Phloxine B Interacts with the Cystic Fibrosis Transmembrane Conductance Regulator at Multiple Sites to Modulate Channel Activity

Cai, Zhiwei; Sheppard, David N.

doi:10.1074/jbc.m108023200

Cited by 52 publications

(65 citation statements)

References 48 publications

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“…The short-term effect of potentiators on mutant CFTR channel activity in detailed patchclamp studies suggests that they may interact directly with the mutant protein (Cai and Sheppard, 2002;Ai et al, 2004). Consistent with this idea, we showed recently that the shortterm addition of 4-methyl-2-(5-phenyl-1H-pyrazol-3-yl)phenol (VRT-532) to isolated membranes expressing the F508del-CFTR protein modified its conformation and susceptibility to trypsin-mediated proteolysis (Wellhauser et al, 2009).…”

Section: Introductionsupporting

confidence: 67%

A Chemical Corrector Modifies the Channel Function of F508del-CFTR

Chiaw¹,

Wellhauser²,

Huan³

et al. 2010

Mol Pharmacol

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The deletion of constitutes the most prevalent cystic fibrosis-causing mutation. This mutation leads to cystic fibrosis transmembrane conductance regulator (CFTR) misfolding and retention in the endoplasmic reticulum and altered channel activity in mammalian cells. This folding defect can however be partially overcome by growing cells expressing this mutant protein at low (27°C) temperature. Chemical "correctors" have been identified that are also effective in rescuing the biosynthetic defect in F508del-CFTR, thereby permitting its functional expression at the cell surface. The mechanism of action of chemical correctors remains unclear, but it has been suggested that certain correctors [including 4-cyclohexyloxy-2-(1-[4-(4-methoxy-benzenesulfonyl)-piperazin-1-yl]-ethyl)-quinazoline (VRT-325)] may act to promote trafficking by interacting directly with the mutant protein. To test this hypothesis, we assessed the effect of VRT-325 addition on the channel activity of F508del-CFTR after its surface expression had been "rescued" by low temperature. It is noteworthy that short-term pretreatment with VRT-325 [but not with an inactive analog, 4-hydroxy-2-(1-[4-(4-methoxy-benzenesulfonyl)-piperazin-1-yl]-ethyl)-quinazoline (VRT-186)], caused a modest but significant inhibition of cAMP-mediated halide flux. Furthermore, VRT-325 decreased the apparent ATP affinity of purified and reconstituted F508del-CFTR in our ATPase activity assay, an effect that may account for the decrease in channel activity by temperature-rescued F508del-CFTR. These findings suggest that biosynthetic rescue mediated by VRT-325 may be conferred (at least in part) by direct modification of the structure of the mutant protein, leading to a decrease in its ATP-dependent conformational dynamics. Therefore, the challenge for therapy discovery will be the design of small molecules that bind to promote biosynthetic maturation of the major mutant without compromising its activity in vivo.

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Section: Introductionsupporting

confidence: 67%

A Chemical Corrector Modifies the Channel Function of F508del-CFTR

Chiaw¹,

Wellhauser²,

Huan³

et al. 2010

Mol Pharmacol

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“…The current findings suggest that it is unlikely that VRT-532 binds at the catalytic site because it does not inhibit the apparent affinity for ATP in this reaction. In agreement with this conclusion, Cai and Sheppard (2002) proposed that phloxine B, another potentiator that increases the channel open time and increases the affinity for ATP in ATP-dependent channel gating, interacted with the protein at a site distinct from the catalytic site. Molecular modeling and mutagenesis studies Zegarra-Moran et al, 2007) also support a model wherein genistein and other potentiators act to stabilize the NBD1-NBD2 dimer interface by binding to a site proximal to, but distinct from, the primary catalytic site.…”

supporting

confidence: 53%

A Small-Molecule Modulator Interacts Directly with ΔPhe508-CFTR to Modify Its ATPase Activity and Conformational Stability

Wellhauser

Chiaw²,

Pasyk³

et al. 2009

Mol Pharmacol

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“…41 Therefore, putative binding sites on the nucleotide-binding domains of CFTR Cl -channels are proposed to be target(s) for many CFTR activators. 42,43 However, our data indicate that the primary mechanism of single CFTR Cl -channel activation by N20 may not involve the nucleotide-binding domains or regulatory domain of CFTR, because these sites have been previously stimulated by ATP and protein kinase A. Therefore, we cannot rule out the possibility that N20 may in some way influence structural changes in the transmembrane domains of CFTR Cl -channels, resulting in channel activation.…”

contrasting

confidence: 38%

Polystyrene nanoparticles activate ion transport in human airway epithelial cells

McCarthy¹,

Gong²,

Nahirney³

et al. 2011

IJN

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Background Over the last decade, nanotechnology has provided researchers with new nanometer materials, such as nanoparticles, which have the potential to provide new therapies for many lung diseases. In this study, we investigated the acute effects of polystyrene nanoparticles on epithelial ion channel function. Methods Human submucosal Calu-3 cells that express cystic fibrosis transmembrane conductance regulator (CFTR) and baby hamster kidney cells engineered to express the wild-type CFTR gene were used to investigate the actions of negatively charged 20 nm polystyrene nanoparticles on short-circuit current in Calu-3 cells by Ussing chamber and single CFTR Clchannels alone and in the presence of known CFTR channel activators by using baby hamster kidney cell patches. Results Polystyrene nanoparticles caused sustained, repeatable, and concentration-dependent increases in short-circuit current. In turn, these short-circuit current responses were found to be biphasic in nature, ie, an initial peak followed by a plateau. EC 50 values for peak and plateau short-circuit current responses were 1457 and 315.5 ng/mL, respectively. Short-circuit current was inhibited by diphenylamine-2-carboxylate, a CFTR Cl − channel blocker. Polystyrene nanoparticles activated basolateral K + channels and affected Cl − and HCO 3 − secretion. The mechanism of short-circuit current activation by polystyrene nanoparticles was found to be largely dependent on calcium-dependent and cyclic nucleotide-dependent phosphorylation of CFTR Cl − channels. Recordings from isolated inside-out patches using baby hamster kidney cells confirmed the direct activation of CFTR Cl − channels by the nanoparticles. Conclusion This is the first study to identify the activation of ion channels in airway cells after exposure to polystyrene-based nanomaterials. Thus, polystyrene nanoparticles cannot be considered as a simple neutral vehicle for drug delivery for the treatment of lung diseases, due to the fact that they may have the ability to affect epithelial cell function and physiological processes on their own.

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Phloxine B Interacts with the Cystic Fibrosis Transmembrane Conductance Regulator at Multiple Sites to Modulate Channel Activity

Cited by 52 publications

References 48 publications

A Chemical Corrector Modifies the Channel Function of F508del-CFTR

A Chemical Corrector Modifies the Channel Function of F508del-CFTR

A Small-Molecule Modulator Interacts Directly with ΔPhe508-CFTR to Modify Its ATPase Activity and Conformational Stability

Polystyrene nanoparticles activate ion transport in human airway epithelial cells

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