Transcriptional Organization of Bovine Papillomavirus Type 4

Abstract

SUMMARYSeven virus-specific RNA transcripts have been identified in tumours induced by bovine papillomavirus type 4 (BPV-4). The RNAs measured 4.2, 3.6, 3.0, 2.8, 1.9, 1.6 and 1.0 kilobases (kb). They were mapped on the viral genome by Northern blot hybridization to subgenomic probes, by cDNA hybridization to viral DNA fragments and by SI analysis of unlabelled and 3' and 5' end-labelled DNA fragments. All the RNA species are transcribed from the same DNA strand, are polyadenylated and with the exception of the 1.0 kb RNA internally spliced. The 3.0, 1.9, 1.6 and 1.0 kb RNAs share the same 3' polyadenylation site at nucleotide 4009 whereas the 4.2 kb RNA and the 2.8 kb RNA terminate near a polyadenylation site at nucleotide 7187. The 4.2 kb and the 2.8 kb RNAs are transcribed from the late open reading frames and encode the structural polypeptides; the 3.0, 1.9, 1.6 and 1.0 kb RNAs are transcribed from the early open reading frames and are the transcripts involved in viral replication and cellular transformation.

Mapping of Two Novel Transcripts of Bovine Papillomavirus Type 4

“…I a, lane 1), as predicted from the previous BPV-4 transcriptional map (Smith et aL, 1986). However, a probe derived from the 5' exon of cDNA 7Ell by cleaving at nt 1009 with restriction endonuclease Sau96I, also hybridized strongly to the 1 kb transcript ( Fig.…”

“…All of these contained sequences from the 3' end of the early region of BPV-4 (Smith et al, 1986) and two also contained sequences from regions further upstream. These two cDNAs were selected for further analysis.…”

“…Total RNA was prepared from bovine oesophageal papillomas according to the method of Chirgwin et al (1979), as described previously (Smith et al, 1986). Polyadenylated RNA [poly(A) RNA] was isolated by passing total RNA twice through an oligo(dT)-cellulose column (Aviv & Leder, 1972).…”

“…S1 nucleaseprotection analysis. This was carried out according to the method of Berk & Sharp (1977) as described previously (Smith et al, 1986), except that ssDNA probes were isolated from polyaerylamide strand-separation gels (Maniatis et al, 1982) and annealing of probes to poly(A) RNA was carried out at predetermined temperatures as above.…”

“…Since no RNA 3' termini had been mapped in this region (Smith et al, 1986) it was concluded that this was a 3' truncated cDNA. A Northern blot of total papilloma RNA was prehybridized to unlabelled cDNA 7E 11 in order to block hybridization to the 3' transcriptional exon of the early region, and then hybridized to radiolabelled cDNA Q, and to radiolabelled cDNA 7E 11 to monitor the effectiveness of the competition.…”