ABCMdb

PMID: 10564718

Pollet JF, Van Geffel J, Van Stevens E, Van Geffel R, Beauwens R, Bollen A, Jacobs P
Expression and intracellular processing of chimeric and mutant CFTR molecules.
Biochim Biophys Acta. 2000 Jan 3;1500(1):59-69., [PubMed]

Sentences

No. Mutations Sentence Comment

6 ABCC7 p.Arg555Lys In addition, while the NBD1 R555K mutation is known to partially correct the processing of CFTR vF508 and to increase activity of both wild-type and vF508 individual channels, it showed no positive effect when introduced into the double NBD1 chimera. Login to comment 27 ABCC7 p.Asp836* Nevertheless, the CFTR truncation mutant (D836X), which contains the 'rst MSD, NBD1, and the R domain, is capable of forming regulated Cl3 channels when expressed in HeLa cells [22] and the protein was shown to cosediment with mature CFTR on a sucrose gradient, suggesting that this 'rst half of the CFTR forms homodimers. Login to comment 127 ABCC7 p.Arg555Lys Lanes: 1 = CHO-mock-transfected, 2 = CHO-CFTR WT, 3 = CHO-CFTR 4M, 4 = CHO-CFTR R555K, 5 = CHO-vNBD2 and 6 = T84 cell line as positive control (derived from human gastrointestinal tract and expressed naturally CFTR protein). Login to comment 128 ABCC7 p.Arg555Lys (B) Iodide e¥ux assays were performed on con£uent CHO cells (as negative control) grown on a plastic support and on CHO cells expressing CFTR WT, CFTR 4M, CFTR 4M-vNBD2 and CFTR 4M-R555K molecules. Login to comment 134 ABCC7 p.Arg555Lys Data points are means of triplicates with S.E.M. Table 2 Expression, maturation and activity of the CFTR variants used in this study CFTR variant Protein Function A B C Wild-type + + + + vF508 3 + 3 3 4M + + + + 4MN + + + + 4M-2UNBD1 3 + 3 3 4MN-2UNBD1 3 + 3 3 4M-R555K 3 + + + 4M-2U(NBD1+R555K) 3 + 3 3 4M-NBD2/NBD1 3 + 3 3 4MN-NBD2/NBD1 3 + 3 3 4M-2UNBD2 3 + 3 3 4MN-2UNBD2 3 + 3 3 4MvNBD1 + + 3 3 4MvNBD2 + + + 3 + indicates the presence and 3 the absence of a CFTR band of the size expected for the A, B and C forms in transiently expressing COS-1 cells. Login to comment 138 ABCC7 p.Arg555Lys To further determine whether the misfolding of 2UNBD1 molecule resulted from the additive e¡ect of the NBD1 intrinsic instability, we looked for a reversion of this phenotype by the R555K mutation. Login to comment 140 ABCC7 p.Arg555Lys The R555K mutation was then introduced into each NBD1 of the 2UNBD1 variant. Login to comment 142 ABCC7 p.Arg555Lys Fig. 3, lane 15, shows that the R555K mutation did not reverse the processing block observed for the 2UNBD1 molecule, suggesting that this defect did not result from an additive e¡ect of the intrinsic NBD1 instability. Login to comment 159 ABCC7 p.Arg555Lys In addition, the R555K mutation, known to reverse the processing abnormality due to the vF508 mutation, did not restore the processing of the 2UNBD1 variant. Login to comment