ABCB4 p.Thr201Met
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PMID: 24594635
[PubMed]
Gordo-Gilart R et al: "Functional analysis of ABCB4 mutations relates clinical outcomes of progressive familial intrahepatic cholestasis type 3 to the degree of MDR3 floppase activity."
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9
Results Eight distinct ABCB4 mutations were identified: one nonsense, one splicing and six missense mutations, four of which (G68R, T201M, P479L, D459H) affected MDR3 expression level.
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ABCB4 p.Thr201Met 24594635:9:132
status: NEW11 Phosphatidylcholine efflux assays indicated that T201M, P479L, S978P and E1118K mutations impaired MDR3 activity to variable degrees.
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ABCB4 p.Thr201Met 24594635:11:49
status: NEW42 Plasmids and site-directed mutagenesis The plasmid pReceiver-M02-MDR3, containing the full open reading frame of ABCB4 (Capital Biosciences, Rockville, Maryland, USA) was used as a template for introducing the substitutions G68R, T201M, P352L, D459H, S978P and E1118K, by site-directed mutagenesis using the QuickChange II system (Agilent Technologies, Santa Clara, California, USA).
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ABCB4 p.Thr201Met 24594635:42:230
status: NEW110 Mutants T201M, P479 L, S978P and E1118 K exhibited surface expression comparable to that of wild-type MDR3.
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ABCB4 p.Thr201Met 24594635:110:8
status: NEW117 MDR3 was detected as a 140-150 kDa band, Table 2 ABCB4 mutations and MDR3 immunohistochemical analysis Mutation allele 1 Mutation allele 2 Patient n Nucleotide change Predicted effect Nucleotide change Predicted effect MDR3 staining 1 c.3559C>T p.R1187X c.3633+2 T>A Splicing defect ABSENT 2 c.202G>A p.G68R c.202G>A p.G68R ABSENT 3 c.202G>A p.G68R c.202G>A p.G68R NA 4 c.1375G>C p.D459H c.1436C>T p.P479L FAINT 5 c.602C>T p.T201M c.3352G>A p.E1118K NORMAL 6 c.2932T>C p.S978P ND NORMAL Immunostaining was carried out on liver biopsy specimens in patients 2-6 and in hepatectomy specimens in patient 1.
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ABCB4 p.Thr201Met 24594635:117:425
status: NEW121 (B) Normal canalicular staining in the tissue of patient 5, who is compound heterozygous for two missense mutations (T201M and E1118K).
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ABCB4 p.Thr201Met 24594635:121:117
status: NEW128 By contrast, expression levels of mutants G68R and D459H were markedly decreased (15% and 20% of wild-type, respectively), consistent with the fact that ER retention usually leads to a premature degradation of the proteins.32 T201M and P479L mutations also caused a significant reduction in the expression of MDR3 (60% and 65% of wild-type, respectively).
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ABCB4 p.Thr201Met 24594635:128:226
status: NEW138 The T201M, P479L and E1118K mutations reduced the release of labelled PC in the presence of the bile salt.
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ABCB4 p.Thr201Met 24594635:138:4
status: NEW140 At the time point of 3 h, the percentage of [3 H]-choline phospholipids effluxed in cells expressing mutants T201M, P479L and E1118K was 27%, 18% and 39% of that in cells expressing the wild-type protein, respectively (figure 4B).
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ABCB4 p.Thr201Met 24594635:140:109
status: NEW159 It seems likely that these mutations lead to misfolded proteins which, typically, exhibit abnormal trafficking and are prematurely degraded by the ER-associated protein degradation.32 This would raise the possibility that these mutants could be rescued to the cell surface by pharmacological chaperones, as has been shown for various other misfolded mutant membrane proteins.37 38 The T201M, P479L, S978P and E1118K mutations affect PC-translocating activity of MDR3, but to different extents.
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ABCB4 p.Thr201Met 24594635:159:385
status: NEW162 (A) Mock-transfected AD-293 cells (control) and cells transfected with wild-type ABCB4 and mutant T201M were stained for MDR3 and analysed by confocal immunofluoresce microscopy.
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ABCB4 p.Thr201Met 24594635:162:98
status: NEW186 binding and ATP hydrolysis.43 Of note, the equivalent residue in mouse Abcb1, S975, directly interacts with specific substrates, as revealed by crystallographic studies.41 Furthermore, a number of mutations within this transmembrane helix has been shown to dramatically affect ATPase activity of ABCB1.44 These observations concur with the complete loss of function of MDR3 caused by mutation S978P .
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ABCB4 p.Thr201Met 24594635:186:45
status: NEW187 PC-translocation activity of the MDR3 mutant T201M was 27% of reference activity for the wild-type MDR3.
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ABCB4 p.Thr201Met 24594635:187:45
status: NEW190 It must be taken into account that the T201M and P479L mutations also lowered the MDR3 protein levels, although the extent to which these levels were decreased did not parallel with the reductions in MDR3 activity obtained in the PC efflux assays.
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ABCB4 p.Thr201Met 24594635:190:39
status: NEW189 It must be taken into account that the T201M and P479L mutations also lowered the MDR3 protein levels, although the extent to which these levels were decreased did not parallel with the reductions in MDR3 activity obtained in the PC efflux assays.
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ABCB4 p.Thr201Met 24594635:189:39
status: NEW