DIFFERENTIAL REGULATION OF CYCLIN-A, CYCLIN-B AND P21 CONCENTRATIONS IN A GROWTH-RESTRICTED HUMAN FIBROBLAST CELL-LINE

When the culture temperature was shifted from 35 degrees C to 39 degre es C, human fibroblasts immortalized by the temperature-sensitive simi an virus 40 T antigen became larger and acquired the morphological cha racteristics of senescent fibroblasts. After culture at 39 degrees C f or 48 h, most cells had ceased to proliferate. A rapid depletion of ce lls with S-phase DNA content was observed after the temperature shift. To elucidate the mechanism governing this rapid arrest of proliferati on, we studied the expression of genes involved in the regulation of c ell cycle progression. Cyclin A, cyclin B and p34(cdc2) concentrations were not changed during growth restriction, whereas p21 was rapidly i nduced in these growth-restricted cells. Transient expression of exoge nous p21 in cells cultured at 35 degrees C led to growth restriction a nd morphological changes characteristic of senescence. Furthermore, we studied the reversibility of growth restriction induced by the temper ature increase. The results showed that senescent morphology and growt h arrest were not reversible. In these cells the p21 concentration rem ained high and p34(cdc2) remained undetectable. This indicates that p2 1 accumulation might be responsible for the maintenance of senescence. Our findings provide information on the use of growth restriction of immortalized fibroblasts induced by a temperature shift as a model sys tem to study senescence.