Approximately 3% of people develop epilepsy at some point during their life and one third of these patients will not have their seizures controlled with antiepileptic medication. Our long term goal is to elucidate mechanisms of genetic epilepsy in an effort to help develop more effective antiepileptic therapies. Juvenile myoclonic epilepsy is one of the most common epilepsy syndromes and it accounts for approximately 5% of all cases of epilepsy. The non-conserved, missense mutation A322D in the GABA-A receptor alpha 1 subunit gene (GABRA1) is associated with an autosomal dominant form of juvenile myoclonic epilepsy (ADJME). Recently, we demonstrated that in a mammalian, non-neuronal expression system, the ADJME mutation reduces alpha 1 subunit expression after protein translation, but before subunit oligomerization, data which suggested that the mutation causes enhanced endoplasmic reticulum associated degradation (ERAD) of the alpha 1 subunit. ERAD is a complex process that utilizes numerous chaperone proteins and thus it is likely that ERAD in neurons would differ from that in fibroblast cell lines. Although neuronal ERAD deficiencies have been studied in relation to some neurodegenerative diseases, its role as a normal response to misfolded mutated proteins is unknown. We propose to elucidate the role of normal ERAD of native alpha 1, recombinant wild type, and mutant alpha 1(A322D) subunits in both a neuronal cell line (HT-22) and in cultured cortical neurons. We propose three Specific Aims. 1) We will determine the effect of the ADJME alpha 1 subunit mutation on alpha 1 subunit biosynthesis and degradation and its association with the ubiquitin proteasome system in both fibroblasts and neurons. 2) We will characterize the effect of this mutation on the GABA-A receptor physiology in neurons. 3) We will characterize neurons'compensatory response to degradation of the alpha 1 subunit resulting from the ADJME mutation;specifically, we will study neurons'alteration of expression of other alpha subunits and the resulting changes in postsynaptic GABA physiology.
