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Phorbol-induced surface expression of NR2A subunit homologues in HEK293 cells

Acta Pharmacologica Sinica 2006年第12期27卷 1580-1585页

作者： Chan-ying ZHENG~2 Xiu-juan YANG~3 Zhan-yan FU~4 Jian-hong LUO~(2,5)2 department of neurobiology,Zhejiang university School of medicine,Hangzhou 310031,China 3 Institute of Neuroscience,Shanghai Institutes for Biological Sciences,the Chinese Academy of Sciences,Shanghai 200031,China 4 department of Physiology and Biophysics,Georgetown university School of medicine Washington 20057,usa departmentofneurobiology ZhejiangUniversitySchoolofMedicineHangzhou310031China InstituteofNeuroscience ShanghaiInstitutesforBiologicalSciencestheChineseAcademyofSciencesShanghai200031China

Aim:N-methyl-D-aspartate receptors(NMDAR)are heteromeric complexes pri-marily assembled from NR1 and NR2 *** normal conditions,NR2 sub-units assemble into homodimers in the endoplasmic reticulum(ER).Thesehomodimers remain in the ER until they coassemble with NR1 dimers and aretrafficked to the cell ***,it still remains unclear whether functionalhomomeric NMDAR exist in physiological or pathological ***:We transfected GFP-NR2A alone into HEK293 cells,treated the cells with PKCactivator 12-myristate-13 acetate(PMA),and then detected surface NR2A sub-units with a live cell immunostaining *** also used a series of NR2Amutants with a partial deletion of its C-terminus to identify the regions that areinvolved in the PMA-mediated surface expression of NR2A ***:NR2A subunits were expressed on the cell membrane after incubation with PMA(200 nmol/L,30 min),although no functional NMDA channels were detected afterPMA-induced membrane *** with an ER marker alsorevealed that NR2A subunits were exported from the ER after PMA ***,the deletion of amino acids between 1149-1347 or 1354-1464 ofNR2A inhibited PMA-induced surface expression ofNR2A ***:First,our data suggests that PMA treatment can induce the surface expression ofhomomeric NR2A ***,this process is probably mediated bythe NR2A C-terminal region between positions 1149 and 1464.

关键词： NMDA receptor NR2A subunit PMA surface expression HEK293 cells

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Effects of enhancing p21^(waf1) on proliferation and expression of G1 cyclins and CDKs in human breast cancer cells

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Chinese Science Bulletin 1999年第16期44卷 1489-1493页

作者： SANG Jianli, WANG Yongchao and YE Yingjiang1. Laboratory of cell biology , department of biology , Beijing Normal university , Beijing 100875 , China 2. The Second Hospital, Beijing Medical university, Beijing 100044, China Laboratory of cell biology Department of Biology Beging Normal University Beijing China The Second Hospital Beijing Medical University Beijing China

The cyclin-dependent kinase inhibitor p21( waf1/cip1/sdi1) is an important negative regulator in control of cell cycle. Its functions of inhibiting cancer cell growth and its effects on expression of Gl phase cyclins and related CDKs are a worthy topic for study. The plasmid expressing p2l with high level was transformed to human breast cancer cells, and the expression of p2l in cells was enhanced, then the cell growth rate, anchorage-independent growth and tu-morigenecity were tested, at the same time the expression levels of cyclinD1, CDK4,

关键词： p21waf1 breast cancer cell cyclin CDK.

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Inducible resistance to Fas-mediated apoptosis in B cells

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cell Research 2000年第4期10卷 245-266页

作者： ROTHSTEIN THOMAS L departments of medicine and Microbiology, and the Evans Memorial department of Clinical Research, Boston university Medical Center, Boston, MA, usa 1departments of medicine and Microbiology and the Evans Memorial Department of Clinical Research Boston University Medical Center Boston MA USA

Apoptosis produced in B cells through Fas (APO-1, CD95) triggering is regulated by signals derived from other surface receptors: CD40 engagement produces upregulation of Fas expres. sion and marked susceptibility to Fas-induced cell death, whereas antigen receptor engagement, or IL-4R engagement, inhibits Fas killing and in so doing induces a state of Fas-resistance, even in otherwise sensitive, CD40-stimulated targets. Surfaceim. munoglobulin and IL-4R utilize at least partially distinct pathways to produce Fas-resistance that differentially depend on PKC and STAT6, respectively. Further, surface immunoglob- "lin signaling for inducible Fas-resistance bypasses Bib, requires NF-GB, and entails new macromolecular synthesis. Terminal effectors of B cell Fas-resistance include the known anti-apoptotic gene products, Bcl-xL and FLIP, and a novel anti-apoptotic gene that encodes FAIM (Fas Apoptosis Inhibitory Molecule). faim was identified by differential display and exists in two alternatively spliced forms; maim-S is broadly expressed, but faim-L expression is tissue-specific. The FAIM sequence is highly evolu tionarily conserved, suggesting an important role for this molecule throughout phylogeny. Inducible resistance to Fas killing is hypothesized to protect foreign antigen-specific B cells during potentially hazardous interactions with FasL-bearing T cells, whereas autoreactive B cells fail to become Fas-resistant and are deleted via Fas-dependent cytotoxicity. Inadvertent or aberrant acquisition of Fas-resistance may permit autoreactive B cells to escape Fas deletion, and malignant lymphocytes to impede anti-tumor immunity.

