of Plant Physiology and Ecology,shanghai institutes for biological sciences.chinese Academy of sciences,shanghai 20003..china CollegeofPlantProtection NorthWestScienceandTechnologyUniversityofAgricultureandForestryShaanxi712100China

Plant viruses encode suppressors of post-transcriptional gene silencing (PTGS), an adaptivedefense response that limits virus replication and its spread in *** helper component proteinase (HC-Pro) of the potato virus A (PVA,genus Potyvirus) suppresses PTGS of silenced ***,the effectof HC-Pro on siRNA-directed interference in the tobacco mosaic virus (TMV) was examined by using atransient Agrobacterium tumefaciens-based delivery system in intact *** was shown that the interferenceeffect was completely blocked by co-infiltration with HC-Pro plus siRNA constructs in both systemic andhypersensitive *** the system host,all plants agro-infiltrated with HC-Pro plus siRNA constructsdisplayed the same symptoms as the negative ***,TMV RNA accumulation was found to beabundant in the upper leaves using reverse transcriptase-PCR (RT-PCR) and Northern blot *** thecontrary,plants agro-infiltrated with the siRNA construct alone were free of ***,ourstudy suggests that the transient expression of HC-Pro inhibited the siRNA-directed host defenses againstTMV infection.

关键词： siRNA HC-Pro tobacco mosaic virus suppressor

pH-Dependent Stability of EGX, A Multi-functional Cellulase from Mollusca, Ampullaria crossean

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Acta Biochimica et Biophysica Sinica 2004年第9期36卷 603-608页

作者： Wen-Ying LI2, Ji WANG1, Yan-Hong LI1, Ming DING1, Gen-Jun XU1,3. Lan-Ying LIU2, and Fu-Kun ZHAO1,3. 1key laboratory of Proteomics, Institute of Biochemistry and Cell biology, {3., the chinese Academy of sciences,Graduate School of the chinese Academy of sciences, shanghai 20003., china 2College of Life Science, Jilin University, Changchun 13.023. china College of Life Science, Zhejiang University of Science and Technology, Hangzhou 3.0018, china key laboratory ofProteomics Institute of Biochemistry and Cell BiologyShanghai Institutes for Biological Sciencesthe Chinese Academy of SciencesGraduate School of the Chinese Academy of SciencesShanghai 200031China

The cellulase activity and stability of EGX, a multi-functional cellulase previously purified from the mollusca Ampullaria crossean, was systematically studied under different pH. The pH induced con- formation and stability change of EGX have been investigated by using the intrinsic fluorescence, ANS fluorescence and CD spectrum. It has been found that the conformation and activity of this cellulase were strongly dependent on the pH. EGX was stable for both the enzyme activity and the conformation from pH 5.6 to pH 7.4. As shown by intrinsic and ANS fluorescence, no red shift of emission maximum occurred and a negligible intensity change was observed at pH 5.6–7.4. The activity of EGX remained about 80% in pH 5.6–7.4 and obviously decreased out of side the pH range. Urea-induced changes in EGX at pH 5.4 and pH 8.0 were measured by intrinsic fluorescence and CD spectrum. At pH 5.4, a significantly red shift of emission maximum occurred when the concentration of urea was 5 M compared to the concentration was 3.M at pH 8.0. The α-helix at pH 5.4 was 40.51% in the absence of urea and 3..04% in the presence of 4 M urea. At pH 8.0 the α-helix was 7.23. in the presence of 4 M urea. The data indicated that EGX was much susceptible to urea-induced unfolding at pH 8.0 and much stable at pH 5.4. The greater pH dependent stability of EGX may allow the enzyme to adequately catalyze the hydrolysis of cellulosic materials under natural or industrial extreme conditions.

