Human epithelial cells immortalized by SV40 retain differentiation capabilities in an in vitro raft system and maintain viral DNA extrachromosomally.
Carcinomas are the predominant type of cancer found in man, yet in vitro studies on the transformation of epithelial cells have been limited. In an attempt to extend our knowledge of the mechanisms involved in the development of epithelial cancers, we have examined the effects of oncogenes on keratinocytes in vitro using both the ability to immortalize and the ability to alter differentiation as criteria for transformation. SV40 T-antigen was observed to be an efficient immortalizing agent in human keratinocytes consistent with previous studies in other human cell types. Using an in vitro cell culture system (rafts) for epithelial stratification at the air-liquid interface, we observed that the morphology of rafts of SV40-immortalized keratinocytes was similar to that of untransfected epithelial cells, demonstrating that although immortal these cells retain differentiation capabilities. The ability to differentiate was lost only upon prolonged passage in culture, suggesting that this effect is separable from immortalization. In these immortalized epithelial cells, SV40 genomes were found to be maintained as a heterogeneous population of extrachromosomal molecules dependent upon the SV40 origin of replication. It is not clear whether these molecules arise continuously as a result of excision events from integrated copies or are stably maintained as episomes.
Lechner
M S
MS
Department of Molecular Genetics and Cell Biology, Howard Hughes Medical Institute, University of Chicago, Illinois 60637.
Laimins
L A
LA
D000952:Antigens, Polyomavirus Transforming / Q000032:analysis; D015139:Blotting, Southern; D015153:Blotting, Western; D002454:Cell Differentiation / Q000235:genetics; D002460:Cell Line; D002472:Cell Transformation, Viral / Q000235:genetics*; D002478:Cells, Cultured; D004279:DNA, Viral / Q000235:genetics; D006801:Humans; D015603:Keratinocytes / Q000166:cytology* / Q000382:microbiology; D010957:Plasmids / Q000235:genetics; D013539:Simian virus 40 / Q000235:genetics*; D014162:Transfection
D000952:Antigens, Polyomavirus Transforming; D004279:DNA, Viral