FACTORS INFLUENCING THE EXPRESSION OF ENDOGENOUS RETROVIRUS-LIKE SEQUENCES IN RAT-6 CELLS

Previous studies from this laboratory have demonstrated increased levels of expression of endogenous rat leukemia virus (RaLV) and 30S retrovirus-like sequences in liver and colon tumors induced by chemical carcinogens in rats. During the process of normal liver regeneration, the levels of RaLV RNA were dramatically increased, whereas the levels of 30S RNA did not change. The present study examined several factors that might influence the expression of these sequences in the Rat 6 embryo fibroblast cell line. Rat 6 cells in either log-phase or confluent cultures were treated with cycloheximide, the tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA), or the activated carcinogen benzo[a]pyrene diol epoxide (BPDE) for various periods of time up to 48 h. Northern blot analyses of total RNA indicated that cells in untreated cultures in log phase had higher levels of RaLV and 30S RNAs than did confluent cells. Within 10 h cycloheximide (2 or 10-mu-g/mL) markedly increased the levels of RaLV and 30S RNAs in both log-phase and confluent cells. BPDE (100 ng/mL) induced a marked increase in the levels of RaLV RNA at 4 to 10 h, which returned to the basal level by 24 h in the log-phase cells; but no significant change was seen in the confluent cells. The level of 30S RNA also increased moderately in the BPDE-treated log-phase cells and was maximal at 24 h; but no change was seen in confluent cells. Treatment with TPA (100 ng/mL) induced no significant increase in either RaLV or 30S RNA levels in the log-phase or confluent cells. The exposure of Rat 6 cells to 5-azacytidine (3-mu-M for 24 h) led to a marked increase in the levels of both RaLV and 30S RNAs, which persisted during at least 15 subsequent passages in the absence of the drug. Thus, inhibition of protein synthesis, DNA damage, or hypomethylation can increase the expression of certain endogenous retrovirus-like sequences, but an activator of protein kinase C does not.