关键词： multi-functional cellulase stability Ampullaria crossean fluorescence spectroscopy pH-dependent structural change CD spectrum

C/EBPαp3. plays transcriptional regulatory roles distinct from C/EBPαp42

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Cell Research 2007年第4期17卷 374-383页

作者： Chunxi Wang~1 Xiaotao Chen~1 Yanping Wang~1 Jialei Gong~1 Gengxi Hu ~1State key laboratory of Molecular biology institute of Biochemistry and Cell biology,shanghai institutes for biological sciences,chinese Academy of sciences,Graduate School of the chinese Academy of sciences,shanghai 20003.,china StatekeylaboratoryofMolecularbiology InstituteofBiochemistryandCellBiologyShanghaiInstitutesforBiologicalSciencesChineseAcademyofSciencesGraduateSchooloftheChineseAcademyofSciencesShanghai200031China

CCAAT/enhancer binding protein α(C/EBPα)is a transcriptional regulatory factor that inhibits cell proliferation,andalternative translational initiation produces two polypeptides,C/EBPcαp3. and C/EBPα*** expression profiling,it was revealed that C/EBPαp3. specifically inhibited a unique set of genes,including MPP11,p84N5 and SMYD2,which were not affected by C/EBPαp42 in both QSG-7701 hepatocyte cell line and QGY-7703.hepatoma ***-quantitative RT-PCR analysis independently confirmed these *** immunoprecipitation assay showed thatC/EBPαp3. bound to the promoters of these genes more strongly than C/EBPα*** clinical hepatoma samples inwhich C/EBPα was downregulated,all three genes specifically inhibited by C/EBPαp3. were ***,repression of MPP11,p84N5 and SMYD2 genes might not be directly involved in C/EBPαp3.-mediated growth *** data suggest that C/EBPαp3. regulates a unique set of target genes and is more than a dominant-negativeregulator of C/EBPαp42.

关键词： C/EBPα expression pattern p84N5 MPP11 SMYD2

一种重组抗胰蛋白酶单体人胰岛素突变体的设计、制备和鉴定

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生物化学与生物物理学报 2000年第6期32卷 633-636页

作者：张弘陈衍冯佑民中国科学院上海生命科学研究院上海生物化学研究所分子生物学国家重点实验室!上海20003.

用缺口双链DNA的定向突变方法分别将胰岛素前体中B链第 2 2、2 8、2 9和 3.0位改变为Asp、Lys、Pro和Lys,酵母分泌表达的前体经胰蛋白酶直接酶切 ,得到重组 [B2 2Asp、B2 8Lys、B2 9Pro、B3.0Lys]人胰岛素。它与受体的结合能力约为猪胰... 详细信息

用缺口双链DNA的定向突变方法分别将胰岛素前体中B链第 2 2、2 8、2 9和 3.0位改变为Asp、Lys、Pro和Lys,酵母分泌表达的前体经胰蛋白酶直接酶切 ,得到重组 [B2 2Asp、B2 8Lys、B2 9Pro、B3.0Lys]人胰岛素。它与受体的结合能力约为猪胰岛素的 6% ,而体内生物活力保留 5 0 %。通过FPLC分子筛测定其自身结合能力 ,在生理条件下浓度达 10 -4mol/L时它以单体形式存在。作为可抗胰蛋白酶酶解的单体胰岛素类似物 ,它可能具有一定的应用前景

关键词：胰岛素突变体单体胰岛素蛋白质工程

Inhibition of genes expression of SARS coronavirus by synthetic small interfering RNAs

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Cell Research 2005年第3期15卷 193-200页

作者： Yi SHI,De Hua YANG,Jie XIONG,Jie JIA,Bing HUANG,You Xin JIN* State key laboratory of Molecular biology,Institute of Biochemistry and Cell biology,shanghai institutes for biological sciences,chinese Academy of sciences,shanghai 20003.,china StatekeylaboratoryofMolecularbiology InstituteofBiochemistryandCellBiologyShanghaiInstitutesforBiologicalSciencesChineseAcademyofSciencesShanghai200031China

