Characterization of human papillomavirus-11 mRNAs expressed in the context of autonomously replicating viral genomes.
We have previously adapted an experimental system which supports autonomous replication of human papillomavirus-11 (HPV-11) genomic DNA in a human squamous carcinoma cell line (SCC-4) following electroporation of linearized viral DNA. This system allows evaluation of HPV-11 transcription in the context of replicating viral genomes. RNA was isolated from cells following transfection, and first-strand cDNA was produced by reverse transcription using HPV-11-specific primers. The cDNAs were amplified using the polymerase chain reaction, and resulting DNA products were cloned and sequenced. Transcripts were identified that utilize the same splice donor and acceptor sites as transcripts characterized previously from human lesions. Three previously unreported splice junctions that employ novel combinations of those splice sites were also identified. We also detected these newly identified transcripts in laryngeal papillomas. A modified version of the 5' rapid amplification of cDNA ends method was used to identify transcriptional start sites of several of the HPV-11 transcripts. The family of mRNAs characterized in this replication system have the potential to encode all of the major HPV-11 E-region proteins described to date. The data support the utility of this system as an experimental model for examining mechanisms of viral replication and transcription.
Renaud
K J
KJ
Department of Infectious Diseases, Isis Pharmaceuticals, Inc., Carlsbad, California 92008, USA.
Cowsert
L M
LM
D001483:Base Sequence; D004279:DNA, Viral / Q000235:genetics*; D016679:Genome, Viral; D006801:Humans; D008969:Molecular Sequence Data; D027383:Papillomaviridae / Q000235:genetics* / Q000502:physiology; D012333:RNA, Messenger; D012367:RNA, Viral / Q000096:biosynthesis / Q000235:genetics*; D014407:Tumor Cells, Cultured; D014779:Virus Replication
D004279:DNA, Viral; D012333:RNA, Messenger; D012367:RNA, Viral