The epilepsy mutation, γ2(R43Q) disrupts a highly conserved inter-subunit contact site, perturbing the biogenesis of GABAA receptors

Hales, Timothy G.; Tang, Haiyan; Bollan, Karen A.; Johnson, Sara; King, Dale P.; McDonald, Neil; Cheng, Aixin; Connolly, Christopher N.

doi:10.1016/j.mcn.2005.01.002

Cited by 70 publications

(85 citation statements)

References 38 publications

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“…Furthermore, loss of ␥2(R43Q) could alter trafficking (39) such that density of receptors at synapses is reduced, causing the observed reduction in mIPSCs. In partial contrast to our in vivo finding, heterologous expression studies performed in HEK cells demonstrate both a reduction in ␥2 subunit expression and in fully formed heteromeric GABA A receptors at the cell surface, suggesting a dominant-negative effect of the R43Q mutation (16,(18)(19)(20). These studies highlight a need for caution when using the results from cell-based experiments to explain the genesis of complex diseases.…”

Section: ␥2(r43q) Cell Surface Reduction Is Common To Mouse and Mancontrasting

confidence: 96%

“…The reduced cortical mIPSCs described above could be explained by intracellular retention of receptors comprising the mutant ␥2(R43Q) subunit. This possibility has been raised in studies that used cell culture systems to heterologously express mutant subunits and assess function and membrane trafficking (16,19,20). To determine whether incorporation of the ␥2(R43Q) subunit causes retention of GABA A receptor complexes by a dominant-negative effect, we cultured primary neurons from C57BL/6 embryonic cortices (E15-E16) and biotin-labeled cell surface receptors.…”

Section: ␥2(r43q) Subunits Have Reduced Cell Surface Expression and Dmentioning

confidence: 99%

“…Epilepsy itself is a complex phenomenon involving the interaction of multiple cell types in networks within and between different brain regions that are likely to be influenced by GABA A receptor dysfunction caused by the R43Q mutation. Furthermore, in vitro analyses of the consequences of this mutation have shown inconsistent findings with a range of deficits in receptor pharmacology, trafficking, kinetics, or assembly (16)(17)(18)(19)(20)(21)(22)(23) potentially implicated in disease pathogenesis.…”

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confidence: 99%

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Reduced cortical inhibition in a mouse model of familial childhood absence epilepsy

Tan

Reid

Single³

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

201

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Mutations in the GABAA receptor ␥2 subunit are associated with childhood absence epilepsy and febrile seizures. To understand better the molecular basis of absence epilepsy in man, we developed a mouse model harboring a ␥2 subunit point mutation (R43Q) found in a large Australian family. Mice heterozygous for the mutation demonstrated behavioral arrest associated with 6-to 7-Hz spike-and-wave discharges, which are blocked by ethosuximide, a first-line treatment for absence epilepsy in man. Seizures in the mouse showed an abrupt onset at around age 20 days corresponding to the childhood nature of this disease. Reduced cell surface expression of ␥2(R43Q) was seen in heterozygous mice in the absence of any change in ␣1 subunit surface expression, ruling out a dominant-negative effect. GABA Amediated synaptic currents recorded from cortical pyramidal neurons revealed a small but significant reduction that was not seen in the reticular or ventrobasal thalamic nuclei. We hypothesize that a subtle reduction in cortical inhibition underlies childhood absence epilepsy seen in humans harboring the R43Q mutation.GABAA receptor ͉ genetics ͉ electroencephalogram ͉ trafficking ͉ synapse G ABA A receptors in the adult brain are important for inhibiting the activity of neurons in which they reside. Dysfunction of these receptors caused by familial mutations can give rise to febrile seizures (FS) and a variety of generalized epilepsy phenotypes (1-3). To date, five mutations have been reported in the GABA A ␥2 subunit gene with an array of seizure types seen in patients (1, 4-7). Childhood absence epilepsy (CAE) and FS were the main phenotypes in a large Australian family with an arginine to glutamine mutation at position 43 (R43Q) in the GABA A ␥2 subunit gene (1,8).Understanding how the GABA A ␥2(R43Q) mutation causes epilepsy is difficult. GABA A receptors themselves serve several roles. They regulate moment-to-moment brain function (9), play an important role in brain development (10), and have key roles in neuronal plasticity (11, 12) and response to brain injury (13-15). Epilepsy itself is a complex phenomenon involving the interaction of multiple cell types in networks within and between different brain regions that are likely to be influenced by GABA A receptor dysfunction caused by the R43Q mutation. Furthermore, in vitro analyses of the consequences of this mutation have shown inconsistent findings with a range of deficits in receptor pharmacology, trafficking, kinetics, or assembly (16-23) potentially implicated in disease pathogenesis.Clearly, the complex nature of epileptogenesis demands in vivo investigation. Genetic epilepsies provide a framework on which to investigate the consequences of causative mutations at a range of organizational levels within the brain, creating a chain of understanding from molecules to behavior. Linking this chain is impossible in humans because of the highly invasive methodology required and is severely limited in heterologous expression systems that lack necessary complexity. Mice mod...

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Section: ␥2(r43q) Cell Surface Reduction Is Common To Mouse and Mancontrasting

confidence: 96%

Section: ␥2(r43q) Subunits Have Reduced Cell Surface Expression and Dmentioning

confidence: 99%

mentioning

confidence: 99%

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Reduced cortical inhibition in a mouse model of familial childhood absence epilepsy

Tan

Reid

Single³

et al. 2007

Proc. Natl. Acad. Sci. U.S.A.

