Molecular Cloning of Bovine Papillomavirus Genomes and Comparison of Their Sequence Homologies by Heteroduplex Mapping

The L-X-C-X-E pRB-binding motif of papillomavirus (PV) E7 proteins has been implicated in the immortalization and transformation of the host cell. However, sequencing of the complete genomes of bovine papillomavirus type 3 (BPV-3), bovine papillomavirus type 5 (BPV-5), equine papillomavirus (EQPV) and reindeer (Rangifer tarandus) papillomavirus (RPV) supports the notion that the pRB-binding motif is not ubiquitous among E7 proteins in the PV proteome. Key among the animal groups that lack the pRB-binding domain are the artiodactyl PVs, including European elk PV (EEPV), deer PV (DPV), reindeer PV (RPV), ovine PVs types 1 and 2 (OvPV-1 and -2) and bovine PVs 1, 2 and 5 (BPV-1, -2 and -5). Whereas the presence of the pRB-binding domain is normally associated with papillomas, the artiodactyl PVs are marked by the development of fibropapillomas on infection. Previous studies emphasized the role of E5 in the pathogenic mechanism of fibropapilloma development, but correlation between the lack of an E7 pRB-binding domain and the unique pathology of the artiodactyl PVs suggests a more complicated mechanism and an early evolutionary divergence from a pRB-binding ancestor. INTRODUCTIONPapillomaviruses (PVs) are highly species-specific pathogens. They form a heterogeneous group of closed-circular, double-stranded DNA viruses measuring about 8 kb in size (Sundberg et al., 1996). Most PVs target the basal cells of dermal or mucosal epithelia (Cheville & Olson, 1964), and infection has been implicated in both benign and malignant lesions of epithelia in a broad range of animals (Sundberg & O'Banion, 1989). Though almost all known human PVs (HPVs) infect dermal or mucosal surfaces, in some animal PVs, most notably in the artiodactyl ruminant PVs, fibroblasts appear to be the primary target cells (Sundberg et al., 1985). However, regardless of species, infection generally manifests as a papilloma or fibropapilloma, and follows a cytopathic mechanism of cell proliferation (Sundberg et al., 1996). Historically, PVs had been classified according to their tissue tropism, and this grouping was supported by later phylogenetic analysis of PV sequence data (Chan et al., 1995;de Villiers, 2001;Myers, 1994). PV phylogenies typically subdivide into mucosal/genital HPVs, cutaneous EV HPVs and three main animal PV clades: the artiodactyl ruminant PVs, the distant avian PV group and a group containing canine, feline, rabbit and rodent PV types. However, sequence analysis has highlighted some significant exceptions. BPV-3, BPV-4 and BPV-6 do not group with the artiodactyl PVs, but instead form an isolated taxon (Jarrett et al., 1984), and HPV-1, HPV-41 and HPV-63 are most closely related to the canine and feline PVs, sharing little similarity to HPVs in either the mucosal or the cutaneous groups (Egawa et al., 1993).Although most work in the area has focused on HPVs, an ever-increasing number of animal PV isolates have been sequenced. These animal PVs share key clinical features of GenBank data deposited: bovine papillomavirus 3, AF486184; bo...

Section: Methodsmentioning

confidence: 99%

Lack of the canonical pRB-binding domain in the E7 ORF of artiodactyl papillomaviruses is associated with the development of fibropapillomas

Narechania

Terai

Chen

et al. 2004

“…Molecularly cloned BPV-4 DNA (Campo & Coggins, 1982) or derived subcl0nes were digested with the appropriate restriction endonucleases, subjected to electrophoresis and transferred to Pall Biodyne A nylon membranes (Southern, 1975). Hybridization was done following the manufacturers' recommendations.…”

Section: Dna Blot Hybridizationmentioning

confidence: 99%

“…The digested DN A was electrophoresed in low melting point (LMP) agarose gels and the required fragment was eluted from the gel and purified (Weislander, 1979). The DNA fragments were ligated to the vector pAT153 (Twigg & Sherratt, 1980) cleaved with the appropriate restriction endonuclease, and recombinant plasmids were identified as described previously (Campo & Coggins, 1982). The subclones are referred to by the first letter of the restriction endonuclease sites bordering the fragments and by the size in kilobases (kb) with nucleotide positions in brackets.…”

Section: Molecular Cloning Of Viral Fragmentsmentioning

confidence: 99%

“…Subgenomic fragments of BPV-4 were prepared by the cleavage of the whole recombinant pBV4 plasmid (Campo & Coggins, 1982) with appropriate restriction endonucleases. The digested DN A was electrophoresed in low melting point (LMP) agarose gels and the required fragment was eluted from the gel and purified (Weislander, 1979).…”

Section: Molecular Cloning Of Viral Fragmentsmentioning

confidence: 99%

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Transcriptional Organization of Bovine Papillomavirus Type 4

Smith

Patel

Campo

1986

Self Cite

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.

“…The DNA sequence of BPV-1 has been determined, and analysis of the open reading frames suggests that the genome is transcribed unidirectionally from one strand (Chen et al, 1982). We have shown previously that the genomes of BPV-1 and 2 are colinear, though mismatched, and thus these viruses probably share a similar genetic organization (Campo & Coggins, 1982). In this paper we compare the genomes of BPV-3 and 4.…”

mentioning

confidence: 99%

The Genomes of Bovine Papillomaviruses Types 3 and 4 are Colinear

Coggins

Hettich

Smith

et al. 1983

Self Cite

SUMMARYThe 7.2 kb genomic DNA of bovine papiUomavirus type 3 (BPV-3) was molecularly cloned using its unique EcoRI site, and the 7.3 kb genome of BPV-4 was cloned using its single BamHI site. The viral genomes were compared by liquid hybridization, Southern blot hybridization and heteroduplex mapping. Low stringency hybridization conditions revealed that the genomes are colinear but the sequences are extensively mismatched. The relative alignment of the restriction endonuclease maps of the two viral genomes has been determined. It was found that the genomes as linearized for cloning are out of phase by 1.7 kb, so that the single EcoRI site of BPV-3 appears to coincide with the BPV-4 EcoRI site at 0.22 map units. It is concluded that the genomes of BPV-3 and BPV-4, both of which cause true epithelial warts, share the same physical organization but exhibit sequence divergence.