Identification of Epstein-Barr virus (EBV) nuclear antigen 2 (EBNA2) target proteins by proteome analysis: activation of EBNA2 in conditionally immortalized B cells reflects early events after infection of primary B cells by EBV.

Abstract

The Epstein-Barr virus (EBV) is a ubiquitous B-lymphotropic herpesvirus associated with several malignant tumors, e.g., Burkitt's lymphoma and Hodgkin's disease, and is able to efficiently immortalize primary B lymphocytes in vitro. The growth program of infected B cells is initiated and maintained by the viral transcription factor EBV nuclear antigen 2 (EBNA2), which regulates viral and cellular genes, including the proto-oncogene c-myc. In our study, patterns of protein expression in B cells with and without EBNA2 were analyzed by two-dimensional polyacrylamide gel electrophoresis and mass spectrometry. For this purpose, we used a conditional immortalization system for EBV, a B cell line (EREB2-5) that expresses an estrogen receptor-EBNA2 fusion protein. In order to discriminate downstream targets of c-Myc from c-Myc-independent EBNA2 targets, we used an EREB2-5-derived cell line, P493-6, in which c-Myc is expressed under the control of a tetracycline-regulated promoter. Of 20 identified EBNA2 target proteins, 11 were c-Myc dependent and therefore most probably associated with proliferation, and one of these proteins was a posttranslationally modified protein, i.e., hypusinylated eIF5a. Finally, to estimate the relevance of EBNA2 targets during early EBV infection, we analyzed the proteomes of primary B cells before and after infection with EBV. The protein expression pattern induced upon EBV infection was similar to that following EBNA2 activation. These findings underscore the value of EREB2-5 cells as an appropriate model system for the analysis of early events in the process of EBV-mediated B-cell immortalization.

• Authors: Martin Schlee, Tanja Krug, Olivier Gires, Reinhard Zeidler, Wolfgang Hammerschmidt, Reinhard Mailhammer, Gerhard Laux, Guido Sauer, Josip Lovric, Georg W Bornkamm
• Journal: Journal of virology (J Virol) Vol. 78 Issue 8 Pg. 3941-52 (Apr 2004) ISSN: 0022-538X [Print] United States
• PMID: 15047810 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• DNA, Viral
• EBNA-2 protein, Human herpesvirus 4
• Epstein-Barr Virus Nuclear Antigens
• MAS1 protein, human
• Proteome
• Proto-Oncogene Mas
• Viral Proteins

Topics

• B-Lymphocytes (cytology, virology)
• Base Sequence
• Cell Transformation, Viral
• DNA, Viral (genetics)
• Epstein-Barr Virus Infections (genetics, metabolism, virology)
• Epstein-Barr Virus Nuclear Antigens (genetics, physiology)
• Gene Expression
• Genes, myc
• Herpesvirus 4, Human (pathogenicity, physiology)
• Humans
• In Vitro Techniques
• Protein Biosynthesis
• Proteome
• Proto-Oncogene Mas
• Viral Proteins