RRC ID 38722
Author Yokoyama K, Yasumoto K, Suzuki H, Shibahara S.
Title Cloning of the human DOPAchrome tautomerase/tyrosinase-related protein 2 gene and identification of two regulatory regions required for its pigment cell-specific expression.
Journal J. Biol. Chem.
Abstract We have cloned and sequenced the human genomic DNA segments encoding the 5'-flanking region and the first two exons of the DOPAchrome tautomerase (DT)/tyrosinase-related protein 2 (TRP-2) gene. The DT gene is a member of the tyrosinase gene family and specifically expressed in melanin-producing cells. A transcriptional initiation site of the DT gene was identified by S1 nuclease-mapping and primer-extension analyses using RNA prepared from human pigmented melanoma cells. To study the mechanism for pigment cell-specific expression of the human DT gene, we analyzed the promoter function of its 5'-flanking region by transient expression assays. The fusion genes, containing the DT gene promoter upstream from a firefly luciferase reporter gene, were introduced into human pigmented melanoma cells and HeLa cells, and the pigment cell-specific promoter activity was evaluated by comparing the luciferase activity expressed in both cell lines. A series of 5' deletion studies of the human DT gene promoter revealed that the 32-bp element, located between -447 and -415, is sufficient to confer pigment cell-specific expression of a reporter gene on a homologous promoter, but not on a heterologous simian virus 40 promoter. Internal deletion studies using a homologous or a heterologous promoter revealed that the pigment cell-specific expression of a reporter gene mediated by the 32-bp element is dependent on the presence of another region of the DT gene spanning from -268 to -56, which was termed the proximal region. However, the proximal region by itself is not sufficient to confer cell type-specific expression. These results indicate that the presence of two regulatory regions, the 32-bp element and the proximal region, is required for pigment cell-specific expression of the DT gene. Both regulatory regions contain a CANNTG motif, a well known binding site for a large family of transcription factors possessing a basic helix-loop-helix structure.
Volume 269(43)
Pages 27080-7
Published 1994-10-28
PMID 7929451
MeSH Base Sequence Cloning, Molecular DNA Mutational Analysis Gene Expression Regulation* Genes, Reporter Humans Indolequinones* Indoles / metabolism Intramolecular Oxidoreductases* Isomerases / genetics* Melanoma, Experimental / metabolism* Molecular Sequence Data Monophenol Monooxygenase / genetics Multigene Family / genetics Promoter Regions, Genetic / genetics* Quinones / metabolism Restriction Mapping Sequence Analysis, DNA Tissue Distribution Transcription, Genetic Tumor Cells, Cultured
IF 4.011
Times Cited 89
WOS Category BIOCHEMISTRY & MOLECULAR BIOLOGY
Resource
Human and Animal Cells