Effects of Mutations within the Herpes Simplex Virus Type 1 DNA Encapsidation Signal on Packaging Efficiency

Abstract

ABSTRACT The cis -acting signals required for cleavage and encapsidation of the herpes simplex virus type 1 genome lie within the terminally redundant region or a sequence. The a sequence is flanked by short direct repeats (DR1) containing the site of cleavage, and quasi-unique regions, Uc and Ub, occupy positions adjacent to the genomic L and S termini, respectively, such that a novel fragment, Uc-DR1-Ub, is generated upon ligation of the genomic ends. The Uc-DR1-Ub fragment can function as a minimal packaging signal, and motifs have been identified within Uc and Ub that are conserved near the ends of other herpesvirus genomes ( pac 2 and pac 1, respectively). We have introduced deletion and substitution mutations within the pac regions of the Uc-DR1-Ub fragment and assessed their effects on DNA packaging in an amplicon-based transient transfection assay. Within pac 2, mutations affecting the T tract had the greatest inhibitory effect, but deletion of sequences on either side of this element also reduced packaging, suggesting that its position relative to other sequences within the Uc-DR1-Ub fragment is likely to be important. No single region essential for DNA packaging was detected within pac 1. However, mutants lacking the G tracts on either side of the pac 1 T-rich motif exhibited a reduced efficiency of serial propagation, and alteration of the sequences between DR1 and the pac 1 T element also resulted in defective generation of Ub-containing terminal fragments. The data are consistent with a model in which initiation and termination of packaging are specified by sequences within Uc and Ub, respectively.