title: Rattlesnake Phospholipase A2 Increases CFTR-Chloride Channel Current and Corrects ∆F508CFTR Dysfunction: Impact in Cystic Fibrosis.
abstract: Deletion of Phe508 in the nucleotide binding domain (∆F508-NBD1) of the cystic fibrosis transmembrane regulator (CFTR; a cyclic AMP-regulated chloride channel) is the most frequent mutation associated with cystic fibrosis. This mutation affects the maturation and gating of CFTR protein. The search for new high-affinity ligands of CFTR acting as dual modulators (correctors/activators) presents a major challenge in the pharmacology of cystic fibrosis. Snake venoms are a rich source of natural multifunctional proteins, potential binders of ion channels. In this study, we identified the CB subunit of crotoxin from Crotalus durissus terrificus as a new ligand and allosteric modulator of CFTR. We showed that CB interacts with NBD1 of both wild type and ∆F508CFTR and increases their chloride channel currents. The potentiating effect of CB on CFTR activity was demonstrated using electrophysiological techniques in Xenopus laevis oocytes, in CFTR-HeLa cells, and ex vivo in mouse colon tissue. The correcting effect of CB was shown by functional rescue of CFTR activity after 24-h ΔF508CFTR treatments with CB. Moreover, the presence of fully glycosylated CFTR was observed. Molecular docking allowed us to propose a model of the complex involving of the ABCβ and F1-like ATP-binding subdomains of ΔF508-NBD1. Hydrogen-deuterium exchange analysis confirmed stabilization in these regions, also showing allosteric stabilization in two other distal regions. Surface plasmon resonance competition studies showed that CB disrupts the ∆F508CFTR-cytokeratin 8 complex, allowing for the escape of ∆F508CFTR from degradation. Therefore CB, as a dual modulator of ΔF508CFTR, constitutes a template for the development of new anti-CF agents.
authors.affiliation: Institut Pasteur, Unité Récepteurs-Canaux, CNRS,UMR 3571, 25, rue du Dr. Roux, F-75015, Paris, France. Electronic address: grazyna.faure-kuzminska@pasteur.fr.
authors.forename: Grazyna
authors.initials: G
authors.lastname: Faure
authors.affiliation: INSERM U1151, team Canalopathies épithéliales: la mucoviscidose et autres maladies, Université Paris Descartes, Paris, France.
authors.forename: Naziha
authors.initials: N
authors.lastname: Bakouh
authors.affiliation: UPMC Université Paris 06, UMRS 872, Laboratoire de génomique, physiologie et physiopathologie rénales, Paris, France.
authors.forename: Stéphane
authors.initials: S
authors.lastname: Lourdel
authors.affiliation: Polish Academy of Sciences, Institute of Biochemistry and Biophysics, Warsaw, Poland.
authors.forename: Norbert
authors.initials: N
authors.lastname: Odolczyk
authors.affiliation: Polish Academy of Sciences, Institute of Biochemistry and Biophysics, Warsaw, Poland.
authors.forename: Aiswarya
authors.initials: A
authors.lastname: Premchandar
authors.affiliation: INSERM U1151, team Canalopathies épithéliales: la mucoviscidose et autres maladies, Université Paris Descartes, Paris, France.
authors.forename: Nathalie
authors.initials: N
authors.lastname: Servel
authors.affiliation: INSERM U1151, team Canalopathies épithéliales: la mucoviscidose et autres maladies, Université Paris Descartes, Paris, France.
authors.forename: Aurélie
authors.initials: A
authors.lastname: Hatton
authors.affiliation: Institut Pasteur, Unité Récepteurs-Canaux, CNRS,UMR 3571, 25, rue du Dr. Roux, F-75015, Paris, France.
authors.forename: Maciej K
authors.initials: MK
authors.lastname: Ostrowski
authors.affiliation: Institut Pasteur, Unité Récepteurs-Canaux, CNRS,UMR 3571, 25, rue du Dr. Roux, F-75015, Paris, France.
authors.forename: Haijin
authors.initials: H
authors.lastname: Xu
authors.affiliation: Institut Pasteur, Plate-forme de Cristallographie, CNRS-UMR 3528, Paris, France.
authors.forename: Frederick A
authors.initials: FA
authors.lastname: Saul
authors.affiliation: INSERM U1151, team Canalopathies épithéliales: la mucoviscidose et autres maladies, Université Paris Descartes, Paris, France.
