Human renin mRNA stability is increased in response to cAMP in Calu-6 cells

The human carcinoma-derived cell line Calu-6 has previously been demonstrat ed to endogenously express human renin (hREN) mRNA and to markedly increase steady-state hREN mRNA levels (100-fold after 24 hours) in response to ana logues of cAMP and postreceptor activators of adenylyl cyclase such as fors kolin. However, both transfection analysis using hREN promoter-reporter con structs and nuclear run-on experiments suggest that transcriptional activit y alone cannot account for this level of induction. We performed primer ext ension, reverse transcription-polymerase chain reaction, and 3' rapid ampli fication of cDNA ends to compare hREN mRNA between unstimulated and forskol in-stimulated cells. We demonstrate that hREN mRNA is identical under both conditions with respect to (1) utilization of the appropriate transcription start site, (2) processing of renin mRNA, and (3) utilization of the prope r polyadenylation site and length of the poly-A tail. To address the mechan ism of induction caused by cAMP, we used transcriptional inhibition and mea sured decay of hREN mRNA before and after forskolin or phorbol ester treatm ent. Experiments with both actinomycin D and 5,6-dichlororibofuranosylbenzi midnzole (DRB) showed that forskolin treatment markedly stabilized hREN mRN A in Calu-6 cells. A 2.3-fold increase in hREN mRNA half-life was also obse rved after treatment of Calu-6 cells with phorbol ester, Experiments with D RB demonstrated a similar robust stabilization of hREN mRNA after forskolin and phorbol ester treatment. These data demonstrate that the induction in hREN mRNA in response to both cAMP and phorbol ester occurs by a mechanism involving a posttranscriptional component.