EPSTEIN-BARR-VIRUS LATENT MEMBRANE-PROTEIN-2 ASSOCIATES WITH AND IS ASUBSTRATE FOR MITOGEN-ACTIVATED PROTEIN-KINASE

The latent membrane protein 2 (LMP2) of Epstein-Barr virus interferes with B-lymphocyte signal transduction through the immunoglobulin (Ig) receptor. Two isoforms of LMP2 exist and differ only in that one isofo rm (LMP2a) contains an N-terminal cytoplasmic domain that the other is oform does not. LMP2a is a phosphoprotein that is phosphorylated on ty rosines and serines in the cytoplasmic domain. GST1-119, a glutathione S transferase (GST) fusion protein containing the 119 amino acids of the cytoplasmic domain, affinity precipitated serine kinase activity f rom BJAB cell extracts. The affinity-precipitated kinase phosphorylate d LMP2a sequences, and kinase activity was increased following inducti on. Probing of Western immunoblots of affinity-precipitated proteins s howed that the Erk1 form of mitogen-activated protein kinase (MAPK) wa s present. Purified MAPK phosphorylated GST fusion proteins containing the cytoplasmic domain of LMP2a and mutational analyses were used to identify S15 and S102 as the sites of in vitro phosphorylation. A poly clonal rabbit antiserum was prepared against a maltose binding protein -LMP2a cytoplasmic domain fusion protein (MBP1-119) and used to immuno precipitate LMP2a from the in vitro-immortalized lymphoblastoid B-cell line B95-8CR. LMP2a immunoprecipitates from B95-8CR contained MAPK as a coprecipitated protein. Cross-linking surface Ig on B95-8CR cells f ailed to induce MAPK activity within the cells. Treatment of B95-8CR w ith phorbol myristate acetate (PMA) was able to bypass the Ig receptor block and activate MAPK activity. Phosphorylation of LMP2a on serine residues increased after PMA induction. The possible role for LMP2a se rine phosphorylation by MAPK in the control of latency is discussed.