title: Cystic fibrosis transmembrane conductance regulator interacts with multiple immunoglobulin domains of filamin A.
abstract: Mutations of the chloride channel cystic fibrosis transmembrane conductance regulator (CFTR) that impair its apical localization and function cause cystic fibrosis. A previous report has shown that filamin A (FLNa), an actin-cross-linking and -scaffolding protein, interacts directly with the cytoplasmic N terminus of CFTR and that this interaction is necessary for stability and confinement of the channel to apical membranes. Here, we report that the CFTR N terminus has sequence similarity to known FLNa-binding partner-binding sites. FLNa has 24 Ig (IgFLNa) repeats, and a CFTR peptide pulled down repeats 9, 12, 17, 19, 21, and 23, which share sequence similarity yet differ from the other FLNa Ig domains. Using known structures of IgFLNa.partner complexes as templates, we generated in silico models of IgFLNa.CFTR peptide complexes. Point and deletion mutants of IgFLNa and CFTR informed by the models, including disease-causing mutations L15P and W19C, disrupted the binding interaction. The model predicted that a P5L CFTR mutation should not affect binding, but a synthetic P5L mutant peptide had reduced solubility, suggesting a different disease-causing mechanism. Taken together with the fact that FLNa dimers are elongated ( approximately 160 nm) strands, whereas CFTR is compact (6 approximately 8 nm), we propose that a single FLNa molecule can scaffold multiple CFTR partners. Unlike previously defined dimeric FLNa.partner complexes, the FLNa-monomeric CFTR interaction is relatively weak, presumptively facilitating dynamic clustering of CFTR at cell membranes. Finally, we show that deletion of all CFTR interacting domains from FLNa suppresses the surface expression of CFTR on baby hamster kidney cells.
authors.affiliation: NHLBI, National Institutes of Health, Bethesda, Maryland 20892, USA.
authors.forename: Martin P
authors.initials: MP
authors.lastname: Playford
authors.forename: Elisa
authors.initials: E
authors.lastname: Nurminen
authors.forename: Olli T
authors.initials: OT
authors.lastname: Pentikäinen
authors.forename: Sharon L
authors.initials: SL
authors.lastname: Milgram
authors.forename: John H
authors.initials: JH
authors.lastname: Hartwig
authors.forename: Thomas P
authors.initials: TP
authors.lastname: Stossel
authors.forename: Fumihiko
authors.initials: F
authors.lastname: Nakamura
mesh_terms: D000199:Actins / Q000737:chemistry; D000595:Amino Acid Sequence; D000818:Animals; D002462:Cell Membrane / Q000378:metabolism; D003285:Contractile Proteins / Q000737:chemistry*; D006224:Cricetinae; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics / Q000378:metabolism*; D019281:Dimerization; D064448:Filamins; D006801:Humans; D007136:Immunoglobulins / Q000737:chemistry*; D008840:Microfilament Proteins / Q000737:chemistry*; D008969:Molecular Sequence Data; D009154:Mutation; D010455:Peptides / Q000737:chemistry; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D012995:Solubility
chemical_list: D000199:Actins; D003285:Contractile Proteins; D064448:Filamins; D007136:Immunoglobulins; D008840:Microfilament Proteins; D010455:Peptides; D019005:Cystic Fibrosis Transmembrane Conductance Regulator