SV40-transformed human cells in crisis exhibit changes that occur in normal cellular senescence.
SV40 T antigen can induce senescent human diploid fibroblasts to synthesize DNA; however, the cells fail to go through mitosis. In this study, we examined the expression of the cdc2 and cyclin B genes, which are required for completion of mitosis, to determine whether defects in their expression occurred when SV40-transformed human cells entered the phase of crisis. If defects were observed it would indicate that immortalization by the virus involved reexpression of these genes. We found that the expression of cdc2 was unimpaired at both the RNA and protein levels, but that cyclin B expression was decreased in cells in crisis when compared with precrisis (mortal) and postcrisis (immortal) cells. Tritiated thymidine uptake demonstrated that the majority of cells in crisis were not actively cycling. Consistent with the latter observation we found that cyclin A, which is required for cells to traverse through S to G2, was downregulated in these cells. Since many of the results obtained with cells in crisis were similar to what is observed in normal human cells when they become senescent, we analyzed the expression of the genes fibronectin and sdi1 (a gene recently cloned from senescent cells that codes for an inhibitor of DNA synthesis). Both genes were overexpressed in cells during crisis, as is the case with senescent cells. The results are discussed in terms of the two-stage model previously proposed to explain the process of immortalization of human diploid fibroblasts by SV40.
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Roy M. and Phyllis Gough, Huffington Center on Aging, Baylor College of Medicine, Houston, Texas 77030.
Pereira-Smith
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D000952:Antigens, Polyomavirus Transforming / Q000032:analysis; D001345:Autoradiography; D002453:Cell Cycle / Q000502:physiology; D002461:Cell Line, Transformed; D016922:Cellular Senescence; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D004247:DNA / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D015536:Down-Regulation; D005347:Fibroblasts / Q000737:chemistry; D005353:Fibronectins / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D005434:Flow Cytometry; D006801:Humans; D008168:Lung / Q000737:chemistry / Q000166:cytology* / Q000502:physiology; D012313:RNA / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D013539:Simian virus 40 / Q000502:physiology*; D013936:Thymidine / Q000378:metabolism; D014162:Transfection; D014316:Tritium
D000952:Antigens, Polyomavirus Transforming; C493956:CDKN1A protein, human; D050759:Cyclin-Dependent Kinase Inhibitor p21; D016213:Cyclins; D005353:Fibronectins; D014316:Tritium; D012313:RNA; D004247:DNA; D013936:Thymidine