5'-Flanking sequence and promoter activity of the rabbit neuronal nitric oxide synthase (nNOS) gene.
We have isolated a rabbit neuronal nitric oxide synthase (nNOS) cDNA encoding a protein of 1,435 amino acids. Using the cDNA clones as probes, the 5'-flanking region of the nNOS gene was isolated from a rabbit genomic DNA library. 5'RACE and primer extension analysis of rabbit brain total RNA mapped multiple transcription initiation sites localized 474-487 bp upstream from the translation start codon. Analysis of 5,197 bp of the 5'-flanking sequence revealed that the rabbit nNOS gene promoter lacks canonical TATA or CCAAT boxes and, instead, contains a GC-rich region and multiple Sp1 sites. Farther from the +1start, various putative cis-elements including AP-1, AP-4, NF-kappaB, STAT, CREB, C/EBP and c-Myc were observed. The functional promoter activity of the 5'-flanking region was demonstrated by its ability to drive the expression of a beta-galactosidase reporter gene in several cell types. Serial deletion analysis of the promoter region revealed that the -291 to -172 region, which contains two Sp1 sites, is essential for basal transcriptional activity. These results suggest that the rabbit nNOS promoter contains characteristics of inducible genes.
Jeong
Y
Y
National Creative Research Initiative Center for Genetic Reprogramming, Institute for Molecular Biology and Genetics, School of Biological Sciences, Seoul National University, Korea.
Won
J
J
Kim
C
C
Yim
J
J
D020121:5' Untranslated Regions / Q000235:genetics*; D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D001921:Brain / Q000201:enzymology*; D003001:Cloning, Molecular; D003062:Codon / Q000235:genetics; D018076:DNA, Complementary; D004268:DNA-Binding Proteins / Q000378:metabolism; D008969:Molecular Sequence Data; D009419:Nerve Tissue Proteins / Q000235:genetics; D019001:Nitric Oxide Synthase / Q000235:genetics*; D052248:Nitric Oxide Synthase Type I; D011401:Promoter Regions, Genetic; D011817:Rabbits; D014158:Transcription, Genetic
D020121:5' Untranslated Regions; D003062:Codon; D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D009419:Nerve Tissue Proteins; D019001:Nitric Oxide Synthase; D052248:Nitric Oxide Synthase Type I