PROMOTER ANALYSIS OF THE RAT BETA(1)-ADRENERGIC RECEPTOR GENE IDENTIFIES SEQUENCES INVOLVED IN BASAL EXPRESSION

The beta(1)-adrenergic receptor (beta(1)-AR) mediates several function s of catecholamines in the heart, including the stimulation of heart r ate and contractility. The expression of the rat beta(1)-AR gene was a ssessed by transiently transfecting chimeric genes containing the beta (1)-AR promoter, driving the luciferase reporter gene into various cel l lines. beta(1)-AR/luciferase vectors containing 3 kb of the 5'-flank ing region and extending to -126 relative to the start site of transla tion were expressed at high levels in ventricular myocytes, SK-N-MC ce lls, and HepG2 cells. The addition of 26 nucleotides from -125 to -100 to the -3311 beta(1)-AR/luciferase chimeric gene reduced expression i n myocytes and SK-N-MC cells while eliminating expression in HepG2 cel ls. This element is located 125 base-pairs 3' to the transcriptional s tart site. The mutation of four nucleotides between -121 and -118 dimi nished the inhibitory effect of this element. The inhibitory activity of the -125 to -100 sequence was completely dependent on promoter cont ext and positioning. In addition to this 3' element, sequences between -3311 and -2740 in the 5'-flanking region of the beta(1)-AR gene were required for the full transcriptional suppression. Using DNase I foot printing and gel mobility assays, it was determined that within the 26 -bp region, rat heart nuclear proteins bound to two sites between nucl eotides -123 and -112 and -106 and -100. Therefore, appropriate basal expression of the beta(1)-AR gene involves widely separated sequences 3' and 5' to the transcriptional start site.