We studied determinants of efficient encapsidation of circular DNA, incorporating a PV early region DNA sequence (nt 584-1978) previously shown to enhance packaging of DNA within papillomavirus (PV)-like particles (VLPs). Insect coelomic cells (Sf-9) and cultured monkey kidney cells (Cos-1) were transfected with an 8-kb reporter plasmid incorporating the putative BPV packaging sequence and infected with BPV1 L1 and L2 recombinant baculovirus or vaccinia virus. Heavy (1.34 g/ml) and light (1.30 g/ml) VLPs were produced, and each packaged some of the input plasmid. In light VLPs, truncated plasmids, which nevertheless incorporated the PV-derived DNA packaging sequence, were more common than full-length plasmids. Packaging efficiency of the plasmid was estimated at 1 plasmid per 10(4) VLPs in both Cos-1 and Sf-9 cells. In each cell type, expression of the BPV1 early region protein E2 in trans doubled the quantity of heavy but not light VLPs and also increased the packaging efficiency of full-length circular plasmids by threefold in heavy VLPs. The resultant pseudovirions incorporated significant amounts of E2 protein. Pseudovirions, comprising plasmids packaged within heavy VLPs, mediated the delivery of packaged plasmid into Cos-1 cells, whereby "infectivity" was blocked by antisera to BPV1 L1, but not antisera to BPV1 E4. We conclude that (a) packaging of DNA within PV L1+L2 pseudovirions is enhanced by BPV1 E2 acting in trans, (b) E2 may be packaged with the pseudovirion, and (c) E2-mediated enhancement of packaging favors 8-kb plasmid incorporation over incorporation of shorter DNA sequences.
Copyright 2000 Academic Press.