Permeability of Wild-Type and Mutant Cystic Fibrosis Transmembrane Conductance Regulator Chloride Channels to Polyatomic Anions

Linsdell, Paul; Tabcharani, Joseph A.; Rommens, Johanna M.; Hou, Yue Xian; Chang, Xiu Bao; Tsui, Lap Chee; Riordan, John R.; Hanrahan, John W.

doi:10.1085/jgp.110.4.355

Cited by 201 publications

(236 citation statements)

References 52 publications

(109 reference statements)

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“…These fast exchanging proteins share the common function of transporting relatively large hydrophilic molecules, and consequently it is expected that they should possess a large hydrophilic pore. Because CFTR has been shown to be permeant not only to chloride but also to polyatomic anions (45) and to glutathione (46,47), it certainly shares the same properties. Moreover, the modification of the exchange rate upon phosphorylation implies that it induces a large conformational change that modifies the accessibility of CFTR.…”

Section: Discussionmentioning

confidence: 99%

Phosphorylation-induced Conformational Changes of Cystic Fibrosis Transmembrane Conductance Regulator Monitored by Attenuated Total Reflection-Fourier Transform IR Spectroscopy and Fluorescence Spectroscopy

Grimard

Ramjeesingh

et al. 2004

Journal of Biological Chemistry

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Cystic fibrosis transmembrane conductance regulator (CFTR) is a member of the ABC protein superfamily. Phosphorylation of a regulatory domain of this protein is a prerequisite for activity. We analyzed the effect of protein kinase A (PKA) phosphorylation on the structure of purified and reconstituted CFTR protein. 1 H/ 2 H exchange monitored by attenuated total reflection Fourier transform IR spectroscopy demonstrates that CFTR is highly accessible to aqueous medium. Phosphorylation of the regulatory (R) domain by PKA further increases this accessibility. More specifically, fluorescence quenching of cytosolic tryptophan residues revealed that the accessibility of the cytoplasmic part of the protein is modified by phosphorylation. Moreover, the combination of polarized IR spectroscopy with 1 H/ 2 H exchange suggested an increase of the accessibility of the transmembrane domains of CFTR. This suggests that CFTR phosphorylation can induce a large conformational change that could correspond either to a displacement of the R domain or to long range conformational changes transmitted from the phosphorylation sites to the nucleotide binding domains and the transmembrane segments. Such structural changes may provide better access for the solutes to the nucleotide binding domains and the ion binding site.Cystic fibrosis is caused by a mutation in the membrane chloride channel CFTR 1 (1). CFTR is a member of the ABC superfamily. As the other members of this family, CFTR contains two transmembrane domains and two nucleotide binding domains (NBD) responsible for ATP hydrolysis. In addition to the common ABC structure, CFTR possesses an R domain (regulatory domain) that contains several consensus phosphorylation sites. Phosphorylation of this domain followed by ATP binding and hydrolysis by the NBDs is necessary to induce chloride permeability (2, 3). Yet, an alternative activation mode has been proposed recently, where glutamate induces chloride channel activity in the absence of PKA and ATP (4), but it still needs further investigation.CFTR contains 10 dibasic (R(R/K)X(S/T)) consensus sequences for PKA phosphorylation as well as several monobasic and low affinity sites, most of them located in the R domain (1). Although the overall sequence identity of CFTR R domains is low, the phosphorylation sites are remarkably conserved among species (5). It is believed that the R domain combines both inhibitory and stimulatory effects. Effectively, deletion of the residues 708 -835 from the R domain (6, 7), and even of the residues 760 -783 (8), generates constitutively active channels. Moreover, overexpression or addition of the unphosphorylated R domain, encompassing residues 590 -858, inhibits chloride transport (9). PKA phosphorylation relieves this inhibition. On the other hand, it has been shown that exogenous phosphorylated R domain (either residues 590 -858, 645-834, or 708 -831) increases the open probability of a CFTR channel construct missing residues 708 -835 from the R domain (7,10,11).Nevertheless, the effect of the ph...

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Section: Discussionmentioning

confidence: 99%

Phosphorylation-induced Conformational Changes of Cystic Fibrosis Transmembrane Conductance Regulator Monitored by Attenuated Total Reflection-Fourier Transform IR Spectroscopy and Fluorescence Spectroscopy

Grimard

Ramjeesingh

et al. 2004

Journal of Biological Chemistry

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“…Thus, anion permeation in the CFTR chloride channel shows a weak dependence on the anion's hydration energy (Table 1). 11,13 We observed a nonlinear dependence of the rate constant for anion transport on the concentration of transporter 3 ( Figure S11). This result indicated that anion transport became more efficient when bis-catechol 3 self-associates in the membrane.…”

