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A major transcript of human papillomavirus type 16 in transformed NIH 3T3 cells contains polycistronic mRNA encoding
1Laboratory of Molecular and Cellular Biology, Kanagawa Cancer Center Research Institute, Yokohama, Japan.
Abstract:
We have cloned cDNA of the major 1.8 kb mRNA from HPV 16-transformed NIH 3T3 cells (PM3T3). The entire nucleotide sequences of this cDNA were determined and compared with prototype HPV 16 genomic DNA sequences. The 5'-end of the cDNA was flanked by approximately 300 bp of cellular sequences, and the 3'-end of the cDNA sequences contained poly A residues following at nt 4230. HPV 16 sequences began at nt 124, downstream of a major viral p97 promoter, within the E6 open reading frame (ORF). The first splice donor site was at nt 226 and the splice acceptor site was at nt 409, suggesting that the E6 gene is inert. Second splice donor and acceptor sites were located at nt 880 and at nt 3357, respectively. This mRNA was thus shown to consist of three exons, resulting in polycistronic mRNA containing three potentially functional virus early genes--E7, E1--E4, and E5--actively transcribed in the transformant.
Insights
Researchers identified a Human Papillomavirus type 16 (HPV 16) mRNA in transformed cells. This polycistronic mRNA contains potentially functional early genes E7, E1-E4, and E5, suggesting active viral gene transcription.
Area of Science:
- Molecular Biology
- Virology
- Cancer Research
Background:
- Human Papillomavirus type 16 (HPV 16) is a major cause of cervical cancer.
- Understanding HPV 16 gene expression is crucial for developing antiviral strategies.
Purpose of the Study:
- To characterize the major 1.8 kb mRNA transcribed from HPV 16 in transformed cells.
- To determine the gene content and potential functionality of this mRNA.
Main Methods:
- Cloning and sequencing of the 1.8 kb cDNA from HPV 16-transformed NIH 3T3 cells (PM3T3).
- Comparison of cDNA nucleotide sequences with prototype HPV 16 genomic DNA.
- Analysis of splice donor and acceptor sites to define mRNA exons and open reading frames (ORFs).
Main Results:
- The cloned cDNA contained cellular sequences at the 5'-end and poly A residues at the 3'-end.
- HPV 16 sequences initiated within the E6 ORF, with splice sites indicating the E6 gene is likely non-functional.
- The mRNA comprises three exons, forming a polycistronic transcript encoding potentially functional E7, E1-E4, and E5 genes.
Conclusions:
- The major 1.8 kb mRNA in HPV 16-transformed cells is a polycistronic transcript.
- This transcript actively transcribes potentially functional early viral genes E7, E1-E4, and E5.
- The E6 gene appears to be transcriptionally inactive in this context.