Nicotine-induced gene expression of proenkephalin in bovine chromaffin cells

The induction of the proenkephalin gene by nicotine has been characterized in bovine adrenal medullary chromaffin cells. Nicotine (10 microM) caused an approximately fourfold increase in the proenkephalin mRNA levels within 24 h. The half-life of the proenkephalin mRNA in nicotine-stimulated cells was similar to that in control cells (about 13 h), indicating that nicotine does not affect mRNA stability but acts at the levels of proenkephalin gene transcription. This was also supported by experiments showing that the expression of a proenkephalin chloramphenicol acetyl transferase reporter gene (PENKCAT-153/+50) containing 153 nucleotides of upstream promoter sequences is increased (about twofold) by nicotine after transient transfection in the chromaffin cells. In addition, nicotine induced a marked elevation of the immediate early gene mRNAs c-fos, c-jun, and jun-B. Maximally increased levels for c-fos mRNA (about 100-fold) were obtained after 20 min. c-jun and jun-B were increased three- to fivefold 60 min after nicotine addition. The expression of PENKCAT-153/+53 and of a proenkephalin gene reporter plasmid which contains a dimer of the enkephalin cAMP responsive element 2 (ENKCRE-2) in front of a minimal promoter was increased by cotransfection of a c-fos expression plasmid, indicating that nicotine may induce the proenkephalin gene in chromaffin cells via c-Fos which binds to the ENKCRE-2 element.