CFTR: Effect of ICL2 and ICL4 amino acids in close spatial proximity on the current properties of the channel

Abstract

These results highlight the critical role of these ICL residues in the assembly of the different domains and/or in the Cl(-) permeation pathway of CFTR.

An Electrostatic Interaction at the Tetrahelix Bundle Promotes Phosphorylation-dependent Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) Channel Opening

“…This difference may be attributable to the fact that a whole cell current measurement is a less direct readout of CFTR channel activity. Interestingly, Billet et al (27) did report that a double mutant in which the charges were swapped between these sites (E267R/K1060E) behaved like wild type CFTR in their assay, which is consistent with an electrostatic interaction between these residues that impacts CFTR activity. In sum, the present findings and these previous reports are in general agreement that E267 and K1060 play an important role in controlling CFTR channel activity that likely involves at least in part an electrostatic interaction between these residues.…”

“…This moderate degree of inhibition by the uncharged K1060T substitution is consistent with our findings. In a brief report Billet et al (27) observed that E267 and K1060 mutations reduced macroscopic (whole cell) CFTR currents in HEK-293 cells without obviously affecting protein maturation. These authors observed greater inhibition by a charge-reversal mutation than by a neutral substitution at the E267 position (E267R versus E267A), as we showed here in our more detailed mechanistic study.…”

“…These authors observed greater inhibition by a charge-reversal mutation than by a neutral substitution at the E267 position (E267R versus E267A), as we showed here in our more detailed mechanistic study. On the other hand, Billet et al (27) reported that a neutral mutation at the K1060 position (A) reduced whole cell CFTR currents to about the same degree as a charge-reversal mutation (E). The apparent lack of charge-dependence for the single K1060 mutants differs from the results of our excised patch experiments.…”

“…The importance of the predicted tetrahelix bundle and the putative E267-K1060 interaction in particular for CFTR gating is not clear. There is a brief report that mutating E267 or K1060 inhibited macroscopic CFTR-mediated currents in transfected HEK-293 cells without apparently reducing CFTR protein expression but the mechanism for this effect was not explored (27). To explore the importance of the predicted E267-K1060 interaction and more generally the tetrahelix bundle in CFTR gating we perturbed the electrostatic interactions between these positions under a variety of conditions.…”

Functional Architecture of the Cytoplasmic Entrance to the Cystic Fibrosis Transmembrane Conductance Regulator Chloride Channel Pore

“…1B). Early homology modeling suggested that the TM extensions that make up the ICLs may form a narrow central "funnel" that forms a pathway for Cl Ϫ movement between the cytoplasm and the transmembrane pore formed by the TMs (9,10). However, such a funnel model was not supported by experiments investigating the functional effects of mutations within the ICLs on Cl Ϫ permeation (11,12).…”