mentioning

confidence: 93%

Catechols as Membrane Anion Transporters

Berezin

Davis

2009

J. Am. Chem. Soc.

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Compounds that transport ions across cell membranes may function as sensors, signal transducers, or antimicrobials. Synthetic transporters may also help us understand how natural systems move ions across hydrophobic barriers. 1 There is an interest in identifying compounds that transport anions, especially Cl -, across lipid membranes. 2 Reports that catechol 1 binds Cl -in organic solvents prompted us to use this group to form new anion transporters. 3,4 While bacterial siderophores and synthetic analogues use catecholates to move Fe 3+ across membranes, 5 this paper is the first report, to our knowledge, of catechols facilitating transmembrane transport of anions.We report that bis-catechol 3 is an anion transporter whose activity depends on the catechol's substitution and amphiphilicity. We also describe a liposomal assay that allows one to readily measure anion transport selectivity. This assay shows that transport facilitated by 3 follows the Hofmeister sequence, 6 wherein anions that are easier to dehydrate are made more permeable to the membrane by this bis-catechol.ESI-MS analysis showed that a dimer, 1 2 •Cl -, was the major complex formed when TBA + Cl -was mixed with excess catechol (1) ( Figure S2). Based on this finding we attached two catechols, as either 2,3-dihydroxy or 3,4-dihydroxy benzoates, to a TREN scaffold. 7 An alkyl amide group was linked to TREN's third position. Syntheses of analogues 2-7 in Chart 1 are described in the Supporting Information (SI). The membrane transport activity of these amphiphiles was first evaluated using an assay with the pH-sensitive HPTS dye. 8 EYPC liposomes filled with NaNO 3 and HPTS were suspended in a buffer containing Na 2 SO 4 as an external electrolyte. Fluorescence measurements monitored changes in intravesicular pH after adding analogues 2-7. Figure 1a indicates that intravesicular alkalinization occurred upon addition of 2-5. Because sulfate is poorly membrane permeable, the increase in intravesicular pH is likely caused by ligand-facilitated NO 3 -efflux coupled with cotransport or diffusion of H + out of the liposome.Analogues 2-5 showed major differences in their ion transport activity ( Figure 1). Bis-catechol 3, with a medium-length alkyl chain, was the most active of the 2,3-catechols, indicating that transport activity depends on the compound's ability to partition into the membrane. Compound 5 with the longer C17 chain likely aggregates in water, limiting its partitioning into the liposomes. Compound 2, with the shortest chain, was the least active analogue. Presumably, the most active analogue 3 does not aggregate too much in water and is hydrophobic enough to partition into liposomes. The bis-catechol's alkyl chain length was not the only structural determinant for transport function. Figure 1b/c shows that the catechol's substitution pattern is also crucial for ion transport. Both 2,3-O-methyl analogue 6 and 3,4-substituted catechol 7 were inactive, indicating that 2,3-OH diols are essential for membrane transport activity. Simple changes i...

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“…A similar model to explain HCO 3 -secretion by the pancreatic duct of non-rodent eutherian mammals has been proposed (Park et al, 2010;Steward et al, 2005). CFTR from eutherian mammals also has a relatively low P HCO3 (Gray et al, 1993;Gray et al, 1990;Linsdell et al, 1997;O'Reilly et al, 2000;Poulsen et al, 1994). However, in the pancreatic duct cells it has been suggested that HCO 3 -secretion occurs via CFTR because there is no mechanism on the basolateral membrane to accumulate Cl -, whereas there is a mechanism for HCO 3 -accumulation (Fernández-Salazar et al, 2004).…”

Section: Discussionmentioning

confidence: 89%

The distribution and expression of CFTR restricts electrogenic anion secretion to the ileum of the brushtail possum, Trichosurus vulpecula

Gill

Bartolo

Demmers

et al. 2011

Journal of Experimental Biology

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SUMMARYIn eutherian mammals, fluid secretion is essential for intestinal function. This is driven by electrogenic Cl -secretion, which involves a NaK2Cl cotransporter (NKCC1) in the enterocyte basolateral membrane and the cystic fibrosis transmembrane conductance regulator (CFTR) in the apical membrane. However, in the possum ileum, NKCC1 expression is low and secretagogues stimulate electrogenic HCO 3 -secretion driven by a basolateral NaHCO 3 cotransporter (pNBCe1). Here we investigated whether electrogenic anion secretion occurs in possum duodenum and jejunum and determined the role of CFTR in possum intestinal anion secretion. Prostaglandin E 2 (PGE 2 ) and forskolin stimulated a large increase in ileal short-circuit current (I sc ), consistent with electrogenic HCO 3 -secretion, but had little effect on the duodenal and jejunal I sc . Furthermore, 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB) and N-(2-naphthalenyl)-[(3,5-dibromo-2,4-dihydroxyphenyl)methylene]glycine hydrazide (GlyH101) inhibited cloned possum CFTR in cultured cells and the PGE 2 -stimulated ileal I sc , implicating CFTR in ileal HCO 3 -secretion. Consistent with this, CFTR is expressed in the apical membrane of ileal crypt and lower villous cells, which also express pNBCe1 in the basolateral membrane. In contrast, duodenal and jejunal CFTR expression is low relative to the ileum. Jejunal pNBCe1 expression is also low, whereas duodenal and ileal pNBCe1 expression are comparable. All regions have low NKCC1 expression. These results indicate that cAMP-dependent electrogenic Cl -secretion does not occur in the possum small intestine because of the absence of CFTR and NKCC1. Furthermore, CFTR functions as the apical anion conductance associated with HCO 3 -secretion and its distribution limits electrogenic HCO 3 -secretion to the ileum.

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Permeability of Wild-Type and Mutant Cystic Fibrosis Transmembrane Conductance Regulator Chloride Channels to Polyatomic Anions

Cited by 201 publications

References 52 publications

Phosphorylation-induced Conformational Changes of Cystic Fibrosis Transmembrane Conductance Regulator Monitored by Attenuated Total Reflection-Fourier Transform IR Spectroscopy and Fluorescence Spectroscopy

Phosphorylation-induced Conformational Changes of Cystic Fibrosis Transmembrane Conductance Regulator Monitored by Attenuated Total Reflection-Fourier Transform IR Spectroscopy and Fluorescence Spectroscopy

Catechols as Membrane Anion Transporters

The distribution and expression of CFTR restricts electrogenic anion secretion to the ileum of the brushtail possum, Trichosurus vulpecula

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