The formation of the cAMP/protein kinase A-dependent annexin 2-S100A10 complex with cystic fibrosis conductance regulator protein (CFTR) regulates CFTR channel function

Lee A. Borthwick, Jean Mcgaw, Gregory E Conner, Christopher J. Taylor, Volker Gerke, Anil Mehta, Louise Robson, Richmond Muimo

Research output: Contribution to journalArticle

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Abstract

Cystic fibrosis results from mutations in the cystic fibrosis conductance regulator protein (CFTR), a cAMP/protein kinase A (PKA) and ATP-regulated Cl_ channel. CFTR is increasingly recognized as a component of multiprotein complexes and although several inhibitory proteins to CFTR have been identified, protein complexes that stimulate CFTR function remain less well characterized. We report that annexin 2 (anx 2)-S100A10 forms a functional cAMP/PKA/calcineurin (CaN)-dependent complex with CFTR. Cell stimulation with forskolin/3-isobutyl-1-methylxanthine significantly increases the amount of anx 2-S100A10 that reciprocally coimmunoprecipitates with cell surface CFTR and calyculin A. Preinhibition with PKA or CaN inhibitors attenuates the interaction. Furthermore, we find that the acetylated peptide (STVHEILCKLSLEG, Ac1-14), but not the nonacetylated equivalent N1-14, corresponding to the S100A10 binding site on anx 2, disrupts the anx 2-S100A10/CFTR complex. Analysis of 4,4′-diisothiocyanatostilbene-2,2′-disulfonic acid (DIDS) and CFTRinh172-sensitive currents, taken as indication of the outwardly rectifying Cl- channels (ORCC) and CFTR-mediated currents, respectively, showed that Ac1-14, but not N1-14, inhibits both the cAMP/PKA-dependent ORCC and CFTR activities. CaN inhibitors (cypermethrin, cyclosporin A) discriminated between ORCC/CFTR by inhibiting the CFTR inh172-, but not the DIDS-sensitive currents, by >70%. Furthermore, peptide Ac1-14 inhibited acetylcholine-induced short-circuit current measured across a sheet of intact intestinal biopsy. Our data suggests that the anx 2-S100A10/CFTR complex is important for CFTR function across epithelia.

Original languageEnglish
Pages (from-to)3388-3397
Number of pages10
JournalMolecular Biology of the Cell
Volume18
Issue number9
DOIs
StatePublished - Sep 1 2007

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Annexins
Cystic Fibrosis Transmembrane Conductance Regulator
Cyclic AMP-Dependent Protein Kinases
Cystic Fibrosis
1-Methyl-3-isobutylxanthine
Multiprotein Complexes
Peptides
Acids
Calcineurin
Colforsin
Protein Kinase Inhibitors
Cyclosporine
Acetylcholine
Epithelium
Adenosine Triphosphate
Binding Sites
Biopsy
Mutation
Proteins

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics
  • Cell Biology

Cite this

The formation of the cAMP/protein kinase A-dependent annexin 2-S100A10 complex with cystic fibrosis conductance regulator protein (CFTR) regulates CFTR channel function. / Borthwick, Lee A.; Mcgaw, Jean; Conner, Gregory E; Taylor, Christopher J.; Gerke, Volker; Mehta, Anil; Robson, Louise; Muimo, Richmond.

In: Molecular Biology of the Cell, Vol. 18, No. 9, 01.09.2007, p. 3388-3397.

Research output: Contribution to journalArticle

Borthwick, Lee A. ; Mcgaw, Jean ; Conner, Gregory E ; Taylor, Christopher J. ; Gerke, Volker ; Mehta, Anil ; Robson, Louise ; Muimo, Richmond. / The formation of the cAMP/protein kinase A-dependent annexin 2-S100A10 complex with cystic fibrosis conductance regulator protein (CFTR) regulates CFTR channel function. In: Molecular Biology of the Cell. 2007 ; Vol. 18, No. 9. pp. 3388-3397.
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abstract = "Cystic fibrosis results from mutations in the cystic fibrosis conductance regulator protein (CFTR), a cAMP/protein kinase A (PKA) and ATP-regulated Cl_ channel. CFTR is increasingly recognized as a component of multiprotein complexes and although several inhibitory proteins to CFTR have been identified, protein complexes that stimulate CFTR function remain less well characterized. We report that annexin 2 (anx 2)-S100A10 forms a functional cAMP/PKA/calcineurin (CaN)-dependent complex with CFTR. Cell stimulation with forskolin/3-isobutyl-1-methylxanthine significantly increases the amount of anx 2-S100A10 that reciprocally coimmunoprecipitates with cell surface CFTR and calyculin A. Preinhibition with PKA or CaN inhibitors attenuates the interaction. Furthermore, we find that the acetylated peptide (STVHEILCKLSLEG, Ac1-14), but not the nonacetylated equivalent N1-14, corresponding to the S100A10 binding site on anx 2, disrupts the anx 2-S100A10/CFTR complex. Analysis of 4,4′-diisothiocyanatostilbene-2,2′-disulfonic acid (DIDS) and CFTRinh172-sensitive currents, taken as indication of the outwardly rectifying Cl- channels (ORCC) and CFTR-mediated currents, respectively, showed that Ac1-14, but not N1-14, inhibits both the cAMP/PKA-dependent ORCC and CFTR activities. CaN inhibitors (cypermethrin, cyclosporin A) discriminated between ORCC/CFTR by inhibiting the CFTR inh172-, but not the DIDS-sensitive currents, by >70{\%}. Furthermore, peptide Ac1-14 inhibited acetylcholine-induced short-circuit current measured across a sheet of intact intestinal biopsy. Our data suggests that the anx 2-S100A10/CFTR complex is important for CFTR function across epithelia.",
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