Complete transformation of embryonal rat fibroblasts by polyomavirus occurs during passage in vitro.
The tumorigenicity of secondary rat embryo fibroblasts transfected with a plasmid harboring a replication origin-defective polyomavirus was found to increase during in vitro propagation. Thus, polyomavirus-transfected cells were found to be more than 10,000-fold more tumorigenic when injected into syngenic rats at 3 months after transfection compared to those injected at an earlier time point. Furthermore, most clones of polyomavirus-transfected cells did not grow in semisolid medium at 52 days after transfection but did grow at 95 days. Addition of glucocorticoid hormones, but not of 25% fetal calf serum, to the growth medium of the early passage cells resulted in limited anchorage-independent growth. An altered level of expression of a number of proteins was found in cells analyzed at different times after transfection. Notably, the expression of a component of the actin filament system, tropomyosin 2, was shown to decrease during growth in vitro. The development of a more fully transformed phenotype at late passages correlated with loss of the requirement for large T-antigen for growth. Thus, cells transfected with a polyomavirus mutant encoding a thermolabile large T-antigen did not grow at the restrictive temperature at 6 weeks after transfection, but grew well at 5 months after transfection. We suggest that these phenomena may be explained by assuming that establishment of rodent fibroblasts, and thereby sensitivity to transformation by middle T-antigen, is not an immediate consequence of expression of large T-antigen but occurs after a period of growth in vitro.
Martens
I
I
Department of Medical Virology, Uppsala University Biomedical Centre, Sweden.
Ramqvist
T
T
Dalianis
T
T
Linder
S
S
D000818:Animals; D002455:Cell Division; D002471:Cell Transformation, Neoplastic; D002478:Cells, Cultured; D002999:Clone Cells; D004261:DNA Replication; D015180:Electrophoresis, Gel, Two-Dimensional; D004622:Embryo, Mammalian; D005347:Fibroblasts / Q000166:cytology; D005814:Genes, Viral; D011905:Genes, ras; D008715:Methionine / Q000378:metabolism; D010957:Plasmids; D011120:Polyomavirus / Q000235:genetics*; D014176:Protein Biosynthesis; D011506:Proteins / Q000302:isolation & purification; D051381:Rats; D013936:Thymidine / Q000378:metabolism; D014162:Transfection
D011506:Proteins; D008715:Methionine; D013936:Thymidine