Organization and expression of the Rat D2A receptor gene: identification of alternative transcripts and a variant donor splice site

Abstract

We have recently reported the creation of a cell line expressing D2 receptors encoded by a gene distinct from that described by Bunzow et al. [Bunzow, J. R., Van Tol, H. H. M., Grandy, D. K., Albert, P., Salon, J., Christie, M., Machida, C. A., Neve, K., & Civelli, O. (1988) Nature 336, 783-787]. To provide a framework for understanding structural differences between these and other G-protein-coupled receptors, the structure of the rat gene coding for the Bunzow et al. cDNA (called D2A here) was delineated. The D2A gene contains eight exons and spans at least 50 kb. Sets of oligonucleotide primers were used in combination with the polymerase chain reaction (PCR) to determine the presence of alternative transcripts within the introns. In contrast to other G-protein-coupled receptors, the D2A gene undergoes alternative RNA processing within intron 5, resulting in an insertion of 29 amino acids to the predicted 415 amino acid sequence of the D2A protein. By use of the PCR assay the relative abundance and tissue distribution of the alternative D2A transcripts (herein termed D2A415 and D2A444) were determined. A variant donor splice site was also identified at the end of exon 4, a GC dinucleotide instead of the canonical GT. The variant dinucleotide was also present in the mouse but not in the human D2A gene.