THE CELLULAR DNA-POLYMERASE ALPHA-PRIMASE IS REQUIRED FOR PAPILLOMAVIRUS DNA-REPLICATION AND ASSOCIATES WITH THE VIRAL E1 HELICASE

Persistent infection by papillomaviruses involves the maintenance of v iral DNA as a nuclear plasmid, the replication of which requires host DNA polymerases. The role of the cellular DNA polymerase alpha-primase holoenzyme was probed by using soluble extracts from rodent cells tha t replicate bovine papilloma virus 1 and human papilloma virus 6b DNA in the presence of the viral E1 helicase and the E2 transcription fact or. Monoclonal antibodies directed against the catalytic 180-kDa subun it of polymerase alpha inhibit DNA synthesis in this system. Addition of purified human polymerase alpha-primase holoenzyme to neutralized e xtracts restores their DNA synthetic activity. The amino-terminal 424 amino acids of E1 forms a specific protein complex with the p180 polym erase subunit. Immune complexes can be isolated with antibodies direct ed against E1 that contain a DNA polymerase activity. Moreover, this p olymerase activity tan be neutralized by anti-polymerase alpha antibod ies. Permissivity barriers were not encountered in this in vitro syste m, as bovine E1 can interface with the murine and human replication ap paratus. Although the large tumor antigens encoded by simian virus 40 and polyoma share limited primary sequence homology with the papilloma virus E1 proteins, the organization of functional moths at the level o f primary protein structure is remarkably similar. In addition to thei r origin-specific DNA-binding activity, each of these helicases may fu nction to help recruit the cellular polymerase alpha-primase complex t o the-viral replication origin.