Characterization of the 5 '-flanking promoter region of the rat somatostatin receptor subtype 3 gene

We investigated the 5'-flanking promoter region of the rat somatostatin rec eptor subtype 3 (rSSTR3), Using a cDNA probe, genomic clones containing the 5'-flanking promoter region of the rSSTR3 gene were isolated. A sequence o f 5.4 kb directly upstream from the start codon mas analyzed and two intron s mere found in the 5' untranslated region (UTR) of the cDNA sequence, The transcriptional initiation site was determined by 5' rapid amplification of cDNA ends (RACE), primer extension and RNase protection analysis with cere bellar RNA. Two major transcriptional initiation sites were found at positi on -1040 (tsp1) and -856 (tsp2) relative to the translational initiation si te. Like a number of other promoters of G-protein-coupled receptors, the rS STR3 gene lacks TATA and CAAT motifs and includes G+C-rich regions. Functio nal analysis of the promoter region by transfecting rSSTR3 luciferase-repor ter gene constructs into rat pituitary GH(3) cells and HEK 293 cells indica ted that a 107-bp region upstream of tsp2 was sufficient to drive transcrip tion. Furthermore a 562-bp region at position -1304 to -1865 upstream of th e ATG start codon exerted a negative regulatory effect on transcriptional a ctivity. (C) 1998 Federation of European Biochemical Societies.