Messenger RNA sequence rather than protein sequence determines the level of self-synthesis and antigen presentation of the EBV-encoded antigen, EBNA1

PLoS Pathog. 2012 Dec;8(12):e1003112. doi: 10.1371/journal.ppat.1003112. Epub 2012 Dec 27.


Unique purine-rich mRNA sequences embedded in the coding sequences of a distinct group of gammaherpesvirus maintenance proteins underlie the ability of the latently infected cell to minimize immune recognition. The Epstein-Barr virus nuclear antigen, EBNA1, a well characterized lymphocryptovirus maintenance protein has been shown to inhibit in cis antigen presentation, due in part to a large internal repeat domain encoding glycine and alanine residues (GAr) encoded by a purine-rich mRNA sequence. Recent studies have suggested that it is the purine-rich mRNA sequence of this repeat region rather than the encoded GAr polypeptide that directly inhibits EBNA1 self-synthesis and contributes to immune evasion. To test this hypothesis, we generated a series of EBNA1 internal repeat frameshift constructs and assessed their effects on cis-translation and endogenous antigen presentation. Diverse peptide sequences resulting from alternative repeat reading frames did not alleviate the translational inhibition characteristic of EBNA1 self-synthesis or the ensuing reduced surface presentation of EBNA1-specific peptide-MHC class I complexes. Human cells expressing the EBNA1 frameshift variants were also poorly recognized by antigen-specific T-cells. Furthermore, a comparative analysis of the mRNA sequences of the corresponding repeat regions of different viral maintenance homologues highlights the high degree of identity between the nucleotide sequences despite very little homology in the encoded amino acid sequences. Based on these combined observations, we propose that the cis-translational inhibitory effect of the EBNA1 internal repeat sequence operates mechanistically at the nucleotide level, potentially through RNA secondary structural elements, and is unlikely to be mediated through the GAr polypeptide. The demonstration that the EBNA1 repeat mRNA sequence and not the encoded protein sequence underlies immune evasion in this class of virus suggests a novel approach to therapeutic development through the use of anti-sense strategies or small molecules targeting EBNA1 mRNA structure.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Amino Acid Sequence / genetics*
  • Antigen Presentation / genetics
  • Antigen Presentation / immunology*
  • Base Sequence / genetics*
  • Cell Line
  • Epstein-Barr Virus Nuclear Antigens* / genetics
  • Epstein-Barr Virus Nuclear Antigens* / immunology
  • Epstein-Barr Virus Nuclear Antigens* / metabolism
  • Frameshift Mutation
  • Genes, MHC Class I
  • HEK293 Cells
  • Humans
  • Immune Evasion
  • Molecular Sequence Data
  • RNA, Messenger / genetics*
  • Repetitive Sequences, Amino Acid / genetics
  • Repetitive Sequences, Amino Acid / immunology
  • Sequence Alignment
  • T-Lymphocytes, Cytotoxic / immunology


  • Epstein-Barr Virus Nuclear Antigens
  • RNA, Messenger
  • EBV-encoded nuclear antigen 1

Grant support

This work was funded by Project Grants from the National Health & Medical Research Council (NH&MRC) Canberra, Australia (#496684; APP1005091) and by a NH&MRC Career Development Award Research Fellowship (#496712). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.