Identification of a determinant of acetylcholine receptor gating kinetics in the extracellular portion of the gamma subunit.
A large body of structure-function studies has identified many of the functional motifs underlying ion permeation through acetylcholine receptor (AChR) channels. The structural basis of channel gating kinetics is, however, incompletely understood. We have previously identified a novel shorter form of the AChR gamma subunit, which lacks the 52 amino acids within the extracellular amino-terminal half, encoded by exon 5. To define the contribution of the missing domain to AChR channel function, we have transiently coexpressed the mouse short gamma subunit [gamma(s)] with alpha, beta and delta subunits in human cells and recorded single-channel currents from the resulting AChRs. Our findings show that replacement of the gamma by the gamma(s) subunit confers a long duration characteristic to AChR channel openings without altering unitary conductance sizes or receptor affinity for the transmitter. We also show that alpha beta gamma(s) delta AChR channels exhibit a peculiar voltage sensitivity characterized by a short opening duration when the membrane potential is hyperpolarized. Together, these findings indicate that the domain in the extracellular amino-terminal half of the gamma subunit that encompasses a conserved disulphide loop and a critical tyrosine residue implicated in receptor oligomerization and insertion at the cell surface is a functional motif that also modulates AChR channel gating kinetics. The results also provide a molecular explanation of the functional diversity exhibited by skeletal muscle AChRs during development.
Fucile
S
S
Istituto Pasteur-Fondazione Cenci Bolognetti, Dipartimento di Medicina Sperimentale e Patologia, Università di Roma, Italy.
Mileo
A M
AM
Grassi
F
F
Salvatore
A M
AM
Alemà
S
S
Eusebi
F
F
D000109:Acetylcholine / Q000494:pharmacology; D000818:Animals; D002460:Cell Line; D004553:Electric Conductivity; D006801:Humans; D015640:Ion Channel Gating; D007473:Ion Channels / Q000378:metabolism*; D007700:Kinetics; D051379:Mice; D011950:Receptors, Cholinergic / Q000737:chemistry / Q000378:metabolism* / Q000502:physiology; D013329:Structure-Activity Relationship
D007473:Ion Channels; D011950:Receptors, Cholinergic; D000109:Acetylcholine