关键词： Apoptosis Fas B lymphocytes FAIM FLIP Bcl-XL surface immunoglobulin IL-4R CD40 autoreactivity

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Dicoumarol Alters cellular Redox State and Inhibits Nuclear Factor Kappa B to Enhance Arsenic Trioxide-Induced Apoptosis

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Acta Biochimica et Biophysica Sinica 2004年第3期36卷 235-242页

作者： Ya-Wu JING, Jing YI, Yu-Ying CHEN, Qing-Shen HU, Gui-Ying SHI, Hue LI, and Xue-Ming TANG* department of cell biology, Shanghai Second Medical university, Shanghai 200025, China department of cell biology Shanghai Second Medical University

The effects of a number of cytotoxic drugs are influenced by cellular reduction/oxidation (redox) state. In the present study, we attempt to explore if dicoumarol, an inhibitor of NADPH: quinone oxidoreductase (NQO1), alters the cellular redox state and how this alteration affects the redox-related apoptosis. Flow cytometry was used to assess the reactive oxygen species (ROS) level and apoptotic rates of HeLa cells treated with arsenic trioxide (As2O3) alone or in combination with natural anthraquinone emodin and dicoumarol or plus N-acetyl-cysteine. Western blot, immunofluorescence, electrophoretic mobility shift assay and luciferase assay were used to detect Nuclear Factor kappa B (NF-κB) activation. The results showed that dicoumarol synergized with emodin to sensitize HeLa cells to As2O3-induced apoptosis through raising the ROS level. More notably, this enhanced susceptibility was associated with a ROS-mediated inhi- bition of NF-κB activation in which the combinative treatment with dicoumarol prevented NF-κB from binding to target DNA. It was suggested that dicoumarol in combination with anthraquinones might be a novel strategy to expand the chemotherapeutic spectrum of As2O3 by means of interfering the cellular redox state.

关键词： dicoumarol arsenic trioxide reduction/ oxidation state apoptosis NF-κB

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microRNAs-powerful repression comes from small RNAs

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Science China(Life Sciences) 2009年第4期52卷 323-330页

作者： MA Cong, LIU YuFei & HE Lin 535 LSA, Division of cell and Developmental biology, MCB department, university of California at Berkeley, Berkeley, CA 94720-3200 535 LSA Division of Cell and Developmental Biology MCB Department University of California at Berkeley Berkeley CA 94720-3200 USA

microRNAs (miRNAs) encode a novel class of small, non-coding RNAs that regulate gene expression post-trancriptionally. miRNAs comprise one of the major non-coding RNA families, whose diverse bio- logical functions and unusual capacity for gene regulation have attracted enormous interests in the RNA world. Over the past 16 years, genetic, biochemical and computational approaches have greatly shaped the growth of the field, leading to the identification of thousands of miRNA genes in nearly all metazoans. The key molecular machinery for miRNA biogenesis and silencing has been identified, yet the precise biochemical and regulatory mechanisms still remain elusive. However, recent findings have shed new light on how miRNAs are generated and how they function to repress gene expression. miRNAs provide a paradigm for endogenous small RNAs that mediate gene silencing at a genome-wide level. The gene silencing mediated by these small RNAs constitutes a major component of gene regu- lation during various developmental and physiological processes. The accumulating knowledge about their biogenesis and gene silencing mechanism will add a new dimension to our understanding about the complex gene regulatory networks.

关键词： miRNA RNAi gene silencing mRNA degradation

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Bradykinin potentiates 5-HT3 receptor-mediated current in rat trigeminal ganglion neurons

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Acta Pharmacologica Sinica 2005年第4期26卷 428-434页

作者： Wang-ping HU~(1,2) Xue-mei LI~2 Ji-liang WU~3 Min ZHENG~3 Zhi-wang LI~4 2 department of Physiology,3 department of Pharmacology,Xianning college,Xianning 437100,China 4 department of neurobiology,Tongji Medical college of Huazhong university of Science and Technology,Wuhan 430030,China departmentofPhysiology XianningCollegeXianning437100China

