FIGURE 1.

Effect of heterozygous α1(AD) subunit expression on surface and total α1 subunit expression. We transfected HEK293T cells with empty plasmid (negative control) or 0.250 μg of β2 and γ2 subunit cDNA and either 0.250 μg of wild type α1 (wt), 0.125 μg α1 (hemizygous (hemi)), 0.125 μg each of α1 and α1(AD) (heterozygous (het)), or 0.250 μg of α1(AD) (homozygous (hom)) cDNA. We biotinylated surface proteins and performed Western blots to determine the surface (A and C) and total (B and D) GABA_AR α1 subunit expression relative to that of the ATPase α1 subunit (loading control). Surface and total hemizygous and heterozygous α1 subunit expression was greater than that of homozygous expression and was significantly reduced relative to wild type expression (*), but there was no significant (ns) difference between them (n = 5). We also performed flow cytometry assays (E–H) as a second method to quantify the surface and total α1 subunit for cells transfected with negative control (filled histogram), wild type (solid line), hemizygous (dotted line), heterozygous (dashed line), or homozygous (data not shown) GABA_AR. There was no significant difference between hemizygous and heterozygous receptors in surface α1 subunit expression (n = 7), but total heterozygous α1 subunit expression was larger than that of hemizygous α1 subunit expression (n = 6, p = 0.03).