F508del-CFTR increases intracellular Ca2+ signaling that causes enhanced calcium-dependent Cl− conductance in cystic fibrosis

Institut für Physiologie, Universität Regensburg, Universitätsstraße 31, D-93053 Regensburg, Germany.
Biochimica et Biophysica Acta (Impact Factor: 4.66). 08/2011; 1812(11):1385-92. DOI: 10.1016/j.bbadis.2011.08.008
Source: PubMed

ABSTRACT In many cells, increase in intracellular calcium ([Ca(2+)](i)) activates a Ca(2+)-dependent chloride (Cl(-)) conductance (CaCC). CaCC is enhanced in cystic fibrosis (CF) epithelial cells lacking Cl(-) transport by the CF transmembrane conductance regulator (CFTR). Here, we show that in freshly isolated nasal epithelial cells of F508del-homozygous CF patients, expression of TMEM16A and bestrophin 1 was unchanged. However, calcium signaling was strongly enhanced after induction of expression of F508del-CFTR, which is unable to exit the endoplasmic reticulum (ER). Since receptor-mediated [Ca(2+)](i) increase is Cl(-) dependent, we suggested that F508del-CFTR may function as an ER chloride counter-ion channel for Ca(2+). This was confirmed by expression of the double mutant F508del/G551D-CFTR, which remained in the ER but had no effects on [Ca(2+)](i). Moreover, F508del-CFTR could serve as a scavenger for inositol-1,4,5-trisphosphate [IP3] receptor binding protein released with IP(3) (IRBIT). Our data may explain how ER-localized F508del-CFTR controls intracellular Ca(2+) signaling.