Aim:To explore the modulatory effect of bradykinin （BK） on 5-HT3receptor-mediated current in trigeminal ganglion （TG） neurons in ***:The whole-cell patch-clamp technique was used to record 5-HT-activated currents (I5-HT) inneurons freshly dissociated from rat *** were applied by rapid ***:The majority of the neurons examined responded to 5-HTapplied externally with an inward current （76.3%,74/97） that could be blocked bythe 5-HT3receptor antagonist,ICS-205,930 (1×10-6mol/L).In 66 of the 74 cellssensitive to 5-HT （89.2%）,pretreatment for 30 s with BK (1×10-6-1×10-10mol/L)could potentiate I5-HTwith the maximal modulatory effect occurring at10-7mol/L BK （71.6%+4.9%）.BK shifted the 5-HT concentration-response curveupwards with an increase of 68.9%±7.2% in the maximal current response,but withno significant change in the EC50value （19.1±3.2 μmol/L vs 20.9±3.5 μmol/L;t-test,P>0.05;n=8）.BK potentiated I5-HTin a holding potential-independent manner anddid not alter the reverse potential of *** BK-induced potentiation ofI5-HTwas almost completely blocked by Hoe 140 (5×10-7mol/L),a selective B2BKreceptor antagonist,and was removed after intracellular dialysis of GF-109203X（2 μmol/L）,a selective protein kinase C （PKC） inhibitor,with the re-patch ***:Pre-application of BK exerts an enhancing effect on I5-HTvia a PKC-dependent pathway in rat TG neurons,which may explain the peripheral'mecha-nism of pain and hyperalgesia caused by,for example,tissue damage andinflammation.

关键词： bradykinin 5-HT3 serotonin receptor regulation patch-clamp techniques trigeminal ganglion

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Those amazing dinoflagellate chromosomes

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cell Research 2003年第4期13卷 215-217页

作者： PETER J RIZZO department of biology Texas A & M university college Station TX 77843 usa

Dinoflagellates are a very large and diverse group of eukaryotic algae that play a major role in aquatic food webs of both fresh water and marine habitats. Moreover, the toxic members of this group pose a health threat in the form of red tides. Finally, dinoflagellates are of great evolutionary importance, because of their taxonomic position, and their unusual chromosome structure and composition. While the cytoplasm of dinoflagellates is typically eukaryotic, the nucleus is unique when compared to the nucleus of other eukaryotes. More specifically, while the chromosomes of all other eukaryotes contain histones, dinoflagellate chromosomes lack histones completely. There are no known exceptions to this observation: all dinoflagellates lack histones, and all other eukaryotes contain histones. Nevertheless, dinoflagellates remain a relatively unstudied group of eukaryotes.

关键词： dinofiagellates chromosomes histones histone-like proteins.

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SH-2-containing protein tyro-sine phosphatase 1 is required for IL-4-induced IL-4R expression in spleen cells

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Science Bulletin 2005年第14期 50-55页

作者： CAO Xin1, HUANG Zan2 & FAN Jingyi3 1. department of cell biology and Medical Genetics, Nanjing Medical university, Nanjing 210029, China 2. Molecular biology Program, Stritch School of medicine, Loyola Uni- versity at Chicago, Maywood, IL 60153, usa 3. department of Organismal biology and anatomy, university of Chi- cago, Chicago, IL 60637, usa

To investigate the role of SH-2-containing pro- tein tyrosine phosphatase 1, SHP-1, in IL-4-induced IL-4 receptor (IL-4R) expression, we examined IL-4 receptor α-chain (IL-4Rα) mRNA expression in Na3VO4-treated wild type (WT) spleen cells and measured IL-4R mRNA in IL-4-stimulated spleen cells of viable motheaten mice (mev/mev). It is found that IL-4-induced IL-4R mRNA ex- pression was impaired in Na3VO4-treated WT spleen cells and IL-4-stimulated mev/mev spleen cells. Here we show that the impaired IL-4-induced IL-4Rα mRNA expression was due to reduced expression of IL-4R that led to impaired STAT6 signaling. We further demonstrate that reduction of IL-4Rα protein expression in mev/mev spleen cells was due to alteration in cell compositions. In mev/mev spleen, the per- centages of CD4+, CD8+, and CD19+ cells expressing rela- tively high levels of IL-4R were reduced dramatically while the percentages of Mac-1+ and Gr-1+ cells with relative low levels of IL-4R increased greatly. Despite the profound effect of reduced expression of IL-4R protein, the IL-4Rα mRNA expression was comparable in spleen cells of littermate con- trol mice (+/?) and mev/mev mice and no differences were found in B cells, T cells, and macrophages, suggesting cell type-specific downregulation of IL-4R expression in macro- phages through a posttranscriptional mechanism. Our study suggests that SHP-1 is required for IL-4-meidated function and indirectly regulates IL-4-meidated function in spleen cells by affecting hematopoiesis.

