The major class of inhibitory neurotransmitter receptors, the GABAA receptors, are modulated by a range of depressant drugs (eg benzodiazepines, ethanol) and their dysfunction has been associated with several neurological disorders. Recently, cDNA cloning has revealed the existence of numerous receptor subunit types. However, in the absence of genomic clones, it has been difficult to establish any link between inappropriate receptor expression and the implicated disease states. In addition, nothing is known of how expression of subunit genes is regulated in a coordinated and localized manner. Studies on the intron/exon structure, DNA sequence and chromosomal localization of the gene encoding the human beta2 subunit have been completed. Repetitive DNA sequences within intronic segments of the gene were used to detect a polymorphic region of the gene. This polymorphism can now be used to screen for any linkage between the beta1 gene and known genetic disorders. These types of studies are being extended to include other GABA receptor genes, in particular those encoding the alpha6 and gamma2 subunits. We have obtained six independent putative gamma2 clones from a human genomic library in our initial screen. We will map the positions of restriction sites and gamma2 introns/exons on these clones. Polymorphisms will be identified by probing these clones with oligonucleotides composed of known polymorphic repeated sequences and by restriction fragment length mapping of human genomic DNA. We will also determine the chromosomal position of these subunit genes by in situ hybridization. Cloned fragments of the human beta3 subunit gene were used to map this gene to the Angelman/Prader-Willi region of human chromosome 15. It is possible that this gene may play a role in the pathogenesis of one or both of these syndromes. As the syndromes appear to result from a genetic imprinting process, an understanding of how expression of the beta3 gene is regulated may be of particular importance. Ongoing studies aim to define the regulatory elements of the beta3 gene using transfected PC12 cells and PC12 nuclear extracts.
