title: A structural requirement for processing the cardiac K+ channel KCNQ1.
abstract: Normal membrane protein function requires trafficking from the endoplasmic reticulum. Here, we studied processing of the KCNQ1 channel mutated in LQT1, the commonest form of the long QT syndrome. Serial C terminus truncations identified a small region (amino acids (aa) 610-620) required for normal cell surface expression. Non-trafficked truncations assembled as tetramers but were nevertheless retained in the endoplasmic reticulum. Further mutagenesis did not identify specific residues mediating channel processing; cell surface expression was preserved with the mutation of known trafficking motifs in the channel and with alanine scanning across aa 610-620. Structural prediction algorithms place aa 610-620 at the C-terminal end of an alpha-helix (aa 586-618) that includes a leucine zipper and is part of a coiled coil. Mutants disrupting the leucine zipper but preserving the predicted coiled coil reached the cell surface, whereas those disrupting the coil did not. These data suggest that specific protein-protein interactions are required for normal channel processing. Further biochemical studies ruled out three candidate proteins, namely KCNE1, yotiao, and KCNQ1 itself, as effectors of this coiled coil-mediated trafficking. Four LQT1 mutations within this helix generated little or no current and were not expressed on the cell surface, whereas LQT1 mutations in adjacent residues, which produce a milder clinical phenotype, generate only slightly reduced current and are expressed on the cell surface. These data suggest that mutations within this domain cause human disease by interfering with normal channel processing. More generally, we have identified a domain whose structural integrity is required for normal surface expression of the KCNQ1 channel.
authors.affiliation: Department of Medicine, Vanderbilt University School of Medicine, Nashville, Tennessee 37232, USA.
authors.forename: Hideaki
authors.initials: H
authors.lastname: Kanki
authors.forename: Sabina
authors.initials: S
authors.lastname: Kupershmidt
authors.forename: Tao
authors.initials: T
authors.lastname: Yang
authors.forename: Sam
authors.initials: S
authors.lastname: Wells
authors.forename: Dan M
authors.initials: DM
authors.lastname: Roden
mesh_terms: D000409:Alanine / Q000235:genetics; D000465:Algorithms; D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000235:genetics; D019281:Dimerization; D004553:Electric Conductivity; D004591:Electrophoresis, Polyacrylamide Gel; D000939:Epitopes / Q000737:chemistry; D005456:Fluorescent Dyes; D017353:Gene Deletion; D007150:Immunohistochemistry; D007163:Immunosorbent Techniques; D051656:KCNQ Potassium Channels; D051657:KCNQ1 Potassium Channel; D016350:Leucine Zippers / Q000235:genetics; D008133:Long QT Syndrome / Q000235:genetics; D008164:Luminescent Proteins / Q000235:genetics; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D018408:Patch-Clamp Techniques; D010446:Peptide Fragments / Q000737:chemistry / Q000235:genetics; D016133:Polymerase Chain Reaction; D015221:Potassium Channels / Q000737:chemistry* / Q000235:genetics / Q000502:physiology*; D024642:Potassium Channels, Voltage-Gated; D017433:Protein Structure, Secondary; D011993:Recombinant Fusion Proteins; D013329:Structure-Activity Relationship; D014162:Transfection
chemical_list: D001426:Bacterial Proteins; D000939:Epitopes; D005456:Fluorescent Dyes; D051656:KCNQ Potassium Channels; D051657:KCNQ1 Potassium Channel; D008164:Luminescent Proteins; D010446:Peptide Fragments; D015221:Potassium Channels; D024642:Potassium Channels, Voltage-Gated; D011993:Recombinant Fusion Proteins; C054562:yellow fluorescent protein, Bacteria; D000409:Alanine