跳到主要內容

臺灣博碩士論文加值系統

(44.220.247.152) 您好!臺灣時間:2024/09/18 23:07
字體大小: 字級放大   字級縮小   預設字形  
回查詢結果 :::

詳目顯示

我願授權國圖
: 
twitterline
研究生:程雅玲
研究生(外文):Chen Ya-Ling
論文名稱:EB病毒蛋白質激脢BGLF4轉錄、轉譯起點及其可能之Ganciclovir激脢活性之探討
論文名稱(外文):Characterization of the Transcription and Translation Initiation Sites of Epstein-Barr Virus BGLF4 Protein Kinase and It's Possible Ganciclovir Kinase Activity
指導教授:陳美如陳美如引用關係
指導教授(外文):Mei Ru ,Chen
學位類別:碩士
校院名稱:國立臺灣大學
系所名稱:微生物學研究所
學門:生命科學學門
學類:微生物學類
論文種類:學術論文
論文出版年:2001
畢業學年度:89
語文別:中文
論文頁數:59
中文關鍵詞:蛋白質激脢轉錄轉譯磷酸化作用ganciclovirEA-D
外文關鍵詞:protein kinasetranscriptiontranslationphosphorylationganciclovirEA-D
相關次數:
  • 被引用被引用:1
  • 點閱點閱:160
  • 評分評分:
  • 下載下載:0
  • 收藏至我的研究室書目清單書目收藏:0
EB病毒開放譯讀架BGLF4,經胺基酸序列比對發現具有ser/thr蛋白質激?功能區保守的胺基酸。在過去的研究中,利用免疫沉澱及蛋白質激?實驗,已證實BGLF4能自我磷酸化,並且能夠磷酸化casein、histone及EB病毒的early-antigen diffuse type (EA-D)。根據EBV基因體圖譜分析中所標示的BGLF4,是位於B95-8病毒的123,692-122,328,由於在123,692並無ATG之轉譯起始密碼,因此在過去的研究中,我們選擇其下游第一個in frame ATG的開放譯讀架進行擇殖及分析其蛋白質激?活性。在本論文中,為瞭解BGLF4基因表現的調控,我們首先利用5'-RACE定位BGLF4轉錄啟始點,經由和B95-8細胞株的EB病毒序列比對的結果,分別在BGLF4開放譯讀架上游255及201 bp處,定位出BGLF4 mRNA 轉錄起始點。由於BGLF4具有長達250 bp的5'端未轉譯區,因此在本論文中也對此段未轉譯區的功能及可能的non-ATG轉譯起始點作探討。另一方面,我們也利用之前實驗室所建立的系統及共轉染實驗,探討ATG上游序列對BGLF4蛋白質激?活性的影響。從蛋白質激?活性實驗看來,包含ATG上游序列的BGLF4的確具有較強的磷酸化訊號。此外,由於BGLF4之類似基因產物HCMV UL97、HHV-8 ORF36已被證實具有ganciclovir激?活性。故在本實驗中利用短暫轉染系統,測試表現BGLF4細胞對於ganciclovir的感受性,結果顯示表現BGLF4細胞相較於HSV-TK對ganciclovir有較弱的感受性。

The Epstein-Barr virus (EBV) open reading frame BGLF4 was identified as a ser/thr protein kinase based on the homology alignment to the conserved motifs of known protein kinases. In a previous study, by using an EBNA-1 tag, the immunoprecipitated BGLF4 was demonstrated for its abilities of autophosphorylation and phosphorylating casein, histone, and EBV early-antigen diffuse type (EA-D). According to the EBV genome map, BGLF4 is located at nt 123,692-122,328 of B95-8 EBV. Since no in frame ATG was identified at 123,692, the first in frame ATG at 123,614 was used to express the BGLF4 protein in our previous studies. One major goal of this study was to characterize the transcription and translation initiation sites of BGLF4. Therefore, 5'-RACE was used to map the 5' end of BGLF4 containing transcripts. Two populations of cDNA clones containing -201 and -255 upstream of the first in frame ATG were identified. Since both transcription initiation sites are relatively far away from the first in frame ATG, the possibilities of the 5'-end region can function as an internal ribosome entry site (IRES) and facilitate a non-ATG translation initiation were also investigated in this study. The possible contributions of the ATG-upstream sequence on BGLF4 protein kinase activity were examined in immuno-complex (IP) autophosphorylation assay and the transphosphorylation effect on EBV early antigen (EA-D) in co-transfected cells. Result of these experiments revealed that an-EBNA-1 tagged BGLF4 protein containing the -78 upstream region demonstrated stronger phosphorylation signals in IP-kinase assay. Finally, this study also demonstrated that, in comparing with HSV-TK, BGLF4 confers relative lower ganciclovir sensitivity in transient transfected 293T cells .

