ABCB4 p.Thr82Asn
| Predicted by SNAP2: | A: N (61%), C: D (66%), D: D (75%), E: D (75%), F: D (66%), G: D (66%), H: D (66%), I: N (78%), K: D (80%), L: D (63%), M: N (57%), N: D (66%), P: D (80%), Q: D (63%), R: D (80%), S: N (82%), V: D (59%), W: D (71%), Y: D (75%), |
| Predicted by PROVEAN: | A: D, C: D, D: D, E: D, F: D, G: D, H: D, I: D, K: D, L: D, M: D, N: D, P: D, Q: D, R: D, S: D, V: D, W: D, Y: D, |
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[hide] Reversal of advanced fibrosis after long-term urso... Ann Hepatol. 2015 Sep-Oct;14(5):745-51. Frider B, Castillo A, Gordo-Gilart R, Bruno A, Amante M, Alvarez L, Mathet V
Reversal of advanced fibrosis after long-term ursodeoxycholic acid therapy in a patient with residual expression of MDR3.
Ann Hepatol. 2015 Sep-Oct;14(5):745-51., [PMID:26256905]
Abstract [show]
INTRODUCTION: Progressive familial intrahepatic cholestasis type 3 (PFIC-3) is a severe liver disorder associated with inherited dysfunction of multidrug resistance protein 3 (MDR3/ABCB4), which functions as a phospholipid floppase, translocating phosphatidylcholine from the inner to the outer hemileaflet of the canalicular membrane of hepatocytes. MDR3 deficiency results in a disbalanced bile which may damage the luminal membrane of cells of the hepatobiliary system. We evaluated clinical, biochemical and histological improvement in a genetically proven PFIC-3 patient after long-term ursodeoxycholic acid (UDCA) administration. MATERIAL AND METHODS: A PFIC-3 patient and a relative with cholestatic liver disease were studied. Hepatic MDR3 expression was analyzed by immunohistochemistry and ABCB4 mutations were identified. The effect of the mutations on MDR3 expression and subcellular localization was studied in vitro. RESULTS: A 23-year-old man presented cholestasis with severe fibrosis and incomplete cirrhosis. Canalicular staining for MDR3 was faint. Sequence analysis of ABCB4 revealed two missense mutations that reduce drastically protein expression levels. After 9 years of treatment with UDCA disappearance of fibrosis and cirrhosis was achieved. CONCLUSION: These data indicate that fibrosis associated with MDR3 deficiency can be reversed by long-term treatment with UDCA, at least when there is residual expression of the protein.
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No. Sentence Comment
71 Site-directed mutagenesis The substitutions R47Q and T82N were introduced into the plasmid pReceiver-M02-MDR3 (Capital Biosciences, Rockville, MD, USA), which contains the full open reading frame of ABCB4, by site-directed mutagenesis using the QuickChange II system (Stratagene, La Jolla, CA, USA).
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ABCB4 p.Thr82Asn 26256905:71:53
status: NEW72 Primers used for mutagenesis were as follows (top strand shown; mutated nucleotides are in lowercase): R47Q, 5`-CATTGTT- TCaATACTCCGATTGGC-3`; T82N, 5`-GGAGAGAT- GAaTGACAAATTTG-3`.
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ABCB4 p.Thr82Asn 26256905:72:143
status: NEW110 Effects of R47Q and T82N mutations on MDR3 subcellular localization and expression.
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ABCB4 p.Thr82Asn 26256905:110:20
status: NEW120 Genetic analysis Sequence analysis revealed that P1 was a compound heterozygote for two missense mutations in exon 4 of ABCB4: c.140G > A and c.245C > A, (p.R47Q and p.T82N respectively; the first of them located in a cytoplasmic domain and the latter in an extracellular domain).
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ABCB4 p.Thr82Asn 26256905:120:168
status: NEW122 Genetic analysis of P2 showed a missense mutation in ABCB4 shared with her nephew, c.245C>A (p.T82N).
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ABCB4 p.Thr82Asn 26256905:122:95
status: NEW123 In vitro studies of ABCB4 mutations In order to determine whether these substitutions affected subcellular localization or expression of MDR3, MDCKII and HEK293T cells were transfected with expression vectors containing wild-type or the mutated versions of MDR3 (R47Q and T82N) and were analyzed by either immunofluorescence or Western blot.
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ABCB4 p.Thr82Asn 26256905:123:272
status: NEW127 Thus, R47Q and T82N mutations do not alter MDR3 localization, but lead to reduced protein levels.
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ABCB4 p.Thr82Asn 26256905:127:15
status: NEW129 In this disorder, the severity of cholestasis, the time of presentation and the favourable response to UDCA therapy seems to depend on the degree of penetrance of the genetic defect.4,6 The phenotypic characterization of the mutations found in our patient confirms this assumption, since R47Q and T82N do not impair the canalicular expression of MDR3, but they result in a dramatic reduction in the levels of the protein, a finding that correlates with data from the immunohistochemical analysis of the liver biopsy.
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ABCB4 p.Thr82Asn 26256905:129:297
status: NEW131 However, it was reported that mutations at R47 impair phosphorylation of N-terminal domain of MDR3, which is determinant for PC secretion.14 It was reported that R47G mutated protein has similar localization and stability to wild-type protein, but PC secretion activity B A KDa 200 150 100 MDR3 Na/K-ATPase Mock WT R47Q T82N Wild-type R47Q T82N resulted markedly decreased because of this lack of phosphorylation of neighboring residues, either Thr44 or Ser49.
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ABCB4 p.Thr82Asn 26256905:131:320
status: NEWX
ABCB4 p.Thr82Asn 26256905:131:340
status: NEW132 The finding of the mutation T82N in the affected aunt with PFIC3 is a strong evidence of the association of this mutation with the cholestatic disease.
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ABCB4 p.Thr82Asn 26256905:132:28
status: NEW