Four of five spontaneous benign equine connective tissue tumors of unknown etiology and a bovine papilloma virus (BPV)induced equine tumor contained BPV-specific DNA sequences as determined by DNA-DNA hybridization of DNA from tumors with BPV DNA labeled in vitro. Analysis of the kinetics of reassociation indicated that 20-75% of the BPV genome was present in the various tumors. The number of partial BPV genome equivalents ranged from 60 to 500 copies per diploid quantity of cellular DNA. Thermal denaturation profiles of duplexes formed between labeled BPV DNA and DNA from tumor cells indicated two tumors contained viral DNA with base sequences identical to BPV DNA. Three tumors (including DNA from the BPV-induced tumor) contained BPV-related DNA sequences that were less thermally stable. The decrease in thermal denaturation temperature may be due to the presence of (adenine + thymine).rich regions of the BPV genome in the tumor cells. The papilloma viruses are a subgroup of the Papovaviridae and produce a self-limiting neoplastic disease in their natural hosts. The bovine papilloma virus (BPV) is the most oncogenic of the papilloma viruses. Intracranial injection of BPV produces meningiomas in calves, while subcutaneous injection of the virus induces fibromas in hamsters and C3H/eB mice (1-3). BPV can transform fetal bovine cells and mouse embryo cells in culture (4,5). Transformation of fetal bovine skin cells by BPV DNA has also been reported (6).Intradermal inoculation of BPV into horses and related species can induce a connective tissue tumor that is histologically similar to equine sarcoid, a spontaneous connective tissue tumor (7, 8). Equine sarcoid is one of the most commonly found tumors in horses and does not appear to metastasize, but excision often results in recurrence of the tumor (9, 10). BPV-induced and naturally occurring equine tumors differ in that induced tumors regress while spontaneous forms do not (7, 10). BPVneutralizing antibodies are not detectable in horses with natural tumors, while neutralizing antibodies are produced in horses with virus-induced disease (11). BPV is equally efficient in inducing sarcoids in normal horses and horses with existing spontaneous tumors (11). Although the epidemiology of equine sarcoid suggests a viral etiology for the disease (12), virus particles have not been detected in or recovered from spontaneous tumors (C. Olson, unpublished observation). However, oncornavirus-like particles have been observed in cell lines derived from spontaneous equine sarcoids (13).Abbreviations: BPV, bovine papilloma virus; A+T, adenine + thymine;Tm, temperature at which 50% of the hybrid was eluted from hydroxyapatite.

Short-lived nucleoprotein complexes (r-py complex) containing replicating polyoma DNA were isolated from infected cells after lysis with Triton X-100. The Triton lysing procedure of Green, Miller, and Hendler (1971) releases most complexes containing supercoiled viral DNA (py complex) from nuclei, but liberates only a portion of r-py complexes. r-py Complexes are associated more strongly with nuclear sites but can be extracted by prolonged incubation of nuclei in lysing solution. Complexes containing replicating polyoma DNA appear to be precursors to stable complexes containing supercoiled DNA. Sedimentation and buoyant density studies indicate that protein is bound to both r-py complexes and py complexes at a ratio of protein to DNA of about 1 to 2/1. Both types of complexes sediment as if the viral DNA is more compact than free DNA and both undergo major reversible configurational changes with increased salt concentration. Changes resulting from enzymatic and chemical treatment indicate that there may be two or more protein components in both r-py complex and py complex. One component is digested by Pronase and trypsin while another is resistant to the enzymes but released by deoxycholate. The abundance and similarity in chemical and physical properties of protein bound to all forms of polyoma DNA suggest that part of the protein molecules may serve in a structural capacity.

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