A 413 bp region upstream the human 5-HT1A receptor gene is sufficient for its in vitro expression

Abstract

The regulation of a specific pattern of neuronal 5-hydroxytryptamine-1A (5-HT1A) receptor gene expression in the brain is hardly understood. Identification of cis-acting DNA sequences that control expression of the human 5-HT1A receptor gene has been undertaken by fusion of a 413 base pairs (bp)-long DNA segment upstream the human 5-HT1A receptor gene start codon to the 5-HT1A receptor gene. When this fusion product was inserted into the expression vector pcDNA3 and transfected into Cos-7 cells, it was shown to drive expression of specific [3H] (+)-8-hydroxy-2-(di-n-propylamino)tetralin ([3H] 8-OH-DPAT) binding sites with a Kd of 1.18 nM and a Bmax of 105 fmol/mg protein. The yield of expression of [3H] 8-OH-DPAT binding sites by fusion of the 5-HT1A receptor gene to strong promoters, such as the Rous sarcoma virus long terminal repeat and the cytomegalovirus promoter, was 2.5 to 8 times greater. The pharmacological binding profile of these [3H] 8-OH-DPAT binding sites was similar to that of cloned human 5-HT1A receptors stably expressed in HeLa cells. The importance of an Initiator sequence and regulatory Spl sites in the 413 bp region to direct transcription of the human 5-HT1A receptor gene is discussed.