Strain-specific transcription and translation of the BamHI Z area of Epstein-Barr Virus

Abstract

The expression of the 1,800-base-pair BamHI Z region of Epstein-Barr virus DNA was analyzed by hybrid-selected translation with several DNA subclones and RNA from different cell lines. Furthermore, large segments of the three reading frames extending in this area were expressed as fusion proteins into Escherichia coli. The fusion proteins were partially purified and used to immunize rabbits. These sera were used to confirm our mapping assignments and to identify the respective posttranslationally modified proteins in in vivo labeling experiments. The reading frame BRLF1 (the first reading frame starting in the BamHI R fragment in leftward orientation) encoded a 93- to 96-kilodalton (kDa) protein depending on the cell line. The molecular weight of in vivo-labeled proteins was increased relative to that of in vitro-translated proteins, indicating that a posttranslational modification had occurred. The BZLF1 reading frame encoded a 35-kDa protein. It was posttranslationally cleaved from a 38-kDa precursor in induced B95-8 and induced Raji cells and from a 40-kDa precursor in induced P3HR1 cells. In Raji cells superinfected with virus derived from P3HR1 cells, the protein seemed to be expressed both from endogenous Raji genomes and from infecting genomes. The transcripts for the 93- to 96-kDa and the 35-kDa protein overlapped partially. The serum against the expressed third reading frame BZLF2 specifically precipitated a 140-kDa protein. This reading frame contains only 650 nucleotides, and therefore further coding sequences were presumably spliced to BZLF2. The latter is deleted in the Raji cell line; therefore, the observed 140kDa protein in superinfected Raji cells was expressed from infecting P3HR1 genomes.