ABCC7 p.Lys68Arg
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PMID: 24777605
[PubMed]
Lee S et al: "Interference with ubiquitination in CFTR modifies stability of core glycosylated and cell surface pools."
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5
N-tail mutants, K14R and K68R, lead to increased mature band C CFTR, which can be augmented by proteasomal (but not lysosomal) inhibition, allowing trafficking to the surface.
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ABCC7 p.Lys68Arg 24777605:5:25
status: NEW73 We hypothe- TABLE 1 Primers for site-directed mutagenesis Mutation Directiona Sequence (5=-3=)b K14R F GGCCAGCGTTGTCTCCAGACTTTTTTTCAGCTGGACC R GGTCCAGCTGAAAAAAAGTCTGGAGACAACGCTGGCC K68R F GGCTTCAAAGAAAAATCCTAGACTCATTAATGCCCTTCGGCG R CGCCGAAGGGCATTAATGAGTCTAGGATTTTTCTTTGAAGCC K710R F CCAATCAACTCTATACGAAGATTTTCCATTGTGCAAAAG R CTTTTGCACAATGGAAAATCTTCGTATAGAGTTGATTGG K716R F GAAAATTTTCCATTGTGCAAAGGACTCCCTTACAAATGAATGG R CCATTCATTTGTAAGGGAGTCCTTTGCACAATGGAAAATTTTC K710/716R F CCAATCAACTCTATACGAAGATTTTCCATTGTGCAAAGGACTCCCTTACAAATGAATGG R CCATTCATTTGTAAGGGAGTCCTTTGCACAATGGAAAATCTTCGTATAGAGTTGATTGG K1041R F CCTCACAGCAATTCAGACAACTGGAATCTGAAG R CTTCAGATTCCAGTTGTCTGAGTTGCTGTGAGG K1080R F GAAACTCTGTTCCACAGAGCTCTGAATTTACATAC R GTATGTAAATTCAGAGCTCTGTGGAACAGAGTTTC K1218R F GATCTCACAGCAAGATACACAGAAGG R CCTTCTGTGTATCTTGCTGTGAGATC a F, forward; R, reverse.
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ABCC7 p.Lys68Arg 24777605:73:181
status: NEW97 Lysine-to-arginine mutations caused minor changes in CFTR conformation, except for the K68R mutation which caused changes in the orientation of third alpha helix, where 68R is located (Fig. 3A).
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ABCC7 p.Lys68Arg 24777605:97:87
status: NEW99 K14R and K68R mutations result in increased levels of immature (A/B band) and mature (C band) forms of CFTR expression (Fig. 2B).
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ABCC7 p.Lys68Arg 24777605:99:9
status: NEW117 To test this hypothesis for K14R and K68R, we expressed these N-tail mutants in the presence and absence of proteasomal inhibition.
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ABCC7 p.Lys68Arg 24777605:117:37
status: NEW119 4A), but proteasome inhibition during expression of the K14R and K68R mutant CFTR cDNAs led to even higher expression levels.
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ABCC7 p.Lys68Arg 24777605:119:65
status: NEW130 This immunoblot demonstrates increased levels of band C CFTR expression from the K14R, K68R, and K1218R cDNA vectors and decreased levels of band C CFTR expression in K710R, K716R, K710/716R, and K1080R vectors.
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ABCC7 p.Lys68Arg 24777605:130:87
status: NEW135 However, unexpectedly, expression levels in K14R- and K68R- transfected cells were not increased by lysosomal inhibition, perhaps because they were already slow to recycle.
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ABCC7 p.Lys68Arg 24777605:135:54
status: NEW136 Densitometry analysis showed that a proteasomal inhibitor increased C bands of wt, K14R, and K68R compared to control levels, but changes in A/B bands were insignificant.
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ABCC7 p.Lys68Arg 24777605:136:93
status: NEW137 Lysosomal inhibitor did not change C bands, but A/B bands of K14R and K68R were decreased compared to control levels.
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ABCC7 p.Lys68Arg 24777605:137:70
status: NEW138 These data suggest that K14R and K68R mutations allow CFTR to bypass the lysosome for degradation so that preventing lysosomal activity does not further increase the expression level of the mutant compared to wild-type CFTR.
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ABCC7 p.Lys68Arg 24777605:138:33
status: NEW144 A K-to-R mutation does not alter the conformational structure of each domain except for the K68R mutant.
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ABCC7 p.Lys68Arg 24777605:144:92
status: NEW163 (A) The results from expression of control vector, wt CFTR, K14R, and K68R with and without proteasomal or lysosomal inhibition are shown in representative blots.
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ABCC7 p.Lys68Arg 24777605:163:70
status: NEW164 MG132 increased total A/B/C bands of CFTR in wild-type, K14R, and K68R proteins.
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ABCC7 p.Lys68Arg 24777605:164:66
status: NEW200 There were decreased levels of cell surface CFTR detection in the K68R, K710R, K716R, K710/ K716R, and K1041R mutant forms of CFTR, but the wild-type, K14R, K1080R, and K1218R mutants accumulated similar levels of visible cell surface CFTR.
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ABCC7 p.Lys68Arg 24777605:200:66
status: NEW201 Analysis of the Z stack of images for the wild type, K68R, K710R, K1080R, and K1218R confirmed that the CFTR immunostaining, shown in red, resided at the apical surface in these nonpermeabilized cells (data not shown).
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ABCC7 p.Lys68Arg 24777605:201:53
status: NEW207 Again, there was a reduction in immunodetectable CFTR (K14R, K68R, K710R, K716R, K710/716R, and K1080R forms) in cell surface membranes compared to wild-type levels.
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ABCC7 p.Lys68Arg 24777605:207:61
status: NEW218 Control vector, wt CFTR, K14R, K68R, K710R, K716R, K710/716R, K1041R, K1080R, and K1218R were expressed in IB3-1 cells for 48 h. CFTR was immunoprecipitated (IP) with M3A7 as described in Materials and Methods and separated by SDS-PAGE.
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ABCC7 p.Lys68Arg 24777605:218:31
status: NEW221 K63-linked ubiquitin on CFTR is reduced in the K68R, K710R, K716R, and K1218R mutants.
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ABCC7 p.Lys68Arg 24777605:221:47
status: NEW253 A lighter signal is visible from cells expressing K68R and the R domain mutants.
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ABCC7 p.Lys68Arg 24777605:253:50
status: NEW275 Since ZO-1 interacts with PDZ domains, the reduced interactions in K14R, K68R, K710R, K716R, K710/716R, and K1041R and the absence of interactions with K1080R and K1218R raise the issue of alternative trafficking routes that are not dependent on the C-tail PDZ domain.
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ABCC7 p.Lys68Arg 24777605:275:73
status: NEW279 K14R and K68R experience some proteasome degradation, but a sizeable fraction is localized near the plasma membrane.
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ABCC7 p.Lys68Arg 24777605:279:9
status: NEW296 Interestingly, ZO-1 was highly abundant in complexes formed with wt CFTR; K14R, K68R, K710R, K716R, K710/716R, and K1041R mutants bound less ZO-1, whereas K1080R and K1218R mutant complexes had none.
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ABCC7 p.Lys68Arg 24777605:296:80
status: NEW