The GABRG2
nonsense mutation, Q40X, is associated with the severe
epilepsy syndrome,
Dravet syndrome, and is predicted to generate a premature translation-
termination codon (PTC) in the
GABA(A) receptor γ2 subunit
mRNA in a position that codes for the first
amino acid of the mutant subunit. We determined the effects of the mutation on γ2 subunit
mRNA and
protein synthesis and degradation, as well as on α1β2γ2
GABA(A) receptor assembly, trafficking and surface expression in HEK cells. Using bacterial artificial chromosome (BAC) constructs, we found that γ2(Q40X) subunit
mRNA was degraded by nonsense mediated mRNA decay (NMD). Undegraded mutant
mRNA was translated to a truncated
peptide, likely the
signal peptide, which was cleaved further. We also found that mutant γ2(Q40X) subunits did not assemble into functional receptors, thus decreasing
GABA-evoked current amplitudes. The GABRG2(Q40X) mutation is one of several
epilepsy-associated
nonsense mutations that have the potential to be rescued by reading through the PTC, thus restoring full-length protein translation. As a first approach, we investigated the use of the
aminoglycoside,
gentamicin, to rescue translation of intact mutant subunits by inducing
mRNA read-through. In the presence of
gentamicin, synthesis of full length γ2 subunits was partially restored, and surface biotinylation and whole cell recording experiments suggested that rescued γ2 subunits could corporate into functional, surface
GABA(A) receptors, indicating a possible direction for future
therapy.