ABCMdb

PMID: 9922375

Sheppard DN, Welsh MJ
Structure and function of the CFTR chloride channel.
Physiol Rev. 1999 Jan;79(1 Suppl):S23-45., [PubMed]

Sentences

No. Mutations Sentence Comment

98 ABCC7 p.Arg347Cys ABCC7 p.Arg334Gln These residues are conserved across spe- because I0 blocks the pore, it appears that I0 is less per- cies, and two are the site of mutations associated with meable under some conditions, including the whole cell CF: R334Q/W and R347C/H/L/P (142). Login to comment 102 ABCC7 p.Lys335Glu ABCC7 p.Lys95Asp This would suggest that like acidic residues (K95D and K335E) altered the whole cell other Cl0 channels, including ligand-gated Cl0 channels in anion permeability sequence by converting CFTR from a neurons (20) and outwardly rectifying Cl0 channels in low I0 permeability pore (Br0 ' Cl0 ú I0 ) to a high I0 epithelia (55), the CFTR pore has a ''weak field strength`` permeability pore (I0 ú Br0 ú Cl0 ) (4). Login to comment 104 ABCC7 p.Arg347Glu ABCC7 p.Arg1030Glu Thus, depending on the experimental tants, R347E and R1030E, did not alter the anion perme- conditions and whether block of the channel by I0 is con- ability sequence, although PI/PCl values were increased. Login to comment 112 ABCC7 p.Lys335Cys ABCC7 p.Lys95Cys On mutants K95C and K335C interact with methanethiosulfo- the basis of these data, the minimum diameter of the nate (MTS) reagents, and mutations that eliminate the CFTR pore was estimated to be Ç5.3 A˚ (77), similar to positive charge at K335 reduce single-channel conduc- that reported for other Cl0 channels (10, 20, 55). Login to comment 114 ABCC7 p.Lys335Glu However, the mutation K335E did is controversial (51, 59, 72, 93, 99, 100, 111, 129). Login to comment 118 ABCC7 p.Arg334Trp ABCC7 p.Arg347Pro When expressed in heterologous epithelial cells, the CF-associatedpermeation is determined by the hydration energy of P27-8/ 9j0e$$ja08 01-13-99 14:54:54 prsa APS-Phys Rev mutants R334W and R347P were correctly processed and proteins play an important structural role by kinking a-helices (22). Login to comment 121 ABCC7 p.Arg334Trp ABCC7 p.Arg347Pro Analysis of the single-channel properties of R334W and R347P demonstrated that both mutants decrease sin- or indirectly to the formation of the CFTR pore (119). Login to comment 124 ABCC7 p.Pro99Leu ABCC7 p.Pro99Ala ABCC7 p.Pro99Gly When the basic arginine at this position was re- gle-channel Cl0 conductance in the rank order: wild type ' P99G ú P99L ' P99A (119). Login to comment 128 ABCC7 p.Arg347His ABCC7 p.Pro99Cys However, because P99C did(128) could control the pore properties of the CF-associated mutant R347H simply by manipulating pH. Login to comment 129 ABCC7 p.Arg347His At pH 5.5, not react with MTS reagents (1), P99 probably does not directly line the CFTR pore.when histidine is positively charged, R347H had normal pore properties (128). Login to comment 130 ABCC7 p.Arg347Pro However, at pH 8.7, when histidine Knowledge of the structure and function of the CFTR pore has also emerged from studies using truncated andis uncharged, single-channel conductance was reduced to a similar extent as that of R347P, and the anomalous mole chimeric proteins (95, 116, 134). Login to comment 131 ABCC7 p.Asp836* The amino-terminal portion of CFTR (D836X, which contains MSD1, NBD1 andfraction behavior of CFTR was lost (128). Login to comment 136 ABCC7 p.Asp836* Based on biochemical and functional data, we speculated that D836X generates a Cl0 channel byveats noted above regarding the contribution of residues to the pore versus less specific alterations of structure forming a homomultimer (116). Login to comment 138 ABCC7 p.Asp836* To identify residues that line the CFTR pore and make below), then in a D836X multimer MSD1 sequences may substitute for MSD2 sequences to form the pore. Login to comment 145 ABCC7 p.Arg352Cys The voltage dependence of the reaction rates of MTS reagents with CFTR (Br0 Å Cl0 ú I0 ), Xenopus CFTR (Br0 Å I0 ú Cl0 ), and human-Xenopus CFTR chimeras in which eithercysteine mutations at these residues and single-channel studies of wild-type CFTR and the mutant R352C suggest MSD1 or MSD2 of human CFTR was replaced with the equivalent region of Xenopus CFTR (hX1-6, Br0 Å I0 úthat the selectivity of CFTR for anions over cations is determined by a site located at the intracellular end of the pore Cl0 , and hX7-12, Br0 ú Cl0 ú I0 , respectively) also suggest that sequences in MSD1 likely determine the anionthat involves the residue R352 (32, 52). Login to comment 148 ABCC7 p.Arg352Cys ABCC7 p.Gln353Cys ABCC7 p.Thr351Cys Therefore, other sequences must account for the differ-end of the pore and R352C is located closer to the extracellular end of the pore than either T351C or Q353C (32). Login to comment 156 ABCC7 p.Arg117Cys First, incorporation of glycosylation sequences into ICL1, ICL3, and ICL4 disrupted protein associated mutations (R117C/H/L/P) (142). Login to comment 157 ABCC7 p.Arg117His When expressed in heterologous epithelial cells, R117H was cor-folding, and there was little Cl0 channel function measured (27). Login to comment 159 ABCC7 p.Arg117His In addition, R117H had an altered sensitivity