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Mutations causing congenital myasthenia reveal principal coupling pathway in the acetylcholine receptor ?-subunit.

ABSTRACT: We identify 2 homozygous mutations in the ?-subunit of the muscle acetylcholine receptor (AChR) in 3 patients with severe congenital myasthenia: ?R218W in the pre-M1 region in 2 patients and ?E184K in the ?8-?9 linker in 1 patient. Arg218 is conserved in all eukaryotic members of the Cys-loop receptor superfamily, while Glu184 is conserved in the ?-, ?-, and ?-subunits of AChRs from all species. ?R218W reduces channel gating efficiency 338-fold and AChR expression on the cell surface 5-fold, whereas ?E184K reduces channel gating efficiency 11-fold but does not alter AChR cell surface expression. Determinations of the effective channel gating rate constants, combined with mutant cycle analyses, demonstrate strong energetic coupling between ?R218 and ?E184, and between ?R218 and ?E45 from the ?1-?2 linker, as also observed for equivalent residues in the principal coupling pathway of the ?-subunit. Thus, efficient and rapid gating of the AChR channel is achieved not only by coupling between conserved residues within the principal coupling pathway of the ?-subunit, but also between corresponding residues in the ?-subunit.

SUBMITTER: Shen XM

PROVIDER: S-EPMC5821208 | biostudies-literature | 2018 Jan

REPOSITORIES: biostudies-literature

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Mutations causing congenital myasthenia reveal principal coupling pathway in the acetylcholine receptor ε-subunit.

Shen Xin-Ming XM Brengman Joan M JM Shen Shelley S Durmus Hacer H Preethish-Kumar Veeramani V Yuceyar Nur N Vengalil Seena S Nalini Atchayaram A Deymeer Feza F Sine Steven M SM Engel Andrew G AG

JCI insight 20180125 2

We identify 2 homozygous mutations in the ε-subunit of the muscle acetylcholine receptor (AChR) in 3 patients with severe congenital myasthenia: εR218W in the pre-M1 region in 2 patients and εE184K in the β8-β9 linker in 1 patient. Arg218 is conserved in all eukaryotic members of the Cys-loop receptor superfamily, while Glu184 is conserved in the α-, δ-, and ε-subunits of AChRs from all species. εR218W reduces channel gating efficiency 338-fold and AChR expression on the cell surface 5-fold, whe ...[more]

PMID: 29367459

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Project description:Congenital hypogonadotropic hypogonadism (CHH) is characterized by low gonadotropins and failure to progress normally through puberty. Mutations in the gene encoding the GnRH receptor (GNRHR1) result in CHH when present as compound heterozygous or homozygous inactivating mutations. This study identifies and characterizes the properties of two novel GNRHR1 mutations in a family in which three brothers display normosmic CHH while their sister was unaffected. Molecular analysis in the proband and the affected brothers revealed two novel non-synonymous missense GNRHR1 mutations, present in a compound heterozygous state, whereas their unaffected parents possessed only one inactivating mutation, demonstrating the autosomal recessive transmission in this kindred and excluding X-linked inheritance equivocally suggested by the initial pedigree analysis. The first mutation at c.845 C>G introduces an Arg substitution for the conserved Pro 282 in transmembrane domain (TMD) 6. The Pro282Arg mutant is unable to bind radiolabeled GnRH analogue. As this conserved residue is important in receptor conformation, it is likely that the mutation perturbs the binding pocket and affects trafficking to the cell surface. The second mutation at c.968 A>G introduces a Cys substitution for Tyr 323 in the functionally crucial N/DPxxY motif in TMD 7. The Tyr323Cys mutant has an increased GnRH binding affinity but reduced receptor expression at the plasma membrane and impaired G protein-coupling. Inositol phosphate accumulation assays demonstrated absent and impaired Gα(q/11) signal transduction by Pro282Arg and Tyr323Cys mutants, respectively. Pretreatment with the membrane permeant GnRHR antagonist NBI-42902, which rescues cell surface expression of many GNRHR1 mutants, significantly increased the levels of radioligand binding and intracellular signaling of the Tyr323Cys mutant but not Pro282Arg. Immunocytochemistry confirmed that both mutants are present on the cell membrane albeit at low levels. Together these molecular deficiencies of the two novel GNRHR1 mutations lead to the CHH phenotype when present as a compound heterozygote.

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Mutations causing congenital myasthenia reveal principal coupling pathway in the acetylcholine receptor ?-subunit.

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Mutations causing congenital myasthenia reveal principal coupling pathway in the acetylcholine receptor ε-subunit.

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