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Cloning of Epstein-Barr virus integration sites in lymphoma cell lines

Research Project

Project/Area Number 15590340
Research Category

Grant-in-Aid for Scientific Research (C)

Allocation TypeSingle-year Grants
Section一般
Research Field Experimental pathology
Research InstitutionOsaka University

Principal Investigator

TAKAKUWA Tetsuya  Osaka University, Graduate School of Medicine, Associate Professor, 医学系研究科, 助教授 (40244933)

Co-Investigator(Kenkyū-buntansha) AOZASA Katsuyuki  Osaka University, Graduate School of Medicine, Professor, 医学系研究科, 教授 (30115985)
中塚 伸一  大阪大学, 医学系研究科, 助手 (90303940)
Project Period (FY) 2003 – 2004
Project Status Completed (Fiscal Year 2004)
Budget Amount *help
¥3,500,000 (Direct Cost: ¥3,500,000)
Fiscal Year 2004: ¥1,700,000 (Direct Cost: ¥1,700,000)
Fiscal Year 2003: ¥1,800,000 (Direct Cost: ¥1,800,000)
KeywordsEpstein-Barr virus / integration / lymphoma / Raji / lymphoblastoid cell line / Rel / Bcl-11A / Bach2
Research Abstract

Epstein-Barr virus (EBV) initially isolated from cultured Burkitt lymphoma (BL) cells, is one of well-known oncogenic virus. The Raji cell line was established from BL tissue and used for research worldwide. Previous study showed that each Raji cell contains an average of 50-60 EBV genome equivalents, and a significant proportion of the EBV genome is linearly integrated into host genome through BamHI-W close to the BamHI-Y fragment. However, a definitive EBV integration site in the chromosome has not been identified as yet. In this study, direct evidence that EBV DNA is integrated into the host genome was provided through cloning of the fragments containing nucleotide sequence of Raji integration sites. Integrated EBV DNA consisted of the BamHI-W fragment at one end and BamHI-D fragment at another end. Both junction sites were highly G/C-rich. The BamHI-W fragment and the adjacent part of chromosome 6 showed 70% homology, while no homology was found between the BamHI-D and adjacent hos … More t sequences. EBV is present at intron 1 of the BACH2 gene which is located on chromosome 6q15. BACH2 mRNA was not expressed in the Raji cell line. Immunohistochemistry confirmed the loss of BACH2 expression at protein level. Because BACH2 is a putative tumor suppressor gene, loss of its expression through EBV integration might contribute to lymphomagenesis. Present findings might indicate a new paradigm for EBV oncogenesis. The NAB-2 line, which was established from a North American Burkitt tumor, was indicated to contain one copy of EBV DNA as the integrated form into chromosome 2p13 of the host genome. To directly demonstrate the integration site of EBV, and to clarify the relation between the integration sites and the oncogenes, fragments containing the nucleotide sequence of NAB-2 integration sites were cloned. EBV were integrated via the terminal repeats. Integration sites located in the clone RP11-440P5 on chromosome 2, between two oncogenes, REL and BCL11A, which is apart from approximately 350 kbp from each other. Expression level of REL was increased in NAB-2 lines. The flanking region of chromosome 2 at the bilateral junction sites showed no homology to the junction sites of EBV. The integration site 2p13 overlaps with common fragile site, FRA2E. NAB-2 cells expressed almost all latent genes but LMP-2A that flanks the terminal repeats, indicating the type III of latent infection of EBV. Integration event in NAB-2 might alter the regulation of the oncogenes and provide advantage for continuous cell proliferation. IB4 is a lymphoblastoid cell line frequently used for the functional analysis of the latent genes of EBV. Previous study indicated that EBV whole genome is integrated tandemly as the linear viral genome into host genome of IB4, although sites of integration have not been determined. Through cloning of the junctional regions between EBV and host genomes, one of the integration sites was identified on the BamHI-C fragments around oriP sequences and another on the EcoRI-I fragment. Both of the integration sites were located on the clone RP11-119H12 of chromosome 4q25 and separated approximately 6.5 kbp from each other. The integration sites identified were apart from the genes of the host genome, indicating that both host gene and EBV latent genes are not altered by the integration event. Less

Report

(3 results)
  • 2004 Annual Research Report   Final Research Report Summary
  • 2003 Annual Research Report
  • Research Products

