New insights into the mechanism of activation of Cys-loop receptors

ANDERSEN N; CORRADI J; BARTOS M; SINE SM; BOUZAT C

Huerta Grande

Congreso; II Reunión Conjunta de Neurociencias; 2010

Sociedad Argentina de Investigacion en Neurociencia

Cys-loop receptors are pentameric neurotransmitter-gated ion channels that contain an extracellular domain, which carries the binding sites, and a transmembrane domain, which forms the ion channel. The transition zone between these domains, called coupling region, is essential for coupling agonist binding to the opening of the channel. To understand this process, we used the chimeric alpha7-5HT3A receptor as a model of homomeric Cys-loop receptors. We constructed a subunit with a mutation that disables the coupling region and a reporter mutation that alters unitary conductance, and co-expressed mutant and non-mutant subunits. By measuring single-channel currents we determined that the amplitude of each opening event reports the number of subunits with intact coupling regions in the pentameric receptor. We found that: i) the coupling region of each subunit contributes additively to channel lifetime of the receptor; ii) channel opening is profoundly impaired in receptors containing only one active interface; and iii) for channel gating each functional binding site requires direct connection to an active interface, indicating that changes at a binding site are confined to the same subunit. Our results provide a framework for defining mechanisms of activation for these receptors.