authors.forename: Christelle
authors.initials: C
authors.lastname: Moquereau
authors.affiliation: INSERM U1151, team Canalopathies épithéliales: la mucoviscidose et autres maladies, Université Paris Descartes, Paris, France.
authors.forename: Sara
authors.initials: S
authors.lastname: Bitam
authors.affiliation: INSERM U1151, team Canalopathies épithéliales: la mucoviscidose et autres maladies, Université Paris Descartes, Paris, France.
authors.forename: Iwona
authors.initials: I
authors.lastname: Pranke
authors.affiliation: INSERM U1151, team Canalopathies épithéliales: la mucoviscidose et autres maladies, Université Paris Descartes, Paris, France.
authors.forename: Gabrielle
authors.initials: G
authors.lastname: Planelles
authors.affiliation: UPMC Université Paris 06, UMRS 872, Laboratoire de génomique, physiologie et physiopathologie rénales, Paris, France.
authors.forename: Jacques
authors.initials: J
authors.lastname: Teulon
authors.affiliation: Department of Molecular Genetics, German Cancer Research Center, D-69120 Heidelberg, Germany; Institute of Neuropathology, University Hospital Erlangen, D-91054 Erlangen, Germany.
authors.forename: Harald
authors.initials: H
authors.lastname: Herrmann
authors.affiliation: Department of Physiology, McGill University, Montreal, Canada.
authors.forename: Ariel
authors.initials: A
authors.lastname: Roldan
authors.affiliation: Polish Academy of Sciences, Institute of Biochemistry and Biophysics, Warsaw, Poland.
authors.forename: Piotr
authors.initials: P
authors.lastname: Zielenkiewicz
authors.affiliation: Polish Academy of Sciences, Institute of Biochemistry and Biophysics, Warsaw, Poland.
authors.forename: Michal
authors.initials: M
authors.lastname: Dadlez
authors.affiliation: Department of Physiology, McGill University, Montreal, Canada.
authors.forename: Gergely L
authors.initials: GL
authors.lastname: Lukacs
authors.affiliation: INSERM U1151, team Canalopathies épithéliales: la mucoviscidose et autres maladies, Université Paris Descartes, Paris, France.
authors.forename: Isabelle
authors.initials: I
authors.lastname: Sermet-Gaudelus
authors.affiliation: INSERM U1151, team Canalopathies épithéliales: la mucoviscidose et autres maladies, Université Paris Descartes, Paris, France.
authors.forename: Mario
authors.initials: M
authors.lastname: Ollero
authors.affiliation: Institut Pasteur, Unité Récepteurs-Canaux, CNRS,UMR 3571, 25, rue du Dr. Roux, F-75015, Paris, France.
authors.forename: Pierre-Jean
authors.initials: PJ
authors.lastname: Corringer
authors.affiliation: INSERM U1151, team Canalopathies épithéliales: la mucoviscidose et autres maladies, Université Paris Descartes, Paris, France.
authors.forename: Aleksander
authors.initials: A
authors.lastname: Edelman
mesh_terms: D000818:Animals; D045744:Cell Line, Tumor; D018118:Chloride Channels / Q000235:genetics*; D017839:Crotalus / Q000235:genetics*; D000242:Cyclic AMP / Q000235:genetics; D003550:Cystic Fibrosis / Q000235:genetics*; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics*; D005260:Female; D006367:HeLa Cells; D006801:Humans; D015640:Ion Channel Gating / Q000235:genetics; D007700:Kinetics; D008297:Male; D051379:Mice; D062105:Molecular Docking Simulation / Q000379:methods; D009154:Mutation / Q000235:genetics; D009865:Oocytes / Q000378:metabolism; D054467:Phospholipases A2 / Q000235:genetics*; D011485:Protein Binding / Q000235:genetics; D017384:Sequence Deletion / Q000235:genetics; D012910:Snake Venoms / Q000235:genetics*; D014982:Xenopus laevis / Q000235:genetics
chemical_list: D018118:Chloride Channels; D012910:Snake Venoms; C093349:cystic fibrosis transmembrane conductance regulator delta F508; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D000242:Cyclic AMP; D054467:Phospholipases A2
keywords: Phospholipase A2 (EC 3.1.1.4); biomolecular interactions; chloride channel; crotoxin; cystic fibrosis; phospholipase A(2)