中文摘要----------------------------------------------------------I
英文摘要----------------------------------------------------------II
緒論----------------------------------------------------------------1
材料與方法-------------------------------------------------------11
實驗結果----------------------------------------------------------22
討論----------------------------------------------------------------29
圖表----------------------------------------------------------------35
參考文獻----------------------------------------------------------50

Baer, R., Bankier, A. J., Biggin, M. D., Deininger, P. L., Farrell, P. J., Gibson, T. J., Hartfull, G., Hudson, G. S., Statchwell, S. C., Seguin, C., Tuffnell, P. S., and Barrell, B. G. (1984). DNA sequence ad expression of the B95-8 Epstein-Barr virus genome. Nature (London) 310, 207-211.
Boeck, R., Curran, J., Matsuoka, Y., Compans, R., and Kolakofsky, D. (1992). The parainfluenza virus type 1 P/C gene uses a very efficient GUG codon to start its C' protein. Journal of Virology 66, 1765-1768.
Burkitt, D. (1958). A sarcoma involving the jaw in African children. British Journal of Surgery 46, 218-223.
Cannon, J. S., Hamzeh, F., Moore, S., Nicholas, J., and Ambinder, R. F. (1999). Human herpesvirus 8-encoded thymidine kinase and phosphotransferase homologues confer sensitivity to ganciclovir. Journal of Virology 73, 4786-4793.
Chavier, P., Gruffat, H., Chevallier, G. A., Buisson, M., and Sergeant, A. (1989). The Epstein-Barr virus (EBV) early promoter DR contains a cis-acting element responsive to the EBV transactivator EB1 and an enhancer with constitutive and inducible activities. Journal of Virology 63, 607-614.
Chee, M., Lawrence, G. L., and Barrell, B. G. (1989). Alpha-, beta- and gammmaherpesviruses encode a putative phosphotransferase. Journal of General Virology 70, 1151-1160.
Chen, C., and Okayama, H. (1987). High efficiency transformation of mammalian cells by plasmid DNA. Molecular Cell Biology 7, 2745-2752.
.
Chen, M. R., Hsu, T. Y., Chen, J. Y. and Yang, C. S. (1990). Molecular characterization of a cDNA clone encoding the Epstein-Barr virus (EBV) DNase. Journal of Virological Methods 29, 127-142.
Chen, M. R., Tsai, C. H., Wu, F. F., Kan, S. H., Yang, C. S., and Chen, J. Y. (1999). The major immunogenic epitopes of Epetein-Barr virus (EBV) nuclear antigen 1 are encoded by sequence domains which vary among nasopharyngeal carcinoma biopsies and EBV-associated cell lines. Journal of General Virology 80, 447-455.
Chen, M. R., Chang, S. J., Huang, H. and Chen, J. Y. (2000). A protein kinase activity associated with Epstein-Barr virus BGLF4 phosphorylates the viral early antigen EA-D in vitro. Journal of Virology 74, 3093-3104.
Cho, M. S., Milman, G., and Hayward, S. D., (1985a). Localization of the coding region for an Epstein-Barr virus early antigen and inducible expression of this 60-kilodalton nuclear protein in transfected fibroblast cell lines. Journal of Virology 56, 852-859.
Cho, M. S., Milman, G., and Hayward, S. D., (1985b). A second Epstein-Barr virus early antigen gene in BamHI fragment M encodes a 48- to 50-kilodalton nuclear protein. Journal of Virology 56, 860-866.
Curran, J., and Kolakofsky, D. (1988). Ribosomal initiation from an ACG codon in the Sendai virus P/C mRNA. EMBO J. 7, 245-251.
Daikoku, T., Shibata, S., Goshima, F., Oshima, S., Tsurumi, T., Yamada, H., Yamashita, Y., and Nishiyama, Y. (1997). Purfication and characterization of the protein kinase encoded by the UL13 gene of herpes simplex virus type 2. Virology 235, 82-93.
De Wind, N., Doman, J., and Berns, A., (1992). Herpesviruses encode an unusual protein-serine/threonine kinase which is nonessential for growth in cultured cells. Journal of Virology 66, 5200-5209.
Epstein, M. A., Achong, B., and Barr, Y. M. (1964). Virus particle in cultured lymphoblasts from Burkitt lymphoma. Lancet 1, 702-703.
