Felis domesticus papillomavirus, isolated from a skin lesion, is related to canine oral papillomavirus and contains a 1·3 kb non-coding region between the E2 and L2 open reading frames

Masanori Terai1 and Robert D. Burk1,2

Department of Microbiology & Immunology1, and Departments of Pediatrics, Obstetrics & Gynecology and Women’s Health, and Epidemiology & Social Medicine2, Comprehensive Cancer Center, Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461, USA

Author for correspondence: Robert Burk. Fax +1 718 430 8975. e-mail burk{at}aecom.yu.edu


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We have characterized the complete genome (8300 bp) of an isolate of Felis domesticus papillomavirus (FdPV) from a domestic cat with cutaneous papillomatosis. A BLAST homology search using the nucleotide sequence of the L1 open reading frame demonstrated that the FdPV genome was most closely related to canine oral papillomavirus (COPV). A 384 bp non-coding region (NCR) was found between the end of L1 and the beginning of E6, and a 1·3 kbp NCR was located between the end of E2 and the beginning of L2. Phylogenetic analysis placed FdPV in the E3 clade with COPV. Both viruses contain the atypical second NCR, which has no homology with sequences in existing databases.


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Papillomaviruses (PVs) are a heterogeneous group of highly species-specific DNA viruses with closed-circular double-stranded DNA genomes about 8 kbp in size. PVs cause benign and malignant proliferative lesions of squamous and mucosal epithelial surfaces in a wide range of animal species (Sundberg et al., 1996 ). Many domestic and native species of mammals can be infected by one or more species-specific PVs (Sundberg et al., 1996 ).

To understand better the evolution and genetic determinants of PV pathogenicity, comparative molecular analysis of a wide variety of PV genomes is required. PVs have been detected in a variety of Felidae, including domestic cats (for review, see Sundberg et al., 2000 ). Domestic cats have been in close proximity to humans for centuries. Thus, they present an opportunity for horizontal transmission. To explore the origins and molecular characteristics of the cutaneous papillomavirus that infects domestic cats (Felis domesticus), we determined the complete nucleotide sequence of an isolate of F. domesticus papillomavirus (FdPV).

An FdPV genome isolated from a domestic cat with cutaneous papillomatosis was cloned in the EcoRI site of pUC18 (Carney et al., 1990 ; Sundberg et al., 2000 ). The recombinant FdPV genome was amplified and purified (Qiagen Plasmid Mini Kit). To determine the nucleotide sequence, the cloned FdPV DNA was directly sequenced, initially with primers selected from the vector sequence and thereafter with additional primers designed by sequence walking. Sequencing was performed in the Einstein DNA sequencing core facility. The overlapping sequences were assembled manually and confirmed by sequencing the complementary strand. Several additional primers were designed and used to clarify sequence ambiguities. Once assembled, the sequence was analysed for homology with other PVs using BLAST software (Altschul et al., 1997 ). The same software was used for protein sequence comparisons. Phylogenetic trees were created using PV sequences available from GenBank and The Human Papillomaviruses 1997 Compendium Online (http://hpv-web.lanl.gov/stdgen/virus/hpv/compendium/htdocs/HTML_FILES/HPVcompintro4.html#comp97). Phylogenetic trees were derived from individual open reading frames (ORFs) and non-coding regions (NCRs) to determine the relationship of FdPV to the available PV sequences using public domain software (Higgins & Sharp, 1988 ).

