Functional characterization of the murine serotonin transporter gene promoter in serotonergic raphe neurons.
We have isolated and characterized the 5'-flanking regulatory region of the murine serotonin 5-HT transporter (5-HTT) gene. A TATA-like motif and several potential binding sites for transcription factors, including two AP1, several AP2 and AP4 binding sites, CCAAT and GC boxes (SP1 binding sites), a nuclear factor-kappaB, and a cyclic AMP response element-like motif, are present in the 5'-flanking region. A approximately 2.2-kb fragment (-2,143 to +51 with respect to the transcription start site), which had been fused to the luciferase reporter gene and transiently expressed in a 5-HTT-expressing cell line and in serotonergic raphe neurons derived from embryonic rat brainstem, displayed both constitutive and inducible promoter activity. Functional promoter mapping revealed two clusters of activating elements from bp -82 to -527 and 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 approximately 0.5 kb of the 5'-flanking sequence is sufficient to confer its cell-selective expression, (3) the promoter responds to cyclic AMP- and protein kinase C-dependent induction, and (4) the expression of the 5-HTT is regulated 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 its cell-selective expression in transgenic strategies.
Heils
A
A
Department of Psychiatry, University of Würzburg, Germany.
Wichems
C
C
Mössner
R
R
Petri
S
S
Glatz
K
K
Bengel
D
D
Murphy
D L
DL
Lesch
K P
KP
D000818:Animals; D001483:Base Sequence; D002273:Carcinogens / Q000494:pharmacology; D002352:Carrier Proteins / Q000235:genetics*; D002540:Cerebral Cortex / Q000737:chemistry / Q000166:cytology; D002822:Choriocarcinoma; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000378:metabolism; D005260:Female; D005333:Fetus / Q000737:chemistry; D017353:Gene Deletion; D018507:Gene Expression Regulation, Developmental / Q000187:drug effects; D006801:Humans; D008562:Membrane Glycoproteins / Q000235:genetics*; D026901:Membrane Transport Proteins; D051379:Mice; D008969:Molecular Sequence Data; D016296:Mutagenesis; D009419:Nerve Tissue Proteins / Q000235:genetics; D009474:Neurons / Q000737:chemistry* / Q000201:enzymology; D010751:Phosphopyruvate Hydratase / Q000032:analysis; D011247:Pregnancy; D011493:Protein Kinase C / Q000378:metabolism; D011903:Raphe Nuclei / Q000737:chemistry / Q000166:cytology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D012701:Serotonin / Q000032:analysis / Q000502:physiology; D050486:Serotonin Plasma Membrane Transport Proteins; D016385:TATA Box / Q000235:genetics*; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014407:Tumor Cells, Cultured / Q000737:chemistry / Q000502:physiology
D002273:Carcinogens; D002352:Carrier Proteins; D008562:Membrane Glycoproteins; D026901:Membrane Transport Proteins; D009419:Nerve Tissue Proteins; C493230:SLC6A4 protein, human; D050486:Serotonin Plasma Membrane Transport Proteins; C493231:Slc6a4 protein, mouse; C493232:Slc6a4 protein, rat; D005576:Colforsin; D012701:Serotonin; D000242:Cyclic AMP; D011493:Protein Kinase C; D010751:Phosphopyruvate Hydratase; D013755:Tetradecanoylphorbol Acetate