Sequence microheterogeneity in the long control region of clinical isolates of human papillomavirus type 16.
The polymerase chain reaction (PCR) has been used to amplify the long control region (LCR) of episomal human papillomavirus type 16 from cervical scrape DNA obtained from a woman with no evidence of cervical disease and a woman with cervical intraepithelial neoplasia grade 3 (CIN 3). An 883 base pair fragment containing the entire LCR was cloned into pUC13 and the DNA sequence determined for both isolates and compared with the prototype HPV type 16 LCR DNA sequence. Nucleotide variation was apparent in the LCRs derived from both women. In the case of the sample derived from the woman with no cervical disease, there were three nucleotide deletions, one insertion, four transversions, and three transitions (overall conservation: 98.7%). In contrast, the LCR derived from the woman with CIN 3 showed significantly more nucleotide variation with two nucleotide deletions, one insertion, nine transversions, and ten nucleotide transitions (overall conservation 97.6%). Using computer analyses coupled with available data from DNA footprint studies, the effects of these sequence variations on established transcription factor binding sequences were investigated. Cloning of the LCR derived from the woman without cervical disease into a chloramphenicol acetyl transferase (CAT) promoter screening vector, followed by transfection of HeLa cells with the LCR-CAT construct, revealed that the LCR was a functional promoter but was 4.6-fold less active than an equivalent SV40 early promoter-CAT construct.
University Division of Communicable Diseases, Royal Free Hospital and School of Medicine, Hampstead, United Kingdom.
P J
PJ
Bavin
P G
PG
Walker
V C
VC
Emery
D001483:Base Sequence; D004252:DNA Mutational Analysis; D004279:DNA, Viral / Q000235:genetics / Q000302:isolation & purification; D005260:Female; D014644:Genetic Variation; D006801:Humans; D008969:Molecular Sequence Data; D027383:Papillomaviridae / Q000235:genetics* / Q000302:isolation & purification; D016133:Polymerase Chain Reaction; D012091:Repetitive Sequences, Nucleic Acid; D014412:Tumor Virus Infections / Q000382:microbiology; D002583:Uterine Cervical Neoplasms / Q000382:microbiology; D014626:Vaginal Smears
D004279:DNA, Viral