Epstein, M. A. (1984). Immunobiochemical characterization with monoclonal antibodies of Epstein-Barr virus-associated early antigens in chemical induced cells. Journal of Virology 50, 372-379.
Farrell, P. J. (1992). Epstein-Barr virus genome. Advances in Viral Oncology 8, 103-132.
Feighny, R. J., Farrell, M. P., and Pagano, J. S. (1980). Polypeptide synthesis and phosphorylation in Epstein-Barr virus-infected cells. Journal of Virology 34, 455-463.
Gruffat, D. E., Holley, G. E., Zhang, Q., Stein, B., Blanar, M. A., Baldwin, A. S., and Kenney, S. C. (1994). The bZIP transactivator of Epstein-Barr virus, BZLF1, functional and physically interacts with the p65 subunit of NF-κB. Molecular Cell Biology 14, 1939-1948.
Gustafson, E. A., Chillemi, A. C., Sage, D. R., and Fingeroth, J. D. (1998). The Epstein-Barr virus thymidine kinase does not phosphorylate ganciclovir or acyclovir and demonstrates a narrow substrate specificity compared to the herpes simplex virus type 1 thymidine kinase. Antimicrobial Agents and Chemotherapy 42, 2923-2931.
Hann, S. R., King, M. W., Bently, D. L., Anderson, C. W., and Eisenman, R. N. (1988). A non-ATG translation initiation in c-myc exon 1 generates an N-terminally distinct protein whose synthesis is disrupted in Burkitt's lymphomas. Cell 52, 185-195.
Hardwick, J. M., Liberman, P. M., and Hayward, S. D. (1988). A new Epstein-Barr virus transactivator, R, induces expression of a cytoplasmic early antigen. Journal of Virology 62, 2274-2284.
Henle, G., and Henle, W. (1966). Immunofluorescence in cells derived from Burkitt's lymphoma. Journal of Bacteriology 91, 1248-1256.
Henle, G., Henle, W., and Diehl, V. (1968). Relation of Burkitt's tumor-associated herpes-type virus to infectious mononucleosis. Proceedings of the National Academy of Sciences USA 59, 94-111.
Henle, G., Henle, W., and Klein, G. (1971). Demonstration of two distinct components in the early antigen complex of Epstein-Barr virus-infected cells. International Journal of Cancer 8, 272-282.
Kallin, B., Sternas, L., Saemundssen, A. K., Luka, J., Jurnvall, H., Eriksson, B., Tao, P. Z., Nilsson, M. T., Klein, G. (1985). Purification of Epstein-Barr virus DNA polymerase from P3HR-1 cells. Journal of Virology 54, 561-568.
Kato, K., Kawaguchi, Y., Tanaka, M., Igarashi, M., Yokoyama, A., Matsuda, G., Kanamori, M., Nakajima, K., Nishimura, Y., Shimojima, M., Phung, H. T. T., Takahashi, E., Hirai, K. (2001). Epstein-Barr virus-encoded protein kinase BGLF4 mediates hyperphosphorylation of cellular elongation factor 1δ (EF-1δ):EF-1δ is universally modified by conserved protein kinases of herpesviruses in mammalian cells. Journal of General Virology 82, 1457-1463.
Kawaguchi, Y., Sant, C. V., and Roizman, B. (1998). Eukaryotic elongation factor 1δis hyperphosphorylated by the protein kinase encodes by the UL13 gene of herpes simplex virus 1. Journal of Virology 72, 1731-1736.
Kieff, E. (1996). Epstein-Barr virus and its replication. In ''Field B. N. and Knipes D. M. (EDS.)'' Virology. Raven Press, Ltd., New York, 2343-2396.
Kikuta, H., Taguchi, Y., Tomizawa, K., Kawamura, N., Ishizaka, A., Sakiyama, Y., Matsumoto, S., Imai, S., and Kinoshita, T. (1988). Epstein-Barr virus genome-positive T lymphocytes in a boy with chronic active EBV infection associated with Kawasaki-like disease. Nature 333, 455-457.
Klein, G., Clifford, P., Klein, E., and Stjernsward, J. (1966). Search for tumor-specific immune reactions in Burkitt lymphoma patients by the membrane immunofluorescence reaction. Proceedings of the National Academy of Sciences U.S.A. 55, 1628-1635.
Leader, D. P. (1993). Viral protein kinases and protein phosphatases. Pharmacology and Therapeutics 59, 343-389.
