FUNCTIONAL-CHARACTERIZATION OF THE MURINE SEROTONIN TRANSPORTER GENE PROMOTER IN SEROTONERGIC RAPHE NEURONS

We have isolated and characterized the 5'-flanking regulatory region o f the murine serotonin 5-HT transporter (5-HTT) gene. A TATA-like moti f and several potential binding sites for transcription factors, inclu ding two AP1, several AP2 and AP4 binding sites, CCAAT and GC boxes (S P1 binding sites), a nuclear factor-kappa B, and a cyclic AMP response element-like motif, are present in the 5'-flanking region. A similar to 2.2-kb fragment (-2,143 to +51 with respect to the transcription st art site), which had been fused to the luciferase reporter gene and tr ansiently expressed in a 5-HTT-expressing cell line and in serotonergi c raphe neurons derived from embryonic rat brainstem, displayed both c onstitutive and inducible promoter activity. Functional promoter mappi ng revealed two clusters of activating elements from bp -82 to -527 an d bp -1,001 to -1,937. A cell/neuron-selective silencer element(s) is contained between bp -294 and -527. Our findings suggest that (1) the murine 5-HTT gene promoter is active in serotonergic raphe neurons but significantly repressed in neuronal cells from frontal cortex that do not express 5-HTT, (2) the information contained within similar to 0. 5 kb of the 5'-flanking sequence is sufficient to confer its cell-sele ctive expression, (3) the promoter responds to cyclic AMP-and protein kinase C-dependent induction, and (4) the expression of the 5-HTT is r egulated by a combination of positive and negative cis-acting elements operating through a basal promoter unit defined by a TATA-like motif. Fusion of the 5-HTT gene promoter unit to a gene of choice may aid it s cell-selective expression in transgenic strategies.