Novel Transcription Start Site for Murine AChE

Abstract

Evidence is accumulating to suggest more than one functional promoter in the gene for murine acetylcholinesterase (Ache). Previously, we detected moderately strong activity (12.5 times background) when a DNA sequence upstream of the established promoter was coupled to a luciferase reporter in neuroblastoma cells. When this sequence and the distal promoter were joined in the same reporter construct, activity was six times greater than with either alone. Elements from the upstream sequence have now been demonstrated in freshly prepared cDNA from mouse brain and in a commercial cDNA preparation designed for RACE (Rapid Amplification of cDNA Ends). With both templates, PCR reactions using appropriate primers amplified a transcript whose 5’ end began at position −626 relative to the translation start codon. Strikingly, this cDNA sequence matched that of murine Ache down through position −335 but, beginning with the next nucleotide, switched to the 5’ end of exon 2 (position −22). Thus, the putative alternative promoter appears to drive transcription in a manner that deletes exon 1, substituting a new exon 1a (as in Torpedo Ache). It is worth noting that the nucleotide sequence of exon 1a contains two ATG codons upstream of the open reading frame. However, a stop codon occurs within thirty nucleotides downstream of each potential translation start site. Therefore, exon 1a is unlikely to participate in protein coding or yield variant amino acid sequences.