Levy J. A., and Henle G. (1966). Indirect immunofluorescence tests with sera from African children and cultured Burkitt lymphoma cells. Journal of Bacteriology 92, 275-276.
Li, J. S., Zhou, B. S., Dutschman, G. E., Grill, S. P., Tan, R. S., and Cheng, Y. C. (1987). Association of Epstein-Barr virus early antigen diffuse component and virus-specified DNA polymerase activity. Journal of Virology 61, 2947-2949.
Lieberman, P. M., and Berk, A. J. (1990). In vitro transcriptional activation, dimerization, and DNA-binding specificity of the Epstein-Barr virus Zta protein. Journal of Virology 64, 2560-2568.
Lieberman, P. M., and Berk, A. J. (1991). The Zta transactivator protein stabilizes TFIID association with promoter DNA by direct protein-protein interaction. Genes and Development 5, 2441-2454.
Littler, E., Stuart, A. D. and Chee, M. S. (1992). Human cytomegalovirus UL97 open reading frame encodes a protein that phosphorylates the antiviral nucleoside analogue ganciclovir. Nature 358, 160-162.
Long, M. C., Leong, V., Schaffer, P. A., Spencer, C. A., and Rice, S. A. (1999). ICP22 and the UL13 protein kinase are both required for the herpes simplex virus-induced modification of the large subunit of RNA polymerase II. Journal of Virology 73, 5593-5604.
Longnecker, R. and Roizman, B. (1987). Clustering of genes dispensable for growth in culture in the S component of the HSV-1 genome. Science 236, 573-576.
McGeoch, D. J., Dolan, A., Donald, S. and Rixon, F. J. (1985).Sequence determination and genetic content of the short unique region in the genome of herpes simplex virus type 1. Journal of Molecular Biology 181, 1-13.
Meignier, B., Longnecker, R., Mavromara-Nazos, P., Sears, A. E., and Roizman, B. (1988). Virulence and establishment of latency by genetically engineered deletion mutants of herpes simplex virus 1. Virology 162, 251-254.
Oguro, M. O., Shimizu, N., Ono, Y., and Takada, K. (1987) Both the rightward and the leftward open reading frames within the BamHI M DNA fragment of Epstein-Barr virus act as trans-activators of gene expression. Journal of Virology 61, 3310-3313.
Overton, H. A., Mcmillan, D. J., Klavinskis, L. S., Hope, L., Ritchie, A. J., and Wong-kai-in, P. (1992). Herpes simplx virus type I gene UL13 encodes a phosphoprotein that is a component of the virion. Virology 190, 184-192.
Pai, S., Ponta, H., Rahmsdorf, H., Hirsch, M., Herrlich, P., and Schweiger, M. (1975a). Protein kinase of bacteriophage T7. 1. Purfication. European Journal of Biochemistry 55, 299-304.
Pai, S., Ponta, H., Rahmsdorf, H., Hirsch, M., Herrlich, P., and Schweiger, M. (1975b). Protein kinase of bacteriophage T7. 2. Proterties, enzyme synthesis in vitro and regulation of enzyme synthesis and activity in vivo. European Journal of Biochemistry 55, 305-314.
Prats, H., Kaghard, M., Prats, M., Klagsburn, M., Lelias, J. M., Liauzun, P., Chalon, P., Tauber, J. P., Amalric, F., Smith, J. A., and Caput, D. (1989). High molecular mass forms of basic fibroblast growth factor are initiated by alternative CUG codons. Proceedings of the National Academy of Sciences USA 86, 1836-1840.
Prichard, M. N., Gao, N., Jairath, S., Mulamba, G., Krosky, P., Coen, D. M., Parker, B. O., and Pari, G. S. (1999). A recombinant human cytomegalovirus with a large deletion in UL97 has a severe replication deficiency. Journal of Virology 73, 5663-5670.
Purves, F. C., Katan, M., Stevely, W. S., and Leader, D.P. (1986). Characteristics of the induction of a new protein kinase in cells infected with herpesviruses. Journal of General Virology 67, 1049-57.
Purves, F. C., and Roizman, B. (1992). The UL13 gene of herpes simplex virus 1 encodes the functions for posttranslational processing associated with phosphorylation of the regulatory protein α22. Proceedings of the National Academy of Sciences USA 89, 7310-7314.