A BLAST homology search using the nucleotide sequence of the FdPV L1 ORF revealed that it was most closely related to canine oral papillomavirus (COPV) (86% identity). The assembled sequence of the viral genome had a total size of 8300 bp with a G+C content of 46·12%. Examination of the FdPV sequence for potential genes showed a typical complement of PV ORFs, including overlaps between E1 and E2 and between L2 and L1, and the inclusion of an E4 ORF within E2. There was no initiation codon in the E4 ORF. The predicted ORFs are summarized in Table 1(a). The presence of an E5 ORF situated between the end of the E2 ORF and the start of the L2 ORF, which is found in some but not all PVs, was sought by comparison of all FdPV ORFs in this region to the complete PV database. None of the small FdPV ORFs in this region showed significant homology with known E5 ORFs. Table 1(b) shows the degree of identity between putative FdPV proteins and the analogous proteins of the closest related PV, COPV. The identity of the amino acid sequence of the L1 ORF between FdPV and COPV was relatively higher (75·3%) than other ORFs, with identity to E1 (60·5%) and L2 (59·4%) also elevated compared with the E7 (53·1 %), E2 (50·5%) and E6 (40·4%) ORFs. This suggests that the E1, L1 and L2 ORFs are more conserved than the other ORFs, consistent with their essential role during PV evolution. To investigate the relationship of the FdPV genome with other PV sequences, the complete nucleotide sequence of FdPV was aligned with the corresponding sequences of other closely related PVs. The resulting phylogenetic tree was calculated based on available full-length sequences (de Villiers, 2001 ). A representative tree is shown in Fig. 1. FdPV was placed into the E3 group.


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Table 1. Characterization of the FdPV genome

 


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Fig. 1. Phylogenetic tree based on the alignment of the full sequences of the indicated and related PV genomes categorized in group E (de Villiers, 2001 ). FdPV, Felis domesticus papillomavirus; COPV, canine oral papillomavirus; HPV1, human papillomavirus type 1; HPV63, human papillomavirus type 63; CRPV, cottontail rabbit papillomavirus; ROPV, rabbit oral papillomavirus.

 
Two NCRs were identified in the FdPV genome. Organization and comparison of the FdPV NCRs with the related COPV NCRs are shown in Fig. 2. The usual NCR (NCR-1), found between the end of L1 and the beginning of E6, is short in the FdPV (384 bp) and COPV (360 bp) genomes (see Fig. 2a). In other PVs, the NCR-1 sequence is also called the long control region (LCR) and upstream regulatory region (URR), since it contains numerous control signals for DNA replication and transcription. The FdPV NCR-1 contains three E2 binding sites (ACCN6GGT), although neither the FdPV nor the COPV NCR-1 regions have a discernible E1 binding site (Lu et al., 1993 ; Sun et al., 1996 ). Papillomavirus NCRs also contain multiple binding sites for transcriptional regulatory factors, such as AP-1 (Chan et al., 1990 ), NF-1 (Apt et al., 1993 ), SP-1 (Gloss & Bernard, 1990 ), transcriptional enhancer factor (TEF)-1 (Ishiji et al., 1992 ) and YY-1 (Dong et al., 1994 ; May et al., 1994 ), among others. The FdPV NCR-1 contains a variety of putative regulatory elements, although the FdPV NCR-1 does not include a TATA box within the E6/E7 promoter region. However, multiple SP-1 binding sites were identified that could serve as the transcription initiation site (Peterson et al., 1990 ). The COPV NCR-1 has a TATA box, TEF-1, a poly(A) signal site and two E2 binding sites. The predicted locations of these sites within the FdPV and COPV NCR-1 are shown in Fig. 2(a). Although the FdPV NCR-1 is closely related to the COPV NCR-1 (53·8% sequence identity), the organization of many regulatory elements is altered, as shown in Fig. 2(a).



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Fig. 2. For legend see page 2306.

 


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Fig. 2. Alignment and comparison of the NCR-1 (a) and the NCR-2 (b) sequences of FdPV and the related COPV. NCR sequences and the positions of multiple binding sites are shown. TATA, TATA box; TEF-1, transcriptional enhancer factor-1; E2, E2 binding domain; Poly (A), poly(A) signal.

