Identification of essential cis-acting regulatory elements for transcription of the rat DAN gene.
The DAN gene was initially isolated as one of the genes whose expression is significantly decreased in a variety of transformed rat fibroblasts 3Y1 cells when compared with the parental 3Y1 cells. In the present study, we have isolated the genomic clone of the DAN gene from a 3Y1 genomic library and characterized the possible regulatory elements responsible for the transcription of the DAN gene. The transcription initiation site was determined by a primer extension experiment. Putative TATA and CAAT-like elements were present 31 and 358 bp upstream from the transcription start site, respectively. Transient transfection of a series of DAN-chloramphenicol acetyltransferase (CAT) gene constructs, which contain different portions of the 5'-flanking region (2,236 bp) of the DAN gene and the CAT gene, was used to localize a regulatory element. These experiments demonstrated the presence of the regions that regulate DAN gene expression positively (-57 to +118) and negatively (-1,232 to -636). The electrophoretic mobility-shift assays revealed that 3Y1 and SR-3Y1 nuclear extracts specifically interact with the positive (-57 to +118) and the negative (-1,226 to -987) regulatory regions, respectively.
Ozaki
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Division of Biochemistry, Chiba Cancer Center Research Institute, Chuoh-ku, Japan.
Sakiyama
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S
D000818:Animals; D001483:Base Sequence; D002460:Cell Line; D015500:Chloramphenicol O-Acetyltransferase / Q000235:genetics; D018076:DNA, Complementary; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins; D011506:Proteins / Q000235:genetics*; D051381:Rats; D012045:Regulatory Sequences, Nucleic Acid; D014158:Transcription, Genetic
D018076:DNA, Complementary; C085763:Nbl1 protein, rat; D009419:Nerve Tissue Proteins; D011506:Proteins; D015500:Chloramphenicol O-Acetyltransferase