RNA interference (RNAi) is triggered by the presence of a double-stranded RNA (dsRNA), and results in the silencing of homologous gene expression through the specific degradation of an mRNA containing the same sequence. dsRNA- mediated RNAi can be used in a wide variety of eucaryotes to induce the sequence-specific inhibition of gene expression. Synthetic 21-23.nucleotide (nt) small interfering RNA (siRNA) with 2 nt 3. overhangs was recently found to mediate efficient sequence-specific mRNA degradation in mammalian cells. Here, we studied the effects of synthetic siRNA duplexes targeted to SARS coronavirus structural proteins E, M, and N in a cell culture system. Among total 26 siRNA duplexes, we obtained 3.siRNA duplexes which could sequence-specifically reduce target genes expression over 80% at the concentration of 60 nM in Vero E6 cells. The downregulation effect was in correlation with the concentrations of the siRNA duplexes in a range of 0~60 nM. Our results also showed that many inactive siRNA duplexes may be brought to life simply by unpairing the 5’ end of the antisense strands. Results suggest that siRNA is capable of inhibiting SARS coronavirus genes expression and thus may be a new therapeutic strategy for treatment of SARS.

关键词： SARS small interfering RNA Vero E6 cells EGFP fusion protein antiviral therapy.

Structural Evidence for α-Synuclein Fibrils Using in Situ Atomic Force Microscopy

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Acta Biochimica et Biophysica Sinica 2005年第2期37卷 113-118页

作者： Feng ZHANG Li-Na JI Lin TANG Jun HU Hong-Yu HU Hong-Jie XU Jian-Hua HE shanghai Institute of Applied Physics,chinese Academy of sciences.shanghai 201800.china key laboratory of Proteomics,Institute of Biochemistry and Cell biology.shanghai institutes for biological sciences,chinese Academy of sciences.shanghai 20003..china Bio-X Research Center.shanghai Jiaotong University.shanghai 20003..china shanghai Institute of Applied Physics Chinese Academy of Sciences Shanghai 201800 China

Human α-synuclein is a presynaptic terminal protein and can form insoluble fibrils that arebelieved to play an important role in the pathogenesis of several neurodegenerative diseases such as Parkinson'sdisease,demen... 详细信息

关键词： results provide indirect evidence on how β-sheets assemble into α-synuclein fibrils on a nanometer scale.α-synuclein atomic force microscopy tapping mode contact mode guanidine hydrochloride

Functional analysis of two Sp1/Sp3.binding sites in murine Nanog gene promoter

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Cell Research 2006年第3期16卷 319-322页

作者： Da Yong Wu Zhen Yao laboratory of Molecular Cell biology,laboratory of Stem Cell biology,Institute of Biochemistry and Cell biology,shanghai institutes for biological sciences,chinese Academy of sciences 3.0 Yue Yang Road,shanghai 20003.,china 1laboratory of Molecular Cell biology Laboratory of Stem Cell Biology Institute of Biochemistry and Cell Biology Shanghai Institutes for Biological Sciences Chinese Academy of Sciences 320 Yue Yang Road Shanghai 200031 China.

Nanog gene plays a key role in maintaining pluripotency of ES cells and early embryonic cells.A 5’ flank sequenceof the Nanog gene has been reported to be regulated differentially,and two regulatory elements within the Nanog pro-moter,namely Oct-4 and Sox-2 binding sites,have been identified to regulate the transcriptional activity ofNanog *** this report,we identified the role of two putative Sp 1 binding sites located in the Nanog gene 5’-flanking region inregulation of murine Nanog gene *** studies showed that the two sites were essential for the Nanogpromoter *** shift and supershift analysis showed that both sites specifically bind Spl and ***,overexpression of dominant-negative Sp 1 or Sp3.mutants significantly inhibits Nanog promoter *** resultssuggest that the transcription factor Spl and Sp3.are important for Murine Nanog gene expression.

