Abstract Details

Title Investigation of Congenital Myasthenic Syndrome (CMS) Reveals Asymmetric Contribution of Invariant Cys-Loop Aspartate Residues Critical for AChR Channel Opening

Topic Muscle Disease/Neuromuscular Junction

Presentation(s) S45 - (-)

Poster/Presentation
Number 002

Objective

Background Most CMS are caused by mutations of AChR, a member of Cys-loop receptor superfamily. The signature Cys-loop at the interface between binding and gating domains of the receptor plays a crucial role in coupling agonist binding to gating but the mechanism is unclear. An invariant aspartate is present at equivalent positions in all subunits of all Cys-loop superfamily receptors of all species, but its functional significance and subunit specificity have not been elucidated.

Design/Methods We used mutation analysis, mutagenesis, and single-channel recordings from wild-type and mutant receptors expressed in HEK cells.

Results An 11-year-old girl with severe myasthenic symptoms and respiratory crises since birth and partially responsive to cholinergic agonists harbors a mutation of the invariant Cys-loop Asp in the AChR ? subunit, ?D140N, not present in the exome variant database, and a heteroallelic splice-site mutation ?IVS7-2A>C; hence ?D140N determines the phenotype. Compared to wild-type, ?D140N reduces AChR expression in HEK cells to 21% and the length of channel opening bursts to 50%, but has little effect on gating efficiency (channel opening rate/channel closing rate). Mutation of the invariant Asp in other subunits reveals that compared to wild-type expression, burst length, and gating efficiency are 50%, 6% and 1% for ?D138N-AChR, 80%, 16%, and 25% for ?D138N-AChR, and close to wild-type for ?D138N-AChR.

Conclusions ?D140N is the first identified mutation of an invariant Cys-loop Asp residue. It compromises neuromuscular transmission by decreasing AChR expression and burst length. The invariant Asp residues contribute to expression, burst length and gating efficiency in a subunit-specific manner; ?-Asp has profound effects on burst length and gating efficiency whereas ?-Asp predominantly affects AChR expression. Thus the ?-subunit Cys-loop Asp emerges as key regulator of transduction of agonist binding to channel opening.

Authors/Disclosures
Xin Ming Shen, PhD, FAAN (Mayo Clinic) PRESENTER	The institution of Dr. Shen has received research support from NIH. The institution of Dr. Shen has received research support from Myasthenia Gravis Foundation of American.
	No disclosure on file
Joan M. Brengman (Mayo Clinic)	No disclosure on file
Andrew G. Engel, MD, FAAN (Mayo Clinic Rochester)	The institution of Dr. Engel has received research support from NIH.
Edward J. Cupler, MD, FAAN (King Faisal Specialist Hospital and Research Center)	Dr. Cupler has received personal compensation in the range of $500-$4,999 for serving on a Speakers Bureau for Sanofi.

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