Structure of the transforming region of human cytomegalovirus AD169

Abstract

The minimum size fragment of human cytomegalovirus AD169 required to initiate transformation was determined by transfection of primary rat embryo cells with deletion fragments constructed by digestion of a cloned fragment containing the transforming region (pCM4000) with exonuclease III and S1 nuclease. The results indicate that the left-hand boundary of the minimum size sequence for transformation must reside between 490 and 318 bases from the HindIll site of pCM4000. The right-hand boundary was defined by the EcoRI site which is 20 bases from the HindIll site. The nucleotide sequence of the transforming fragment of human cytomegalovirus was determined by the chemical degradation technique of Maxam and Gilbert and with the dideoxynucleoside triphosphate chain termination method. The sequence of pCM4000 comprises 2,848 base pairs and has an A * T composition of 59.5%. Reading frame analysis of the sequence indicated the longest open reading frame was 118 amino acids in length. Northern blot analysis of polyadenylated and non-polyadenylated RNA extracted from cells at immediate early and late times after infection with human cytomegalovirus indicate that one 5.0-kilobase RNA species hybridized to pCM4000 (Jahn et al., J. Virol, in press). The direction of transcription was determined by hybridization of this transcript with M13 singlestranded probes, and S1 analysis of this RNA did not detect introns. Human cytomegalovirus (HCMV) a member of the human herpesvirus group, is capable of producing a permissive or latent infection in the human host. HCMV is also able to transform rodent cells in vitro to a malignant phenotype. Albrecht and Rapp (1) transformed hamster embryo fibroblasts with UV-irradiated HCMV. These transformed cells were tumorigenic in weanling Syrian hamsters, but less than 0.5% of the cells stained for HCMV-specific antigen by indirect immunofluorescence. Recently, we have demonstrated transformation of NIH 3T3 and primary rat embryo * Corresponding author.

Low tumorigenicity of canine cells transformed by the human cytomegalovirus

“…Because our results were obtained with a cellular sys-'tem allowing the study of immortalization, it is not surprising that they differ from those of Nelson et al [15,16]. Data obtained with the canine cells indicate that, in contrast with the indications of Clanton et al [3], the HindIII-E fragment of the virus strain AD-169 does not provoke the immortalization of cell cultures.…”

“…Antiserum purchased from Flow Laboratories was used, on the other hand, to detect the large T antigen in SV40-transformed cells. Restriction maps of HSV-1 and HSV-2 (a) as well as those of HCMV (b) were constructed according to [4,7,10,16,19]. imm, immortalization; tum, complete oncogenic transformation (tumorigenic).…”

“…As far as HCMV is concerned, the same situation apparently prevails (fig 1). Work carried out by Nelson et al [15,16] with immortalized cell lines shows that the HindIII-E fragment of the viral genome (strain AD-169) carries DNA sequences related to tumorigenic transformation. Data obtained by Jariwalla et al [3,13] suggest, on the other hand, that the same viral DNA fragment carries a distinct region associated with immortalization.…”

“…A number of experiments have shown that the human cytomegalovirus (HCMV), a member of the herpes virus family, could transform cells in vitro and give them the capacity to induce tumors in vivo [1,2,6]. The viral DNA sequence responsible for neoplastic or tumorigenic transformation of the cells has been identified [15,16].…”

History, precedent, and progress in the development of mammalian cell culture systems for preparing vaccines: Safety considerations revisited

“…The literature on viral carcinogenesis is extensive [Marx, 1989;Lambowitz, 1989;Dyson et al, 1989;Whyte et al, 1988;Hansen and Cavenee, 1988;Klein, 1987;Spriggs, 1986;Bishop, 1985;Rawls, 1985;Tevethia, 1985;Galloway et al, 1984;Nelson et al, 1984;Schafer et al, 1984;Fung et al, 1983;Ruley, 1983;zur Hausen, 1982;Lane and Crawford, 19791; and yet the mechanisms for viral carcinogenesis are incompletely or little understood. It is recognized, however, that cells respond to both positive and negative signals in the regulation of cell proliferation and that the proteins encoded by cellular and foreign genetic materials can be o r are involved in cell signalling.…”