title: Pore-to-gate coupling of HCN channels revealed by a pore variant that contributes to gating but not permeation.
abstract: Although ample evidence suggests the presence of an intracellular activation gate in HCN (pacemaker) channels, mutations in the outer pore can alter gating properties. Here we investigated the role of the outer pore residue A354 in HCN1 gating by systematically converting it to the equivalent residues (T, Y, and F) found in K(+)-channels. A354T negatively shifted steady-state activation (DeltaV(1/2) approximately -25 mV), decelerated gating kinetics (by up to 8-fold), and abolished the effects of external ions on gating. A354Y and A354F did not yield functional currents when expressed alone, although immunofluorescence microscopy indicated the presence of these channel proteins on the membrane surface. Currents recorded after co-expressing A354Y with WT HCN1 were reduced in amplitude (relative to WT alone) and had changes in gating similar to those of A354T. We conclude that the pore variant at position 354 contributes to gating but not permeation, and that the HCN outer pore may be involved in gating via a pore-to-gate coupling mechanism.
authors.affiliation: Department of Medicine, Johns Hopkins University, USA.
authors.forename: Ezana M
authors.initials: EM
authors.lastname: Azene
authors.forename: Dongpei
authors.initials: D
authors.lastname: Sang
authors.forename: Suk-Ying
authors.initials: SY
authors.lastname: Tsang
authors.forename: Ronald A
authors.initials: RA
authors.lastname: Li
mesh_terms: D000409:Alanine / Q000235:genetics / Q000378:metabolism; D000595:Amino Acid Sequence; D019943:Amino Acid Substitution / Q000235:genetics*; D000818:Animals; D002463:Cell Membrane Permeability; D054815:Cyclic Nucleotide-Gated Cation Channels; D015870:Gene Expression; D064428:Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels; D015640:Ion Channel Gating / Q000502:physiology*; D007473:Ion Channels / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D007700:Kinetics; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009865:Oocytes / Q000378:metabolism; D015221:Potassium Channels; D021122:Protein Subunits / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D016415:Sequence Alignment; D014982:Xenopus laevis
chemical_list: D054815:Cyclic Nucleotide-Gated Cation Channels; D064428:Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels; D007473:Ion Channels; D015221:Potassium Channels; D021122:Protein Subunits; D000409:Alanine