Two key enzymes involved in the synthesis of melatonin, hydroxyindole-O-methyltransferase (HIOMT) and arylalkylamine N-acetyltransferase (AA-NAT), are present in Y79 human retinoblastoma cells. Under certain conditions these cells produce melatonin and secrete it into the culture medium. In a previous study, it was observed that melatonin levels increase dramatically over control levels after the addition of dibutyryl cyclic AMP (dbcAMP), whereas after treatment with butyrate melatonin levels decreased. The changes in melatonin levels appeared to be the result of increases in AA-NAT activity or decreases in HIOMT activity, following dbcAMP or butyrate treatment. In this study, mechanisms by which these agents influence HIOMT and AA-NAT gene expression were examined. Levels of AA-NAT and HIOMT RNA expression in response to treatment of Y79 cultures with 4 mM dbcAMP or 2 mM butyrate were measured by semi-quantitative reverse-transcription/polymerase chain reaction. Butyrate and dbcAMP showed no effect on AA-NAT gene expression, whereas HIOMT gene expression was reduced by treatment with these agents. Levels of beta-actin RNA were increased following dbcAMP or butyrate treatment. This analysis suggests that the reduction in HIOMT activity caused by dbcAMP or butyrate treatment is the result of a decrease in HIOMT RNA synthesis or accumulation. Conversely, since AA-NAT RNA levels were unaffected by dbcAMP or butyrate treatment, the increase in AA-NAT activity previously observed may be the result of changes in the activational state of the AA-NAT protein.