The promoter regions of human choline acetyltransferase (ChAT) and aromatic L-amino acid decarboxylase (AADC) genes have been analyzed by transient transfection assays. AADC gene is transcribed from two alternative noncoding first exons, 1N and 1NN, expressed in pheochomocytoma and hepatoma cells, respectively. 5' flanking sequences of exon 1 N (from 9000 to 147 bp) display promoter activity in SK-N-BE neuroblastoma cells, but not in MC-I-XC cholinergic neuroepithelioma cells, and in AADC-rich non-neuronal cells. On the contrary, 5' flanking sequences of exon 1 NN (from 1117 to 119 bp) display high promoter activity in human hepatoma cells HepG2, but not in SK-N-BE cells, suggesting high degrees of specificity of promoters N and NN for AADC-expressing neuronal and non-neuronal cells, respectively. Preliminary evidence suggests that leukemia inhibitory factor suppresses the activity of the neuronal promoter in cultured sympathetic neurons. Two alternative first exons, R and M, have been localized in human ChAT gene, and the corresponding promoters characterized in cholinergic PC12 and NG-108-15 cells, and in non-cholinergic neuro2A cells. Several positively or negatively acting cis elements have been localized in the two promoters, as well as a cAMP-inducible, enhancer-like element in the second intron. Among the various cell lines studied, there was no correlation between promoter activities and the expression of the endogenous ChAT gene, suggesting that the fine-tuning of ChAT gene expression is controlled by silencer elements which remain to be localized.