Coestablishment of persistent infection and oncogenic transformation of hamster embryo cells by equine cytomegalovirus

Abstract

Semipermissive, primary hamster embryo (HE) cells were morphologically transformed in vitro by infection with UV-irradiated equine cytomegalovirus (equine herpesvirus type 2; ECMV). Cell lines (designated EC-1-3) were established independently from foci and were shown to exhibit growth and biological properties typically associated with transformed cells: altered morphology, loss of contact inhibition, increased saturation density, decreased generation time, immortality in culture, normal growth in low concentrations of serum, colony formation in soft agar, and resistance to ECMV superinfection. All ECMV transformed cells were restrictive for the replication of equine herpesvirus type 1 (EHV-1) which shares 2.9% homology with ECMV and replicated to high titers in normal HE cells. All EC cell lines were oncogenic in immunocompetent syngeneic LSH hamsters. Tumor cell lines were established from selected malignant fibrosarcomas that developed at the site of injection. Two of the transformed cell lines (EC-2 and EC-3) were found to be persistently infected and to release infectious ECMV from 0.5 to 2% (EC-2) and 0.8 to 5% (EC-3) of the total cell populations. The transformed cell line EC-1 as well as all tumor cell lines were virus nonproducers. Data from DNA-DNA reassociation analyses indicated the presence of 8-32 ECMV genome equivalents per productive EC-2 cell and greater than 300 ECMV genome equivalents per productive EC-3 cell. Small amounts of subgenomic ECMV DNA sequences were detected in the nonproducer EC-1 transformed cells and in all tumor cell lines (EC-1T, -2T, -3T). Some of these DNA sequences must be expressed since ECMV-specific polypeptides were demonstrated in all transformed and tumor cell lines by indirect immunofluorescence using antiserum to ECMV-infected cell extracts and since the sera of tumor-bearing hamsters contained ECMV antibody as detected by an ECMV plaque reduction assay.