Cloning and cell type-specific regulation of the human tyrosine hydroxylase gene promoter.
Tyrosine hydroxylase (TH), the rate-limiting enzyme of catecholamine biosynthesis, is predominantly expressed in several cell groups within the brain, including the dopaminergic (DA) neurons of the substantia nigra and ventral tegmental area, and the noradrenergic neurons of the locus coeruleus. To investigate the regulation of cell type-specific TH expression, we cloned and sequenced a 5.5kb fragment of human genomic DNA immediately 5(') of the TH coding region. This 5(')-flanking region does not contain either a CAAT box or a GC-rich region, but does contain a TATA box and consensus binding sequences for basal (TATA and CRE), and DA neuron-specific (NBRE, Gli, and BBE) transcription factors. Sequence analysis showed low overall homology with the rat and mouse TH promoter regions, with the exception of two high-homology domains, which encompassed -2384 to -2323 and -123 to -65, respectively. Interestingly, these distal and proximal domains contained NBRE, BBE, CRE, and TATA boxes, which are known to play important roles in DA neurogenesis. To further localize the TH promoter region responsible for transcriptional activity, we fused a 3301-bp human TH promoter fragment (-3174 to +127) to a luciferase reporter gene, and used this to assess promoter activity in neuronal and non-neuronal cell lines. Consistent with endogenous TH expression, this promoter construct was active in SH-SY5Y human neuroblastoma cells but not F3 human neural stem cells (NSCs). Deletion analysis of TH promoter/luciferase constructs revealed the presence of the repressor element in -1232 to -1210 upstream of transcription initiation site. While this region repressed 85% of promoter activity when transfected into F3 cells, it was not active in SH-SY5Y cells. These data suggest that the repressor element may play an important role in neuron cell-specific expression of the TH gene. Our results may provide insight into neuronal cell-specific expression of the human TH gene and allow a better understanding of catecholaminergic neuron disorders such as Parkinson's disease and schizophrenia.
Kim
Tae Eun
TE
Brain Disease Research Center, School of Medicine, Ajou University, Suwon, South Korea.
Park
Mi Jung
MJ
Choi
Eun Jung
EJ
Lee
Hack Sup
HS
Lee
Sung-Ho
SH
Yoon
Soo Han
SH
Oh
Chang-Kwon
CK
Lee
Byung Ju
BJ
Kim
Seung U
SU
Lee
Young Seek
YS
Lee
Myung Ae
MA
D000328:Adult; D000818:Animals; D001921:Brain / Q000378:metabolism*; D002460:Cell Line; D045744:Cell Line, Tumor; D003001:Cloning, Molecular / Q000379:methods*; D005260:Female; D015971:Gene Expression Regulation, Enzymologic / Q000235:genetics*; D006801:Humans; D051379:Mice; D009474:Neurons / Q000378:metabolism*; D009928:Organ Specificity; D011401:Promoter Regions, Genetic / Q000235:genetics; D051381:Rats; D016415:Sequence Alignment; D020539:Sequence Analysis, Protein; D017386:Sequence Homology, Amino Acid; D013045:Species Specificity; D013329:Structure-Activity Relationship; D014446:Tyrosine 3-Monooxygenase / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*
D014446:Tyrosine 3-Monooxygenase