P32/TAP, a cellular protein that interacts with EBNA-1 of Epstein-Barr virus

Virology. 1997 Sep 15;236(1):18-29. doi: 10.1006/viro.1997.8739.

Abstract

The Epstein-Barr virus (EBV) EBNA-1 protein has a central role in the maintenance of a latent EBV infection and is the only virus-encoded protein expressed in all EBV-associated tumors. EBNA-1 is required for replication of the episomal form of the latent viral genome and transactivates the latency C and LMP-1 promoters. The mechanisms by which EBNA-1 performs these functions are not known. Here we describe the cloning, expression, and characterization of a cellular protein, P32/TAP, which strongly interacts with EBNA-1. We show that P32/TAP is expressed at high levels in Raji cells and is synthesized as a proprotein of 282 amino acids (aa) that is posttranslationally processed by a two-step cleavage process to yield a mature protein of 209 aa. It has been previously reported that P32/TAP is expressed on the cell surface. Our transient expression assays detected full-length P32/TAP (1-282 aa) in the cytoplasm while mature P32/TAP protein localized to the nucleus. Three lines of evidence support P32/TAP interaction with EBNA-1. First, in the yeast two-hybrid system we mapped two interactive N-terminal regions of EBNA-1, aa 40-60 and aa 325-376, each of which contains arginine-glycine repeats. These regions interact with the C-terminal half of P32/TAP. Second, the full-length cytoplasmic P32/TAP protein can translocate nuclear EBNA-1 into the cytoplasm. Third, P32/TAP co-immunoprecipitated with EBNA-1. We have confirmed that a Gal4 fusion protein containing the C-terminal region of P32/TAP (aa 244-282) transactivates expression from a reporter containing upstream Gal4-binding sites. Deletion of the P32/TAP interactive regions of EBNA-1 severely diminished EBNA-1 transactivation of FRTKCAT in transient expression assays. Our data suggest that interaction with P32/TAP may contribute to EBNA-1-mediated transactivation.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Amino Acid Sequence
  • Animals
  • Arginine
  • Binding Sites
  • Carrier Proteins
  • Cell Line
  • Chloramphenicol O-Acetyltransferase / biosynthesis
  • Chlorocebus aethiops
  • Cloning, Molecular
  • Consensus Sequence
  • DNA Primers
  • DNA, Complementary
  • Epstein-Barr Virus Nuclear Antigens / biosynthesis
  • Epstein-Barr Virus Nuclear Antigens / chemistry
  • Epstein-Barr Virus Nuclear Antigens / metabolism*
  • Gene Library
  • Genome, Viral
  • Glycine
  • Herpesvirus 4, Human / genetics
  • Herpesvirus 4, Human / physiology*
  • Humans
  • Hyaluronan Receptors*
  • Lymphocytes / metabolism
  • Membrane Glycoproteins*
  • Mice
  • Mitochondrial Proteins
  • Molecular Sequence Data
  • Polymerase Chain Reaction
  • Receptors, Complement / biosynthesis
  • Receptors, Complement / chemistry
  • Receptors, Complement / metabolism*
  • Recombinant Fusion Proteins / biosynthesis
  • Recombinant Fusion Proteins / chemistry
  • Recombinant Fusion Proteins / metabolism
  • Saccharomyces cerevisiae
  • Sequence Alignment
  • Transfection

Substances

  • C1QBP protein, human
  • C1qbp protein, mouse
  • Carrier Proteins
  • DNA Primers
  • DNA, Complementary
  • Epstein-Barr Virus Nuclear Antigens
  • Hyaluronan Receptors
  • Membrane Glycoproteins
  • Mitochondrial Proteins
  • Receptors, Complement
  • Recombinant Fusion Proteins
  • complement 1q receptor
  • Arginine
  • Chloramphenicol O-Acetyltransferase
  • Glycine