The Epstein-Barr virus Pol catalytic subunit physically interacts with theBBLF4-BSLF1-BBLF2/3 complex

The Epstein-Barr virus (EBV)-encoded replication proteins that account for the basic reactions at the replication fork are thought to be the EBV Pol h oloenzyme, consisting of the BALFS Pol catalytic and the BMRF1 Pol accessor y subunits, the putative helicase-primase complex, comprising the BBLF4, BS LF1, and BBLF2/3 proteins, and the BALF2 single-stranded DNA-binding protei n, Immunoprecipitation analyses using anti-BSLF1 or anti-BBLF2/3 protein-sp ecific antibody with clarified lysates of B95-8 cells in a viral productive cycle suggested that the EBV Pol holoenzyme physically interacts with the BBLF4-BSLF1-BBLF2/3 complex to form a large complex. Although the complex w as stable in 500 mM NaCl and 1% NP-40, the BALF5 protein became dissociated in the presence of 0.1% sodium dodecyl sulfate. Experiments using lysates from insect cells superinfected with combinations of recombinant baculoviru ses capable of expressing each of viral replication proteins showed that no t the BMRF1 Pol accessory subunit but rather the BALF5 Pol catalytic subuni t directly interacts with the BBLF4-BSLF1-BBLF2/3 complex. Furthermore, dou ble infection with pairs of recombinant viruses revealed that each componen t of the BBLF4-BSLF1-BBLF2/3 complex makes contact with the BALFS Pol catal ytic subunit, The interactions of the EBV DNA polymerase with the EBV putat ive helicase-primase complex warrant particular attention because they are thought to coordinate leading- and lagging-strand DNA synthesis at the repl ication fork.