title: Channel Gating Regulation by the Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) First Cytosolic Loop.
abstract: In this study, we present data indicating a robust and specific domain interaction between the cystic fibrosis transmembrane conductance regulator (CFTR) first cytosolic loop (CL1) and nucleotide binding domain 1 (NBD1) that allows ion transport to proceed in a regulated fashion. We used co-precipitation and ELISA to establish the molecular contact and showed that binding kinetics were not altered by the common clinical mutation F508del. Both intrinsic ATPase activity and CFTR channel gating were inhibited severely by CL1 peptide, suggesting that NBD1/CL1 binding is a crucial requirement for ATP hydrolysis and channel function. In addition to cystic fibrosis, CFTR dysregulation has been implicated in the pathogenesis of prevalent diseases such as chronic obstructive pulmonary disease, acquired rhinosinusitis, pancreatitis, and lethal secretory diarrhea (e.g. cholera). On the basis of clinical relevance of the CFTR as a therapeutic target, a cell-free drug screen was established to identify modulators of NBD1/CL1 channel activity independent of F508del CFTR and pharmacologic rescue. Our findings support a targetable mechanism of CFTR regulation in which conformational changes in the NBDs cause reorientation of transmembrane domains via interactions with CL1 and result in channel gating.
authors.affiliation: From the Gregory Fleming James Cystic Fibrosis Research Center and; the Department of Pediatrics, Emory University, Atlanta, Georgia 30322.
authors.forename: Annette
authors.initials: A
authors.lastname: Ehrhardt
authors.affiliation: From the Gregory Fleming James Cystic Fibrosis Research Center and; Departments of Neurobiology.
authors.forename: W Joon
authors.initials: WJ
authors.lastname: Chung
authors.affiliation: From the Gregory Fleming James Cystic Fibrosis Research Center and.
authors.forename: Louise C
authors.initials: LC
authors.lastname: Pyle
authors.affiliation: Cellular, Integrative, and Developmental Biology.
authors.forename: Wei
authors.initials: W
authors.lastname: Wang
authors.affiliation: Cellular, Integrative, and Developmental Biology.
authors.forename: Krzysztof
authors.initials: K
authors.lastname: Nowotarski
authors.affiliation: the Hospital for Sick Children Research Institute, Toronto M5G 1X8, Canada.
authors.forename: Cory M
authors.initials: CM
authors.lastname: Mulvihill
authors.affiliation: the Hospital for Sick Children Research Institute, Toronto M5G 1X8, Canada.
authors.forename: Mohabir
authors.initials: M
authors.lastname: Ramjeesingh
authors.affiliation: From the Gregory Fleming James Cystic Fibrosis Research Center and; Cellular, Integrative, and Developmental Biology.
authors.forename: Jeong
authors.initials: J
authors.lastname: Hong
authors.affiliation: Chemistry, University of Alabama at Birmingham, Birmingham, Alabama 35294.
authors.forename: Sadanandan E
authors.initials: SE
authors.lastname: Velu
authors.affiliation: Bristol-Myers Squibb, Princeton, New Jersey 08543.
authors.forename: Hal A
authors.initials: HA
authors.lastname: Lewis
authors.affiliation: Eli Lilly & Co., San Diego, California 92121.
authors.forename: Shane
authors.initials: S
authors.lastname: Atwell
authors.affiliation: From the Gregory Fleming James Cystic Fibrosis Research Center and; Pharmacology, and.
authors.forename: Steve
authors.initials: S
authors.lastname: Aller
authors.affiliation: the Hospital for Sick Children Research Institute, Toronto M5G 1X8, Canada,; the Departments of Biochemistry and; Physiology, University of Toronto, Toronto M5G 1X8, Canada, and.
authors.forename: Christine E
authors.initials: CE
authors.lastname: Bear
authors.affiliation: the Department of Physiology, McGill University, Montreal H3G 1Y6, Canada.
authors.forename: Gergely L
authors.initials: GL
authors.lastname: Lukacs
authors.affiliation: From the Gregory Fleming James Cystic Fibrosis Research Center and; Cellular, Integrative, and Developmental Biology.
authors.forename: Kevin L
authors.initials: KL
authors.lastname: Kirk
authors.affiliation: the Department of Pediatrics, Emory University, Atlanta, Georgia 30322,. Electronic address: esorscher@emory.edu.
authors.forename: Eric J
authors.initials: EJ
authors.lastname: Sorscher
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism; D000595:Amino Acid Sequence; D001665:Binding Sites; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D006868:Hydrolysis; D007700:Kinetics; D008969:Molecular Sequence Data; D017434:Protein Structure, Tertiary
chemical_list: D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D000255:Adenosine Triphosphate
keywords: ABC transporter; channel activation; chloride channel; cystic fibrosis; cystic fibrosis transmembrane conductance regulator (CFTR); domain interaction; drug discovery; epithelial cell; pore formation