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Available from: Rainer Schreiber, Sep 26, 2015
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    • "(1) This might be a consequence of the unfolded protein response [57] triggered in cells expressing misfolded proteins that allows them to be cleared by the ER associated degradation machinery [58]. (2) Calumenin might be mobilized from the ER in response to intra-cellular Ca2+ signaling, that has been found to be increased in cells expressing F508del-CFTR [59]. (3) Calumenin might undergo alternative splicing into various isoforms that might in turn translocate to the cytoplasm and nucleus [22], [44]. "
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    ABSTRACT: The cystic fibrosis transmembrane regulator (CFTR) is a cyclic-AMP dependent chloride channel expressed at the apical surface of epithelial cells lining various organs such as the respiratory tract. Defective processing and functioning of this protein caused by mutations in the CFTR gene results in loss of ionic balance, defective mucus clearance, increased proliferation of biofilms and inflammation of human airways observed in cystic fibrosis (CF) patients. The process by which CFTR folds and matures under the influence of various chaperones in the secretory pathway remains incompletely understood. Recently, calumenin, a secretory protein, belonging to the CREC family of low affinity calcium binding proteins has been identified as a putative CFTR chaperone whose biophysical properties and functions remain uncharacterized. We compared hydropathy, instability, charge, unfoldability, disorder and aggregation propensity of calumenin and other CREC family members with CFTR associated chaperones and calcium binding proteins, wild-type and mutant CFTR proteins and intrinsically disordered proteins (IDPs). We observed that calumenin, along with other CREC proteins, was significantly more charged and less folded compared to CFTR associated chaperones. Moreover like IDPs, calumenin and other CREC proteins were found to be less hydrophobic and aggregation prone. Phylogenetic analysis revealed a close link between calumenin and other CREC proteins indicating how evolution might have shaped their similar biophysical properties. Experimentally, calumenin was observed to significantly reduce F508del-CFTR aggregation in a manner similar to AavLEA1, a well-characterized IDP. Fluorescence microscopy based imaging analysis also revealed altered trafficking of calumenin in bronchial cells expressing F508del-CFTR, indicating its direct role in the pathophysiology of CF. In conclusion, calumenin is characterized as a charged protein exhibiting close similarity with IDPs and is hypothesized to regulate F508del-CFTR folding by electrostatic effects. This work provides useful insights for designing optimized synthetic structural correctors of CFTR mutant proteins in the future.
    PLoS ONE 08/2014; 9(8):e104970. DOI:10.1371/journal.pone.0104970 · 3.23 Impact Factor
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    • "Nevertheless, CFTR inhibition did significantly reduce the constriction of WT aorta to KCl. This contrasts with studies in epithelial cells where CFTR activation antagonizes IP3-mediated smooth muscle cell constriction and promotes vasodilation, possibly by hyperpolarizing the endoplasmic reticulum membrane [21], [22]. This dichotomy may originate from the differing resting membrane potentials and intracellular chloride content of the 2 cells types. "
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    ABSTRACT: The F508del mutation of the cystic fibrosis transmembrane conductance regulator (CFTR) is the most common cause of cystic fibrosis (CF). Both CF patients and F508del carriers have decreased blood pressure. While this has been attributed to salt depletion, recent studies have shown F508del expression interferes with smooth muscle cell calcium mobilization. We tested the hypothesis that carriers of the F508del mutation have lower adult blood pressures and reduced aortic contractility without a reduction in circulating blood volume. By radiotelemetry, F508del heterozygous mice had significantly lower arterial pressures than wild-type C57BL/6 controls, with the greatest effect seen at the time of dark-to-light cycle transition (mean difference of 10 mmHg). To replicate the vascular effects of sympathetic arousal, isoproterenol and epinephrine were co-infused, and F508del mice again had significantly reduced arterial pressures. Aortas isolated from F508del heterozygous mice had significantly decreased constriction to noradrenaline (0.9±0.2 versus 2.9±0.7 mN). Inhibition of wild-type CFTR or the inositol triphosphate receptor replicated the phenotype of F508del aortas. CFTR carrier status did not alter circulating blood volume. We conclude the CFTR-F508del mutation decreases aortic contractility and lowers arterial pressures. As a cAMP-activated chloride channel that facilitates calcium mobilization, we speculate wild-type CFTR co-activation during adrenergic receptor stimulation buffers the vasodilatory response to catecholamines, and loss of this compensatory vasoconstrictor tone may contribute to the lower arterial pressures seen in heterozygote carriers of a CFTR-F508del mutation.
    PLoS ONE 05/2014; 9(5):e96756. DOI:10.1371/journal.pone.0096756 · 3.23 Impact Factor
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    • "Conversely, carbachol can amplify the response to VIP by stimulating Ca 2+ -activated high conductance and intermediate conductance K + channels, thereby hyperpolarizing the cell membrane and increasing the driving force for apical anion efflux. cAMP and Ca 2+ signalling during transepithelial transport has been discussed in a recent review article (Kunzelmann & Mehta, 2013), which emphasizes the upregulation of Ca 2+ signalling in CF cells (Antigny et al. 2007; Balghi et al. 2011; Martins et al. 2011) and functional interactions between CFTR and the Ca 2+ -activated Cl − channel TMEM16A (Kunzelmann et al. 2012). Cross-talk and synergy between VIP and cholinergic signalling have been appreciated since 1982 (Schultzberg et al. 1982) and are clearly important, but they do not explain why cholinergic responses are 60% smaller in CF glands than in non-CF glands. "
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    ABSTRACT: cAMP-stimulated anion conductance is defective in cystic fibrosis. The regulatory domain of CFTR, the anion channel protein encoded by the CF gene, possesses an unusually high density of consensus sequences for phosphorylation by PKA (14 in a stretch of <200 amino acids). Thus it is not surprising that CFTR is viewed primarily as a cAMP-stimulated anion channel, and most studies have focused on this mode of activation. However there is growing evidence that CFTR also responds to Ca2+ mobilizing secretagogues and contributes substantially to cholinergic and purinergic responses in native tissues. G protein coupled receptors that signal through Gαq can stimulate CFTR channels by activating Ca2+-dependent adenylyl cyclase and tyrosine kinases, and also by inhibiting PP2A. Here we review evidence for these novel mechanisms of CFTR activation and discuss how they may help explain previous observations.
    The Journal of Physiology 08/2013; 591(21). DOI:10.1113/jphysiol.2013.261909 · 5.04 Impact Factor
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