to external pH, suggesting thatof human CFTR (36, 149). Login to comment 161 ABCC7 p.Arg117His R117H caused a small decrease in single-channel conductance but a dramatic change informs a cAMP-stimulated Cl0 current (46, 58, 61). Login to comment 168 ABCC7 p.Ser945Leu ABCC7 p.Gly970Arg Although, Xenopus CFTR hadships of the mutations S945L and G970R in ICL3 showed weak outward rectification in contrast to that of wild-type a pattern of gating similar to human CFTR, the human-Xenopus CFTR chimera hX1-6 showed a pattern of gatingCFTR (114, 148). Login to comment 169 ABCC7 p.Arg117His These results suggest that ICL2 and ICL3 may be located close to the intracellular mouth of the CFTR characterized by a shortened open time, similar to that of R117H (95). Login to comment 176 ABCC7 p.Ile148Thr ABCC7 p.Gly178Arg Furthermore, the mutations I148T and G178R could not be locked open by the nonhydrolyzable ATP analog 5؅- adenylylimidodiphosphate (AMP-PNP) (112), the mutation Information about the CFTR pore has also emerged from studies using Cl0 channel inhibitors. Login to comment 177 ABCC7 p.Arg1066Leu ABCC7 p.Phe1052Val Because DPCA1067T altered the relationship between open probability (Po) and ATP concentration (33), and the response of inhibition of CFTR was voltage dependent and enhanced when the external Cl0 concentration was reduced,R1066L and F1052V to pyrophosphate (PPi) was less than wild type (33). Login to comment 183 ABCC7 p.Thr1134Phe site could be transferred from S341 to S1141 in M12 while another mutation in M12, T1134F, increased the affinityStudies of CFTR variants lacking the consensus glycosylation sequences in extracellular loop (ECL) 4 of DPC block (87). Login to comment 193 ABCC7 p.Arg347Asp Moreover, the mutant R347D significantly weak- main have had little discernible effect on conduction and permeation, although they have frequently had profoundened the binding of DNDS and DIDS to CFTR, suggesting that R347 contributes to the binding site for disulfonic effects on gating behavior (see below). Login to comment 232 ABCC7 p.Arg347Asp Because the mutant R347D significantly weakened the activity of CFTR Cl0 channels by using excised inside-out membrane patches from cells expressing recombinantbinding of DNDS and DIDS to CFTR, R347 likely contributes to this site (74). Login to comment 248 ABCC7 p.Thr338Cys ABCC7 p.Thr339Cys However, because the mutations T338C and T339C did not react with MTS reagents, the side PKA phosphorylation but did not substitute for ATP in opening phosphorylated CFTR Cl0 channels.chains of these residues do not interact with permeating ions (31, 77). Login to comment 375 ABCC7 p.Lys464Ala ABCC7 p.Gln552Ala This is consistent with the finding that in the absence predicted to decrease the rate of hydrolysis at NBD1, for example, K464A and Q552A, would thus increase the in-of ATP, the channel does not open. Login to comment 382 ABCC7 p.His1350Gln In addition, the observation that the H1350Q mutation, which would be predicted to increase the ratechannel. Login to comment 497 ABCC7 p.Ser753Ala The mutation 10SA-S753A eliminates much Additional evidence for phosphorylation-dependent control of CFTR channel activity comes from observa-of the residual PKA-dependent phosphorylation of the 10SA mutant and decreases the Po of the 10SA mutant by tions that there appear to be both high and low phosphorylation states correlating with a high and low Po (43, 64).40% (115). Login to comment 504 ABCC7 p.Asp836* First, those sites were phosphor- interference between R domains in a D836X homomultimer.ylated in vivo after stimulation with cAMP agonists. Login to comment 521 ABCC7 p.Ser660Ala When S660 in CFTRDR was mutated to alanine dan (42) proposed a two-domain model of the R domain: RD1 (amino acids 587-672) and RD2 (amino acids 679-(CFTRDR-S660A), PKA failed to stimulate channel activity. Login to comment 526 ABCC7 p.Ser660Ala ABCC7 p.Asp836* In addition, deletion studies of portions of the R domain suggest thatout that PKA cannot interact with CFTRDR-S660A, even though it does interact with CFTRDR. there are two halves to the R domain with the first conserved in other ABC transporters and the second uniqueLike CFTRDR, the amino-terminal portion of CFTR (D836X, which contains MSD1, NBD1, and the R domain) to CFTR (102, 103). Login to comment 528 ABCC7 p.Asp836* The activation of D836X Cl0 channels by PKA indicates that E. A Model of R Domain Functionthe MSD1-NBD1 motif must contain many of the sequences with which the R domain interacts to control CFTR. Login to comment 529 ABCC7 p.Asp836* The PKA-independent activity of D836X Cl0 chan- The R domain was originally proposed to regulate CFTR by keeping the channel closed at rest. Login to comment 533 ABCC7 p.Ser660Ala ABCC7 p.Asp836* Alternatively, the PKA-independent activity of D836X could be due to As described above, phosphorylation by PKA, deletion of FIG. 11 Effect of a recombinant R domain (R1, residues 645-834) on CFTRDR/S660A channel in presence of PKA. Login to comment 535 ABCC7 p.Ser660Ala B and C: effect of R1 on rates of entry into and exit from burst for CFTR-DR/S660A channels in presence of PKA. Login to comment