    (13 results)

All 2005 2004 2003 Other

All Journal Article (10 results) Publications (3 results)

  • [Journal Article] Identification of Epstein-Barr virus integrated sites in lymphoblastoid cell line (IB4)2005

    • Author(s)
      Tetsuya Takakuwa
    • Journal Title

      Virus Research 108(1-2)

      Pages: 133-138

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2004 Annual Research Report 2004 Final Research Report Summary
  • [Journal Article] Identificatioan of Epstein-Barr virus integrated sites in lymphoblastoid cell line (IB4).2005

    • Author(s)
      Takakuwa T.
    • Journal Title

      Virus Research 108

      Pages: 133-138

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2004 Final Research Report Summary
  • [Journal Article] Integration of Epstein-Barr virus into chromosome 6q15 of Burkitt lymphoma cell line (Raji) induce loss of BACH2 expression2004

    • Author(s)
      Tetsuya Takakuwa
    • Journal Title

      American Journal of Pathology 164(3)

      Pages: 967-974

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2004 Annual Research Report 2004 Final Research Report Summary
  • [Journal Article] Epstein-Barr virus is integrated between REL and BCL-11A in American Burkitt lymphoma cell line (NAB-2)2004

    • Author(s)
      Wen-Juan Luo
    • Journal Title

      Laboratory investigation 84(9)

      Pages: 1193-1199

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2004 Annual Research Report 2004 Final Research Report Summary
  • [Journal Article] A 50-bp Insertion from Intron 2 Between Exons 2 and 3 of NBS1 May be a Spliced Variant.2004

    • Author(s)
      Tetsuya Takakuwa
    • Journal Title

      Genes Chrom Cancer 39

      Pages: 341-342

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2004 Final Research Report Summary
  • [Journal Article] Integration of Epstein-Barr virus into chromosome 6q15 of Burkitt lymphoma cell line (Raji) induce loss of BACH2 expression.2004

    • Author(s)
      Takakuwa T.
    • Journal Title

      Am J Pathol 164

      Pages: 967-974

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2004 Final Research Report Summary
  • [Journal Article] Epstein-Barr virus is integrated between REL and BCL-11A in American Burkitt lymphoma cell line (NAB-2).2004

    • Author(s)
      Luo WJ.
    • Journal Title

      Lab Invest. 84

      Pages: 1193-1199

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2004 Final Research Report Summary
  • [Journal Article] A 50-bp Insertion from Intron 2 Between Exons 2 and 3 of NBS1 May be a Spliced Variant.2004

    • Author(s)
      Takakuwa T.
    • Journal Title

      Genes Chrom Cancer 39

      Pages: 341-342

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2004 Final Research Report Summary
  • [Journal Article] Establishment and characterization of unique cell lines derived from pyothorax-associated lymphoma which develops in Ion g-standing pyothorax and strongly associated Epstein-Barr virus infection.2003

    • Author(s)
      Tetsuya Takakuwa
    • Journal Title

      Cancer Science 94

      Pages: 858-863

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      2004 Final Research Report Summary
  • [Journal Article] Establishment and characterization of unique cell lines derived from pyothorax-associated lymphoma which develops in long-standing pyothorax and strongly associated Epstein-Barr virus infection.2003

    • Author(s)
      Takakuwa T.
    • Journal Title

      Cancer Sci 94

      Pages: 858-863

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      2004 Final Research Report Summary
  • [Publications] Tetsuya Takakuwa: "Integration of Epstein-Barr virus into chromosome 6q15 of Burkitt lymphoma cell line (Raji) induce loss of BACH2 expression"American Journal of Pathology. 164(3). 967-974 (2004)

    • Related Report
      2003 Annual Research Report
  • [Publications] Tetsuya Takakuwa: "Identification of Epstein-Barr virus integrated sites in lymphoblastoid cell line (IB4)"Virology. in press.

    • Related Report
      2003 Annual Research Report
  • [Publications] Wen-Juan Luo: "Epstein-Barr virus is integrated between REL and BCL-11A in American Burkitt lymphoma cell line (NAB-2)"Laboratory investigation. in press.

    • Related Report
      2003 Annual Research Report

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Published: 2003-04-01   Modified: 2016-04-21  

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