 
FdPV and COPV have a unique and longer NCR (NCR-2), which is located between the end of E2 and the beginning of L2. The FdPV NCR-2 is 1271 bp in length. The NCR-2 sequence of FdPV was found to be similar to that of COPV (46·7% sequence identity). The A/T content of FdPV and COPV NCR-2 are 60·8 and 64·6%, whereas those of NCR-1 are 51·8 and 57·0%, respectively. Except for the relatedness with COPV, FdPV NCR-1 and the atypical FdPV NCR-2 had no homology with sequences in the existing databases using the BLAST software. The NCR-2 region either originated from an ancient integration event into the PV genome prior to the split of felines and canines, occurred independently in both viruses, or was acquired by host switching (Novacek, 2001 ). Although most closely related to FdPV, COPV was isolated from a mucosal lesion, florid oral papillomatosis, and is grouped into the cutaneous papillomaviruses based on nucleotide sequence homology (Delius et al., 1994 ; Sundberg et al., 1994 ). However, since immunosuppressed dogs will develop cutaneous and oral papillomas (Sundberg et al., 1994 ), it is possible that COPV at one time also infected skin. Whether FdPV can infect mucosal epithelium will require additional study. Based on epitope differences, there appear to be at least two distinct PVs that infect domestic cats (Sundberg et al., 2000 ). We analysed the divergence between the ORFs of FdPV and COPV by amino acid and nucleotide sequence differences as shown in Table 1(c). The nucleotide substitutions of the L1 ORF were predominantly found in the third position; however, codons with extensive third position changes were found in all ORFs.

We have determined the complete nucleotide sequence of a PV isolated and cloned from a domestic cat with cutaneous papillomatous lesions. FdPV is most closely related to COPV by amino acid and nucleotide sequence homology and contains the novel NCR-2 region. PVs are considered highly species-specific and are not thought to cross the species barrier; however, there are exceptions in the veterinary literature (Perrott et al., 2000 ; Sundberg & O'Banion, 1989 ). Nevertheless, a host switch or horizontal movement of virus from one species to another cannot be ruled out, although the divergence between these genomes indicates that they split long ago. In fact, 53–66% of all codons had a third position change, suggesting that the genomes have been saturated by mutations. Subsequently each genome may have evolved as an independent entity by genetic drift. The lack of significant homology with human PVs indicates that recent horizontal transmission has not occurred. The cloning and characterization of FdPV has the potential to be utilized for development of an additional animal model of PV infection.


   Acknowledgments
 
We thank Drs John Sundberg and Marc Van Ranst for providing reagents and helpful suggestions on the manuscript.


   Footnotes
 
The complete nucleotide sequence of FdPV is available in GenBank, accession number AF377865.


   References
Top
Abstract
Main text
References
 
Altschul, S. F., Madden, T. L., Schaffer, A. A., Zhang, J., Zhang, Z., Miller, W. & Lipman, D. J. (1997). Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Research 25, 3389-3402.[Abstract/Free Full Text]

Apt, D., Chong, T., Liu, Y. & Bernard, H. U. (1993). Nuclear factor I and epithelial cell-specific transcription of human papillomavirus type 16. Journal of Virology 67, 4455-4463.[Abstract]

Carney, H. C., England, J. J., Hodgin, E. C., Whiteley, H. E., Adkison, D. L. & Sundberg, J. P. (1990). Papillomavirus infection of aged Persian cats. Journal of Veterinary Diagnostic Investigation 2, 294-299.[Medline]

Chan, W. K., Chong, T., Bernard, H. U. & Klock, G. (1990). Transcription of the transforming genes of the oncogenic human papillomavirus-16 is stimulated by tumor promotors through AP1 binding sites. Nucleic Acids Research 18, 763-769.[Abstract]

Delius, H., Van Ranst, M. A., Jenson, A. B., zur Hausen, H. & Sundberg, J. P. (1994). Canine oral papillomavirus genomic sequence: a unique 1·5-kb intervening sequence between the E2 and L2 open reading frames. Virology 204, 447-452.[Medline]

de Villiers, E. M. (2001). Taxonomic classification of papillomaviruses. Papillomavirus Report 12, 57-63.

