The role of mRNA stability and transcription in O6-methylguanine DNA methyltransferase (MGMT) expression in Mer+ human tumor cells.

We have recently reported that following depletion of O6-methylguanine DNA methyltransferase (MGMT) activity by acute streptozotocin (STZ) treatment to sensitize innately chloroethylnitrosourea (CENU)-resistant HT-29 cells, the eventual repletion of activity occurs with no concommitant alterations in steady-state MGMT mRNA levels. This suggestion of a potentially stable transcript prompted studies to define the relative contributions of MGMT mRNA stability and transcription to cellular MGMT expression. Northern analysis of MGMT mRNA in actinomycin D-treated HT-29, MR-1 and A2182 cells, ranging in relative MGMT expression from high to low respectively, demonstrates relatively long MGMT mRNA half-lives of > 10-12 h. Cell lines with low and moderate levels of MGMT mRNA appear to have longer mRNA half-lives than those with high levels. Run-on transcription in nuclei isolated from cells with low to moderate MGMT mRNA levels demonstrates undetectable basal MGMT transcription rates. Collectively these data suggest that a very low transcription rate, coupled with a stable mRNA molecule, might result in the translation of pre-existing mRNA molecules. This translation may be responsible for the gradual recovery of MGMT and CENU resistance over 24 h following MGMT depletion.