201

230

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“…Another ␥2 subunit mutation, ␥2(R43Q), reduced surface expression of GABA A receptors Sancar and Czajkowski, 2004;Hales et al, 2005), and like the ␥2(R43Q) mutation, both ␦(E177A) and ␦(R220H) variants are located in the subunit N terminus. The ␥2 subunit N terminus interacts with ␣ and ␤ subunits during receptor assembly (Klausberger et al, 2000;Jin et al, 2004), and the ␥2(R43Q) subunit mutation resulted in abnormal receptor assembly and trafficking Hales et al, 2005). The present demonstration that surface expression of ␣4␤2␦(R220H) and ␣4␤2␦(E177A) receptors were reduced suggests that the N terminus of the ␦ subunit may also be involved in receptor assembly and/or trafficking.…”

Section: Alterations Of Surface Expression Of ␣4␤2␦ Receptors and Thementioning

confidence: 99%

δ Subunit Susceptibility Variants E177A and R220H Associated with Complex Epilepsy Alter Channel Gating and Surface Expression of α4β2δ GABA_AReceptors

et al. 2006

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Most human idiopathic generalized epilepsies (IGEs) are polygenic, but virtually nothing is known of the molecular basis for any of the complex epilepsies. Recently, two GABA A receptor ␦ subunit variants (E177A, R220H) were proposed as susceptibility alleles for generalized epilepsy with febrile seizures plus and juvenile myoclonic epilepsy. In human embryonic kidney 293T cells, recombinant h␣1␤2␦(E177A) and h␣1␤2␦(R220H) receptor currents were reduced, but the basis for the current reduction was not determined. We examined the mechanistic basis for the current reduction produced by these variants using the h␣4␤2␦ receptor, an isoform more physiologically relevant and linked to epileptogenesis, by characterizing the effects of these variants on receptor cell surface expression and single-channel gating properties. Expression of variant ␣4␤2␦(R220H) receptors resulted in a decrease in surface receptor proteins, and a smaller, but significant, reduction was observed for variant ␣4␤2␦(E177A) receptors. For both variants, no significant alterations of surface expression were observed for mixed population of wild-type and variant receptors. The mean open durations of ␣4␤2␦(E177A) and ␣4␤2␦(R220H) receptor single-channel currents were both significantly decreased compared to wild-type receptors. These data suggest that both ␦(E177A) and ␦(R220H) variants may result in disinhibition in IGEs by similar cellular and molecular mechanisms, and in heterozygously affected individuals, a reduction in channel open duration of ␦ subunit-containing GABA A receptors may be the major contributor to the epilepsy phenotypes.

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“…It is somewhat less clear how salt bridge formation between Arg-32 and ␥2(Asp-123) could discourage incorporation of ␥2 subunits; however, it is important to note that salt bridges can be destabilizing (43) and that ␥2 subunit (122-129) integrity was essential for ␥2-␤3 subunit interaction (35). It is also worth mentioning that the epilepsy- associated mutation ␥2(R82Q), which has been shown to disrupt receptor assembly (36,44), is located in the ␥2 subunit loop closest to the N-terminal domain of the ␤3 subunit ␣-helix. Indeed, point mutations throughout this loop impaired ␥2 subunit incorporation.…”

Section: Altered Salt Bridge Formation and Receptor Conformationmentioning

confidence: 99%

GABRB3 Mutation, G32R, Associated with Childhood Absence Epilepsy Alters α1β3γ2L γ-Aminobutyric Acid Type A (GABAA) Receptor Expression and Channel Gating

Gurba

Hernández

et al. 2012

Journal of Biological Chemistry

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Background: A GABRB3 mutation has been associated with childhood absence epilepsy and ␤3 subunit hyperglycosylation. Results: The mutation altered subunit expression and reduced GABA A receptor function independent of N-glycosylation. Conclusion: The mutation introduced a charged residue predicted to alter subunit interactions. Significance: The distal N terminus of GABA A receptor subunits may play an unexpected role in receptor assembly and channel gating.

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The epilepsy mutation, γ2(R43Q) disrupts a highly conserved inter-subunit contact site, perturbing the biogenesis of GABAA receptors

Cited by 70 publications

References 38 publications

Reduced cortical inhibition in a mouse model of familial childhood absence epilepsy

Reduced cortical inhibition in a mouse model of familial childhood absence epilepsy

δ Subunit Susceptibility Variants E177A and R220H Associated with Complex Epilepsy Alter Channel Gating and Surface Expression of α4β2δ GABA_AReceptors

GABRB3 Mutation, G32R, Associated with Childhood Absence Epilepsy Alters α1β3γ2L γ-Aminobutyric Acid Type A (GABAA) Receptor Expression and Channel Gating

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The epilepsy mutation, γ2(R43Q) disrupts a highly conserved inter-subunit contact site, perturbing the biogenesis of GABAA receptors

Cited by 70 publications

References 38 publications

Reduced cortical inhibition in a mouse model of familial childhood absence epilepsy

Reduced cortical inhibition in a mouse model of familial childhood absence epilepsy

δ Subunit Susceptibility Variants E177A and R220H Associated with Complex Epilepsy Alter Channel Gating and Surface Expression of α4β2δ GABAAReceptors

GABRB3 Mutation, G32R, Associated with Childhood Absence Epilepsy Alters α1β3γ2L γ-Aminobutyric Acid Type A (GABAA) Receptor Expression and Channel Gating

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δ Subunit Susceptibility Variants E177A and R220H Associated with Complex Epilepsy Alter Channel Gating and Surface Expression of α4β2δ GABA_AReceptors