关键词： Nanog promoter Sp1 Sp3

Developmental regulation of intracellular calcium transients during cardiomyocyte differentiation of mouse embryonic stem cells

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Acta Pharmacologica Sinica 2006年第7期27卷 901-910页

作者： Ji-dong FU Hui-mei YU Rong WANG Ji LIANG Huang-tian YANG~2 key laboratory of Stem Cell biology,Institute of Health {3.,shanghai institutes for biological sciences,chinese Academy of {3. and shanghai Jiao Tong University School of Medicine,shanghai 200025,china keylaboratoryofStemCellbiology InstituteofHealthSciencesShanghaiInstitutesforBiologicalSciencesChineseAcademyofSciencesandShanghaiJiaoTongUniversitySchoolofMedicineShanghai200025China

Aim:To investigate the developmental regulation of intracellular Ca2+transients,an essential event in excitation-contraction coupling,during ***:Using the embryonic stem(ES)cell in vitro differentia-tion system and pharmacological intervention,we investigated the molecular andfunctional regulation of Ca2+handling proteins on the Ca2+transients at early,intermediate and later differentiation stages of ES cell-derived cardiomyocytes(ESCM).Results:Nifedipine,a selective antagonist of L-type Ca2+channels,totally blocked Ca2+transients even in the condition of field-electric stimulation inESCM at three differentiation *** Ca2+transients of ESCM were alsoinhibited by both ryanodine[an inhibitor of ryanodine receptors(RyRs)]and 2-aminoethoxydipheylborate[2-APB,an inhibitor of inositol-1,4,5-trisphosphatereceptors(IP3.s)].The inhibitory effect of ryanodine increased with the time ofdifferentiation,while the effect of 2-APB decreased with the ***,an inhibitor of SR Ca2+-pump ATPase,inhibited Ca2+transientsequally at three differentiation stages that matched the expression ***~+free solution,which inhibits Na~+-Ca2+exchanger(NCX)to extrude Ca2+from cytosol,did not affect the amplitude of Ca2+transients of ESCM until the latter differentia-tion stage,but it significantly enhanced the basal Ca2+***:The Ca2+transients in ESCM depend on both the sarcolemmal Ca2+entry viaL-type Ca2+channels and the SR Ca2+release from RyRs and IP3.s even at theearly differentiation stage;but NCX seems not to regulate the peak of Ca2+tran-sients until the latter differentiation stage.

关键词： Ca2+ transients cardiac differentiation embryonic stem cells Ca2+ handing proteins

水稻齿叶矮缩病毒S8基因编码的结构蛋白质及其自聚集性质

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生物化学与生物物理学报 2002年第5期34卷 565-570页

作者：吕慧涓邵朝纲龚祖埙中国科学院上海生命科学研究院生物化学与细胞生物学研究所、蛋白质组学重点实验室中国科学院上海生命科学研究院生物化学与细胞生物学研究所、蛋白质组学重点实验室上海20003.

水稻齿叶矮缩病毒是一种以水稻为侵害寄主的植物呼肠孤病毒 ,其基因组 1 0条dsRNA编码的蛋白质产物的功能除S9外的大部分还未阐明。报道了该病毒菲律宾分离株S8的开放阅读框序列 ,在大肠杆菌中表达和纯化出了其 6 7kD的蛋白质产物 ,并... 详细信息

水稻齿叶矮缩病毒是一种以水稻为侵害寄主的植物呼肠孤病毒 ,其基因组 1 0条dsRNA编码的蛋白质产物的功能除S9外的大部分还未阐明。报道了该病毒菲律宾分离株S8的开放阅读框序列 ,在大肠杆菌中表达和纯化出了其 6 7kD的蛋白质产物 ,并进一步研究了该产物的功能。实验结果表明 ,S8的蛋白质产物是病毒的一种主要结构蛋白质 ,在体外具有自剪切活性和自聚集性质 ,并推测可能是病毒的内层衣壳蛋白

关键词： RRSV P8 结构蛋白质自剪切自聚集