关键词： SHP-1 IL-4 receptor STAT6 signaling viable mot- heaten.

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Tetrandrine inhibits Ca2+-activated chloride channel in cultured human umbilical vein endothelial cells

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Acta Pharmacologica Sinica 2004年第03期25卷 73-79页

作者： Qi-zhi FANG1,2,Ning ZHONG1,3,Yi ZHANG1,Zhao-nian ZHOU1,4 1Physiological Lab of Hypoxia,Shanghai Institute of BiologicalSciences,Chinese Academy ofSciences,Shanghai200031,China 3department of Molecular and cell biology,university of California at Berkeley,Berkeley,CA 94720,usa 2 Now in CardiovascularResearch Institute,university of Cali- fornia at San Francisco,San Francisco,CA 94143,usa Physiological Lab of Hypoxia Shanghai Institute of Biological Sciences Chinese Academy of Sciences Shanghai 200031 China department of Molecular and cell biology University of California at Berkeley

AIM: To characterize the electrophysiological and kinetic properties of Ca2+-activated chloride channel (CaCC) in cultured human umbilical vein endothelial cell line (HUVEC), and test the inhibitory effects of tetrandrine (Tet) on CaCC. METHODS: Ca2+-activated Cl- currents (ICl,Ca) were recorded by patch-clamp whole cell configurations. [Ca2+]i was measured via intracellular Fura-2 fluorescence intensities. RESULTS: ICl,Ca was activated by increasing [Ca2+]i via direct elevation of intracellular calcium. ICl,Ca showed an apparent outward rectification properties, and it was activated in a voltage- and calcium-dependent mode. Tet dose-dependently inhibited ICl,Ca, the IC50 was (5.2±0.4) μmol/L (n=8 cells). Tet suppressed both voltage-dependent and calcium-dependent activation of ICl,Ca. The activa- tion time constant was (326±12) ms [in the presence of 10 μmol/L Tet, compared to control (175±17) ms, at +100 mV], and Ca2+ concentration for half maximal activation was (387±61) nmol/L for Tet (compared to control (287±36) nmol/L. CONCLUSIONS: Tet effectively blocked ICl,Ca, and such effects might be due to its inhibitory effects on the activation process of Ca2+-activated chloride channel.

关键词： chloride channels endothelium tetrandrine patch-clamp techniques calcium

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A novel model-free approach for reconstruction of time-delayed gene regulatory networks

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中国科学：生命科学英文版 2006年第2期49卷 190-200页

作者： JIANG Wei1,2, LI Xia1,2,3,4, GUO Zheng1,2,3, LI Chuanxing1, WANG Lihong1 & RAO Shaoqi1,5 1. department of Bioinformatics, Harbin Medical university, Harbin 150086, China 2. department of Computer Science, Harbin Institute of Technology, Harbin 150080, China 3. School of Life Science and Technology, Tongji university, Shanghai 200092, China 4. Biomedical Engineering Institute, Capital university of Medical Sciences, Beijing 100054, China 5. departments of Cardiovascular medicine and Molecular Cardiology, Cleveland Clinic Foundation, Cleveland, ohio 44195, usa department of Bioinformatics Harbin Medical University Harbin 150086 China department of Computer Science Harbin Institute of Technology Harbin 150080 China School of Life Science Technology Tongji University Shanghai 200092 China Biomedical Engineering Institute Capital University of Medical Sciences Beijing 100054 China departments of Cardiovascular medicine Molecular Cardiology Cleveland Clinic Foundation Cleveland Ohio 44195 USA

Reconstruction of genetic networks is one of the key scientific challenges in functional genomics. This paper describes a novel approach for addressing the regulatory dependencies be-tween genes whose activities can be delayed by multiple units of time. The aim of the proposed ap-proach termed TdGRN (time-delayed gene regulatory networking) is to reversely engineer the dy-namic mechanisms of gene regulations, which is realized by identifying the time-delayed gene regu-lations through supervised decision-tree analysis of the newly designed time-delayed gene expres-sion matrix, derived from the original time-series microarray data. A permutation technique is used to determine the statistical classification threshold of a tree, from which a gene regulatory rule(s) is ex-tracted. The proposed TdGRN is a model-free approach that attempts to learn the underlying regula-tory rules without relying on any model assumptions. Compared with model-based approaches, it has several significant advantages: it requires neither any arbitrary threshold for discretization of gene transcriptional values nor the definition of the number of regulators (k). We have applied this novel method to the publicly available data for budding yeast cell cycling. The numerical results demonstrate that most of the identified time-delayed gene regulations have current biological knowledge supports.

关键词： gene regulatory networks, time delayed, decision tree.

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