Genetic variants causing the fast-channel congenital myasthenic syndrome (CMS) have been identified in the α, δ, and ε but not the β subunit of acetylcholine receptor (AChR). A 16-year-old girl with severe myasthenia had low-amplitude and fast-decaying miniature endplate potentials. Mutation analysis revealed two heteroallelic variants in CHRNB1 encoding the AChR β subunit: a novel c.812C>T (p.P248L) variant in M1-M2 linker (p.P271L in HGVS nomenclature), and a ~430 bp deletion causing loss of exon 8 leading to frame-shift and a premature stop codon (p.G251Dfs*21). P248 is conserved in all β subunits of different species, but not in other AChR subunits. Measurements of radio-labeled α-bungarotoxin binding show that βP248L reduces AChR expression to 60% of wild-type. Patch clamp recordings of ACh-elicited single channel currents demonstrate that βP248L shortens channel opening bursts from 3.3 ms to 1.2 ms, and kinetic analyses predict that the decay of the synaptic response is accelerated 2.4-fold due to reduced probability of channel reopening. Substituting βP248 with threonine, alanine or glycine reduces the burst duration to 2.3, 1.7, and 1.5 ms, respectively. In non-β subunits, substituting leucine for residues corresponding to βP248 prolongs the burst duration to 4.5 ms in the α subunit, shortens it to 2.2 ms in the δ subunit, and has no effect in the ε subunit. Conversely, substituting proline for residues corresponding to βP248 prolongs the burst duration to 8.7 ms in the α subunit, to 4.6 ms in the δ subunit, but has no effect in the ε subunit. Thus, this fast channel CMS is caused by the dual defects of βP248L in reducing expression of the mutant receptor and accelerating the decay of the synaptic response. The results also reveal subunit-specific contributions of the M1-M2 linker to the durations of channel opening bursts.
Keywords: Acetylchoine receptor; Channel gating; Congenital myasthenic syndromes; Fast-channel myasthenia; Intracellular linker; M1-M2 linker; Neuromuscular disease; Neuromuscular junction; Receptor activation; β subunit.
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