Two molecular pathways (NMD and ERAD) contribute to a genetic epilepsy associated with the GABA(A) receptor GABRA1 PTC mutation, 975delC, S326fs328X.

Approximately one-third of human genetic diseases are caused by premature translation-termination codon (PTC)-generating mutations. These mutations in sodium channel and GABA(A) receptor genes have been associated with idiopathic generalized epilepsies, but the cellular consequences of the PTCs on the mutant channel subunit biogenesis and function are unknown. The PTCs could result in translation of a truncated subunit, or more likely, trigger mRNA degradation through nonsense-mediated mRNA decay (NMD), thus preventing or reducing production of mutant subunit at the transcriptional level. The GABA(A) receptor alpha1 subunit mutation, 975delC, S326fs328X, is an autosomal dominant mutation associated with childhood absence epilepsy that generates a PTC in exon 8 of the 9 exon GABRA1 gene that is 74 bp upstream of intron 8. Using an intron 8-inclusion minigene that supports NMD, we demonstrated that mutant mRNA was substantially reduced, but not absent. Loss of mutant transcripts was blocked by ribosome inhibition or by silencing the NMD-essential gene hUPF-1. In both neurons and non-neuronal cells, the PTC caused substantial loss of mutant alpha1(S326fs328X) subunit mRNA through NMD with a minor portion of the mRNA escaping NMD and producing a mutant protein. The translated mutant protein had reduced stability due to endoplasmic reticulum associated degradation (ERAD) and had enhanced association with molecular chaperones. This study suggests that loss of mRNA due to activation of NMD and activation of ERAD by the mutant protein may contribute to epileptogenesis. The molecular mechanisms outlined here delineate a model for the pathogenesis of many PTC-generating mutations.