Kaposi's sarcoma-associated herpesvirus ORF6 gene is essential in viral lytic replication.

ABSTRACT: Kaposi's sarcoma associated herpesvirus (KSHV) is associated with Kaposis's sarcoma (KS), primary effusion lymphoma and multicentric Castleman's disease. KSHV encodes at least 8 open reading frames (ORFs) that play important roles in its lytic DNA replication. Among which, ORF6 of KSHV encodes an ssDNA binding protein that has been proved to participate in origin-dependent DNA replication in transient assays. To define further the function of ORF6 in the virus life cycle, we constructed a recombinant virus genome with a large deletion within the ORF6 locus by using a bacterial artificial chromosome (BAC) system. Stable 293T cells carrying the BAC36 (wild type) and BAC?6 genomes were generated. When monolayers of 293T-BAC36 and 293T-BAC?6 cells were induced with 12-O-tetradecanoylphorbol-13-acetate (TPA) and sodium butyrate, infectious virus was detected from the 293T-BAC36 cell supernatants only and not from the 293T- BAC?6 cell supernatants. DNA synthesis was defective in 293T-BAC?6 cells. Expression of ORF6 in trans in BAC?6-containing cells was able to rescue both defects. Our results provide genetic evidence that ORF6 is essential for KSHV lytic replication. The stable 293T cells carrying the BAC36 and BAC?6 genomes could be used as tools to investigate the detailed functions of ORF6 in the lytic replication of KSHV.

SUBMITTER: Peng C 

PROVIDER: S-EPMC4050029 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature