Mutation causing congenital myasthenia reveals acetylcholine receptor β/δ subunit interaction essential for assembly

Quiram, Polly A.; Ohno, Kinji; Milone, Margherita; Patterson, Marc C.; Pruitt, Ned J.; Brengman, Joan M.; Sine, Steven M.; Engel, Andrew G.

doi:10.1172/jci8179

Cited by 76 publications

(54 citation statements)

References 31 publications

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“…The Role of C-terminal Motifs in the Assembly of Other Cysloop Family Members-Although this is the first report implicating a region other than the N-terminal domain in GABA A receptor assembly, the M3-M4 loop and M4 domain are known to be important for assembly of nAChRs (35). A three-amino acid deletion in the nicotinic acetylcholine receptor ␤ subunit M3-M4 loop, ␤426⌬EQE, associated with congenital myasthenic syndromes has been demonstrated to impair the interaction between ␤ and ␦ subunits (35).…”

Section: Mutation Of the Conserved Aspartate Residue Compromised Pentmentioning

confidence: 94%

“…A three-amino acid deletion in the nicotinic acetylcholine receptor ␤ subunit M3-M4 loop, ␤426⌬EQE, associated with congenital myasthenic syndromes has been demonstrated to impair the interaction between ␤ and ␦ subunits (35). Moreover mutation of histidine 408 in the ␣ subunit that is adja-FIGURE 11.…”

Section: Mutation Of the Conserved Aspartate Residue Compromised Pentmentioning

confidence: 99%

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A Conserved Cys-loop Receptor Aspartate Residue in the M3-M4 Cytoplasmic Loop Is Required for GABAA Receptor Assembly

Botzolakis

Tang

et al. 2008

Journal of Biological Chemistry

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Members of the Cys-loop superfamily of ligand-gated ion channels, which mediate fast synaptic transmission in the nervous system, are assembled as heteropentamers from a large repertoire of neuronal subunits. Although several motifs in subunit N-terminal domains are known to be important for subunit assembly, increasing evidence points toward a role for C-terminal domains. Using a combination of flow cytometry, patch clamp recording, endoglycosidase H digestion, brefeldin A treatment, and analytic centrifugation, we identified a highly conserved aspartate residue at the boundary of the M3-M4 loop and the M4 domain that was required for binary and ternary ␥-aminobutyric acid type A receptor surface expression. Mutation of this residue caused mutant and partnering subunits to be retained in the endoplasmic reticulum, reflecting impaired forward trafficking. Interestingly although mutant and partnering wild type subunits could be coimmunoprecipitated, analytic centrifugation studies demonstrated decreased formation of pentameric receptors, suggesting that this residue played an important role in later steps of subunit oligomerization. We thus conclude that C-terminal motifs are also important determinants of Cys-loop receptor assembly.The Cys-loop superfamily of ligand-gated ion channels, which includes ␥-aminobutyric acid type A (GABA A ) 2 and type C (GABA C ), nicotinic acetylcholine, glycine, and 5-hydroxytryptamine type 3 receptors, mediates fast synaptic transmission in the nervous system. Mutations that alter Cys-loop receptor surface density by affecting receptor biogenesis have been associated with idiopathic generalized epilepsies (1-4), congenital myasthenic syndromes (5), and psychiatric disorders (6). Unfortunately because the structural and cellular determinants of receptor biogenesis are poorly understood, development of effective treatment strategies remains a significant challenge.A wealth of evidence suggests that Cys-loop receptors are assembled as heteropentamers from a large repertoire of neuronal subunits (7-10). Subunits share a similar topology that includes an extracellular N-terminal domain, four transmembrane domains, three loops including a large cytoplasmic loop, and a variable length extracellular C-terminal tail (7, 11). Receptor assembly is thought to occur in the endoplasmic reticulum (ER) following glycosylation and folding of de novo synthesized subunits (12-15). Assembly is closely monitored by ER quality control machinery, and consequently subunits that fail to assemble properly are retained and degraded (15)(16)(17). Although N-terminal motifs are known to be important for subunit assembly (18,19), recent studies in nicotinic acetylcholine receptors (nAChRs) suggest that C-terminal motifs may also play a role (20).GABA A receptors are the most abundant Cys-loop receptor in the mammalian brain and are responsible for the majority of fast inhibitory neurotransmission. Like other Cys-loop superfamily receptors, they are pentamers assembled from combinations of 16 subunit subtypes ...

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Section: Mutation Of the Conserved Aspartate Residue Compromised Pentmentioning

confidence: 94%

Section: Mutation Of the Conserved Aspartate Residue Compromised Pentmentioning

confidence: 99%

A Conserved Cys-loop Receptor Aspartate Residue in the M3-M4 Cytoplasmic Loop Is Required for GABAA Receptor Assembly

Botzolakis

Tang

et al. 2008

Journal of Biological Chemistry

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“…There was robust expression of αV132L-AChR compared with wild type (mean ± SD: 135% ± 12%, n = 3), but essentially no expression of α381delC-AChR (0.21% ± 0.22%, n = 3; P < 0.0001 vs. wild type). Robust expression of αV132L-AChR indicates that all three non-α subunits incorporate into surface receptors because omission of one or more subunits would abolish or markedly reduce expression (35,36). Thus α381delC is a null mutation and αV132L determines the phenotype.…”

Section: Characteristics Of Cms Patientmentioning

confidence: 99%

Mutation causing severe myasthenia reveals functional asymmetry of AChR signature cystine loops in agonist binding and gating

Shen¹,

Ohno²,

Tsujino³

et al. 2003

J. Clin. Invest.

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IntroductionCongenital myasthenic syndromes (CMSs) are heterogeneous disorders caused by presynaptic, synaptic, or postsynaptic defects (1). Postsynaptic CMSs can be broadly classified according to whether the response to ACh is increased or decreased relative to control responses. An increased response is observed in slowchannel syndromes due to prolonged activation episodes of AChR. A decreased response is observed with AChR deficiency due to mutations in AChR subunit genes (2) or in rapsyn (3), and in the fast-channel syndromes. Fast-channel syndromes are characterized by attenuated and abnormally rapidly decaying endplate (EP) currents, abnormally brief single-channel currents, and decreased probability of channel opening.Several molecular mechanisms underlying fastchannel syndromes have been identified. At the ligand-binding site formed by α and ε subunits, the mutation εP121L reduces ACh affinity for the open channel state and slows the rate of channel opening, resulting in a moderately severe myasthenic phenotype (4). Also at the α-ε site, the mutation εD175N reduces ACh affinity of the resting closed state and impairs gating efficiency, while the nearby mutation εN182Y increases ACh affinity for the resting closed state and also impairs gating (5). The phenotypic consequences of these combined mutations are also moderately severe. At the ligand-binding site formed by the α and δ subunits, the mutation δE59K likely reduces ACh affinity (6), but the altered steps in the activation process have not been identified. This mutation was shown to result in hypomotility in utero, multiple congenital joint contractures, and neonatal respiratory distress, but the patient subsequently improved and could walk short distances. In the third transmembrane domain of the α subunit, the mutation αV285I impairs gating efficiency (7) but results in a mild phenotype. Finally, in the amphipathic helix of the long cytoplasmic loop of the ε subunit, either a duplication of codons 413-418 (8) or an αA411P missense mutation (9) result in heterogeneous channel gating kinetics; the phenotypic consequences are mild to moderately severe. We describe a highly disabling congenital myasthenic syndrome (CMS) associated with rapidly decaying, low-amplitude synaptic currents, and trace its cause to a valine to leucine mutation in the signature cystine loop (cys-loop) of the AChR α subunit. The recently solved crystal structure of an ACh-binding protein places the cys-loop at the junction between the extracellular ligand-binding and transmembrane domains where it may couple agonist binding to channel gating. We therefore analyzed the kinetics of ACh-induced single-channel currents to identify elementary steps in the receptor activation mechanism altered by the αV132L mutation. The analysis reveals that αV132L markedly impairs ACh binding to receptors in the resting closed state, decreasing binding affinity for the second binding step 30-fold, but attenuates gating efficiency only about twofold. By contrast, mutation of the equivalent valine...

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“…By contrast, mutations in subunits of the fetal receptor that result in a reduced response to ACh might be expected to inhibit seriously fetal movement during crucial developmental periods. However, features of AMC were not noted in a case of AChR deficiency due to a 3-codon deletion of the β subunit that affects AChR assembly (27) or in a case of fast-channel syndrome due to a missense mutation in the α subunit M3 region (9). It may be that AMC was present in these cases but not documented or that these mutations did not cause sufficient dysfunction of fetal AChR to result in AMC.…”

Section: Discussionmentioning

confidence: 93%

Acetylcholine receptor δ subunit mutations underlie a fast-channel myasthenic syndrome and arthrogryposis multiplex congenita

Brownlow¹,

Webster²,

Croxen³

et al. 2001

J. Clin. Invest.

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Mutation causing congenital myasthenia reveals acetylcholine receptor β/δ subunit interaction essential for assembly

Cited by 76 publications

References 31 publications

A Conserved Cys-loop Receptor Aspartate Residue in the M3-M4 Cytoplasmic Loop Is Required for GABAA Receptor Assembly

A Conserved Cys-loop Receptor Aspartate Residue in the M3-M4 Cytoplasmic Loop Is Required for GABAA Receptor Assembly

Mutation causing severe myasthenia reveals functional asymmetry of AChR signature cystine loops in agonist binding and gating

Acetylcholine receptor δ subunit mutations underlie a fast-channel myasthenic syndrome and arthrogryposis multiplex congenita

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