Abstract:Dopamine β‐hydroxylase (DBH) catalyzes the synthesis of the neurohormone norepinephrine and is expressed only in noradrenergic and adrenergic cells of the nervous and endocrine systems. We have previously described a positive‐acting genetic regulatory element of the DBH that contributes to the restricted expression of this gene. In the study described here, we identify and characterize a negative‐acting regulatory element within the 5′‐flanking region of the rat DBH gene. This negative regulatory element, which is located between −282 and −232, represses transcription from the DBH promoter in cell lines of noncatecholaminergic origin and binds to nuclear factors found in extracts from both the catecholaminergic cell line SHSY‐5Y and the noncatecholaminergic cell lines JEG‐3 and C6. The negative regulatory region will reduce transcription from a heterologous promoter in two noncatecholaminergic cell lines. These experiments demonstrate that the selective expression of the DBH gene is controlled by both positive‐ and negative‐acting genet