Purves, F. C., Ogle, W. O., and Roizman, B. (1993). Processing of the herpes simplex virus regulatory protein α22 mediated by the UL13 protein kinase determines the accumulation of a subset of α and γmRNAs and proteins in infected cells. Proceedings of the National of Sciences USA 90, 6701-6705.
Rahmsdorf, H. J., Pai, S. H., Ponta, H., Herrlich, P., Roskoski, R., Schweiger, M., and Studier, F. W. (1974). Protein kinase induction in Escherichia coli by bacteriophage T7. Proceedings of the National Academy of Sciences USA 71, 586-589.
Rice, S. A., Long, M. C., Lam, V., Schaffer, P. A., and Spencer, C. A. (1995). Herpes simplex virus immediate-early protein ICP22 is required for viral modification of host RNA polymerase II and establishment of the normal viral transcription program. Journal of Virology 69, 5550-5559.
Sasaki, J., and Nakashima, N., (1999). Translation initiation at the CUU codon is mediated by the internal ribosome entry site of an insect picorna-like virus in vitro. Journal of Virology 73, 1219-1226.
Schmitz, J., Prufer, D., Rohde, W., and Tacke, E. (1996). Non-canonical translation mechanisms in plants: efficient in vitro and in planta initiation at AUU codons of the tobacco mosaic virus enhancer sequence. Nucleic Acids Res. 24, 257-263.
Sixbey, J. W., Vesterinen, E. H., Nedrud, J. G., Raab-Traub, N., Wlton, L. A., and Pagano, J. S. (1983). Replication of Epstein-Barr virus in human epithelial cells infected in vitro. Nature 306, 480-483.
Smith, R. F. and Smith, T. F. (1989). Identification of a new protein kinase-related genes in three herpesviruses, herpes simplex virus, varicella-zoster virus, and Epstein-Barr virus. Journal of Virology 63, 450-455.
Sugihara, H., Andrisani, V., and Salvaterra, P. M. (1990). Drosophila choline acetyltransferase uses a non-AUG codon and full length RNA is inefficiently translated. Journal of Biological Chemistry 265, 21714-21719.
Sullivan, V., Talarico, C. L., Stanat, S. C., Davis, M., Coen, D. M. and Biron, K. K., (1992). A protein kinase homologue controls phosphorylation of ganciclovir in human cytomegalovirus-infected cells. Nature 358, 162-164.
Talarico, C. L., Burnette, T. C., Miller, W. H., Smith S. L., Davis, M. G., Stanat, S. C., Hg, T. I., He, Z., Coen, D. M., Roizman, B., and Biron, K. K. (1999). Acyclovir is phosphoryalted by the human cytomegalovirus UL97. Antimicrobial Agents and Chemotherapy 43, 1941-1946.
Tsai, C. H., and Glaser, R. (1991). A comparison of Epstein-Barr virus specific proteins expressed by three Epstein-Barr virus isolates using specific monoclonal antibodies. Intervirology 32, 376-382.
Tsurumi, T., Daikoku, T., and Nishiyama, Y. (1993). Functional interaction between Epstein-Barr virus DNA polymerase catalytic subunit and its accessory subunit in vitro. Journal of Virology 67, 7648-7653.
Westphal, E., Mauser, A., Swenson, J., Davis, M. G., Talarico, C. L., and Kenney, S. C. (1999). Induction of lytic Epstein-Barr virus (EBV) infection in EBV-associated malignancies using adenovirus vectors in vitro and in vivo. Cancer Research 59, 1485-1491.
Wolf, D. G., Courcelle, C. T., Prichard, M. N., and Mocarski, E. S. (2001). Distinct and separate roles for herpesvirus-conserved UL97 kinase in cytomegalovirus DNA synthesis and encapsidation. Proceedings of the National Academy of Sciences USA 98, 1895-1900.
張新杰 (1997). EB病毒BGLF4之基因產物具有蛋白質激?活性。台大微生物學研究所碩士論文
黃曉雯 (1999). EB病毒BGLF4基因產物相關之蛋白質激?活性的研究。台大微生物學研究所碩士論文

QRCODE
 
 
 
 
 
                                                                                                                                                                                                                                                                                                                                                                                                               
第一頁 上一頁 下一頁 最後一頁 top
無相關期刊