Dong, X. P., Stubenrauch, F., Beyer-Finkler, E. & Pfister, H. (1994). Prevalence of deletions of YY1-binding sites in episomal HPV 16 DNA from cervical cancers. International Journal of Cancer 58, 803-808.

Gloss, B. & Bernard, H. U. (1990). The E6/E7 promoter of human papillomavirus type 16 is activated in the absence of E2 proteins by a sequence-aberrant Sp1 distal element. Journal of Virology 64, 5577-5584.[Medline]

Higgins, D. G. & Sharp, P. M. (1988). CLUSTAL: a package for performing multiple sequence alignment on a microcomputer. Gene 73, 237-244.[Medline]

Ishiji, T., Lace, M. J., Parkkinen, S., Anderson, R. D., Haugen, T. H., Cripe, T. P., Xiao, J. H., Davidson, I., Chambon, P. & Turek, L. P. (1992). Transcriptional enhancer factor (TEF)-1 and its cell-specific co-activator activate human papillomavirus-16 E6 and E7 oncogene transcription in keratinocytes and cervical carcinoma cells. EMBO Journal 11, 2271-2281.[Abstract]

Lu, J. Z., Sun, Y. N., Rose, R. C., Bonnez, W. & McCance, D. J. (1993). Two E2 binding sites (E2BS) alone or one E2BS plus an A/T-rich region are minimal requirements for the replication of the human papillomavirus type 11 origin. Journal of Virology 67, 7131-7139.[Abstract]

May, M., Dong, X. P., Beyer-Finkler, E., Stubenrauch, F., Fuchs, P. G. & Pfister, H. (1994). The E6/E7 promoter of extrachromosomal HPV16 DNA in cervical cancers escapes from cellular repression by mutation of target sequences for YY1. EMBO Journal 13, 1460-1466.[Abstract]

Novacek, M. J. (2001). Mammalian phylogeny: genes and supertrees. Current Biology 11, R573-R575.[Medline]

Perrott, M. R., Meers, J., Greening, G. E., Farmer, S. E., Lugton, I. W. & Wilks, C. R. (2000). A new papillomavirus of possums (Trichosurus vulpecula) associated with typical wart-like papillomas. Archives of Virology 145, 1247-1255.[Medline]

Peterson, M. G., Tanese, N., Pugh, B. F. & Tjian, R. (1990). Functional domains and upstream activation properties of cloned human TATA binding protein. Science 248, 1625-1630.[Medline]

Sun, Y. N., Lu, J. Z. & McCance, D. J. (1996). Mapping of HPV-11 E1 binding site and determination of other important cis elements for replication of the origin. Virology 216, 219-222.[Medline]

Sundberg, J. P. & O'Banion, M. K. (1989). Animal papillomaviruses associated with malignant tumors. Advances in Viral Oncology 8, 55-71.

Sundberg, J. P., Smith, E. K., Herron, A. J., Jenson, A. B., Burk, R. D. & Van Ranst, M. (1994). Involvement of canine oral papillomavirus in generalized oral and cutaneous verrucosis in a Chinese Shar Pei dog. Veterinary Pathology 31, 183-187.[Abstract]

Sundberg, J. P., Van Ranst, M., Burk, R. D. & Jenson, A. B. (1996). The nonhuman (animal) papillomaviruses: host range, epitope conservation, and molecular diversity. In Human Papillomavirus Infections in Dermatology and Venereology , pp. 47-68. Edited by G. von Krogh & G. Gross. Boca Raton, FL: CRC Press.

Sundberg, J. P., Van Ranst, M., Montali, R., Homer, B. L., Miller, W. H., Rowland, P. H., Scott, D. W., England, J. J., Dunstan, R. W., Mikaelian, I. & Jenson, A. B. (2000). Feline papillomas and papillomaviruses. Veterinary Pathology 37, 1-10.[Abstract/Free Full Text]

Received 15 February 2002; accepted 19 April 2002.



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