STRUCTURE OF THE HUMAN ALPHA(2) SUBUNIT GENE OF THE GLYCINE RECEPTOR - USE OF VECTORETTE AND ALU-EXON PCR

The alpha subunit of the glycine receptor is encoded by multiple genes that display developmental and tissue-specific expression. The alpha( 1) subunit gene is expressed predominantly in the adult brain stem and spinal cord, whereas the alpha(1) subunit gene is expressed in fetal brain and spinal cord. We wished to determine the genomic organization of the human alpha(2) subunit gene as well as to define the 5' ends o f the alpha(1), and alpha(2) subunit genes. Gene structure can be defi ned rapidly from yeast artificial chromosome (YAC) DNA sources by the use of vectorette-exon polymerase chain reaction (PCR). However, YACs frequently contain small deletions that complicate the determination o f the complete exon-intron structure of a gene, and this often necessi tates the isolation of additional clones. In this study we have used v ectorette-exon PCR from YAC DNA to define exons of the glycine recepto r alpha(2) subunit gene. To define those exons that were absent in the isolated YACs, we used Alu-exon PCR on genomic DNA, using nested prim ers to obtain specificity in the PCR reactions. The alpha(2) subunit g ene was found to contain nine exons varying in size from 68 bp (exons 3A and 3B) to 581 bp (exon 1). All of the intron-exon boundary sequenc es conform to consensus splice donor and acceptor sites. In addition, we have defined the 5' end of this gene as well as that of the alpha(1 ) subunit gene by RACE-PCR. The structures of the alpha subunit glycin e receptor genes in humans are very similar to each other and to the a lpha subunit genes in mice.