ABCMdb

ABCB4 p.Ile541Phe

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Publications

PMID: 21567408 [PubMed] Nakagawa H et al: "Ubiquitin-mediated proteasomal degradation of ABC transporters: a new aspect of genetic polymorphisms and clinical impacts."

No. Sentence Comment

155 Effect of Mutations and Nonsynonymous SNPs on Protein Trafficking, Maturation, or ERAD of ABC Transporters Protein AA Mutation/SNP Effect on Protein Reference ABCA1 W590S Mutation Functional defect 115 R587W Mutation Impaired glycol processing 115 Q597R Mutation Impaired glycol processing, ERAD 115,116 Y1532C Mutation Altered protein trafficking 117 R1925Q Mutation Altered protein trafficking 118 ABCA3 R43L Mutation Altered protein trafficking 119 L101P Mutation Altered protein trafficking 119 R280C Mutation Altered protein trafficking 119 ABCA4 L541P Mutation Mislocalization 120 R602W Mutation Mislocalization 120 A1038V Mutation Mislocalization 120 C1490Y Mutation Mislocalization 120 ABCB1a G268V Mutation ERAD 121 G341C Mutation ERAD 121 I1196S Mutation Reduced glycosylation 122 ABCB4 I541F Mutation Accumulation in ER 123 ABCB11a E135K Mutation Reduced level of mature protein 124 L198P Mutation Reduced level of mature protein 124 E297G Mutation Reduced level of mature protein 124 L413W Mutation Reduced level of mature protein 124 R432T Mutation Reduced level of mature protein 124 D482G Mutation Immature protein in ER 124,125 N490D Mutation Reduced level of mature protein 124 A570T Mutation Reduced level of mature protein 124 T655I Mutation Reduced level of mature protein 124 Y818F SNP Moderate reduction of protein 124 G982R Mutation Retention in ER 125 R1153C Mutation ERAD 125 R1286Q Mutation Retention in ER 125 ABCC2a R768W Mutation Impaired protein trafficking 126 I1173F Mutation Impaired protein maturation 127 R1392 Mutation Impaired protein maturation 128 M1393 Mutation Impaired protein maturation 129 ABCC4a E757K SNP Altered protein trafficking 23 ABCC7 F508 Mutation Misfolding, ERAD 36-39,130 G85E Mutation Impaired protein maturation 130-132 G91R Mutation Impaired protein maturation 130-132 N1303K Mutation Impaired protein maturation 130-132 ABCC8 WT Wild type Ubiquitin-proteasome degradation 133 A116P Mutation Ubiquitin-proteasome degradation 133 V187D Mutation Ubiquitin-proteasome degradation 133 F1388 Mutation Impaired protein trafficking 134 L1544P Mutation Impaired protein trafficking 135,136 ABCC11a G180R SNP Ubiquitin-proteasome degradation 50 27 Mutation Ubiquitin-proteasome degradation 50 ABCG2a V12M SNP Altered protein localization 96 Q141K SNP Ubiquitin-proteasome degradation 102 F208S SNP Ubiquitin-proteasome degradation 78,99 S441N SNP Ubiquitin-proteasome degradation 78,99 Mutations of ABCA1, ABCA3, ABCA4, ABCB4, ABCB11, ABCC2, ABCC7 (CFTR), and ABCC8 are associated with Tangier disease, fatal surfactant deficiency, Stargardt disease, progressive familial intrahepatic cholestasis type 3 (PFIC-3), progressive familial intrahepatic cholestasis type 2 (PFIC-2), Dubin-Johnson syndrome, cystic fibrosis, and familial hyperinsulinism, respectively. Login to comment

PMID: 19185004 [PubMed] Delaunay JL et al: "A missense mutation in ABCB4 gene involved in progressive familial intrahepatic cholestasis type 3 leads to a folding defect that can be rescued by low temperature."

No. Sentence Comment

133 These expression experiments show that mutation I541F causes a trafficking defect both in ABCB1 and ABCB4, and that ABCB1-GFP chimera provides an interesting model to further investigate the effect of I541F mutation. Login to comment
4 Expression studies showed that ABCB4 was localized at the bile canalicular membrane in HepG2 cells and at the apical surface in MDCK cells, whereas the I541F mutant was intracellular. Login to comment
5 In MDCK cells, ABCB1-I541F also accumulated intracellularly in compartments, which were identified as the endoplasmic reticu- lumandcis-Golgi,andremainedpartiallyendoH-sensitive.Aftershiftingcellsto27°C,ABCB1-I541F wasexpressedattheapicalcellsurfaceinamatureandactiveform.Similarly,ABCB4wassignificantly trafficked to the membrane of bile canaliculi in HepG2 cells. Login to comment
6 Conclusion: Mutation I541F causes mislocalizationofbothABCB4andABCB1.IntracellularretentionofABCB4-I541Fcanexplainthe disease in PFIC3 patients bearing this mutation. Login to comment
31 We have studied the effect of mutation I541F, located in the first nucleotide-binding domain (NBD), which has been described in a homozygous patient with PFIC3.4 This female patient experienced chronic cholestasis since 1 year of age and developed cirrhosis. Login to comment
100 Results Expression of ABCB4 and ABCB4-I541F in HepG2 and MDCK Cells. Login to comment
102 To examine the cellular localization of ABCB4 and ABCB4-I541F, HepG2 cells were transiently transfected with the pcDNA3 plasmid encoding wild-type ABCB4 or ABCB4-I541F or with the empty plasmid. Login to comment
105 Cells transfected with the plasmid encoding ABCB4 expressed the protein exclusively on the membranes of bile canaliculi, which correspond to the apical surface (Fig. 1A), consistent with the expected canalicular localization of ABCB4 in hepatocytes.15 By contrast, cells transfected with the plasmid encoding ABCB4-I541F did not express the protein at the membrane of bile canaliculi. Login to comment
109 MDCK cells were transfected with the plasmids encoding ABCB4 and ABCB4-I541F or the empty plasmid. Login to comment
114 By contrast, ABCB4-I541F was detected around the nuclei (Fig. 1D). Login to comment
116 These expression studies show that ABCB4 is expressed at the apical membrane, whereas ABCB4-I541F leads to a major trafficking defect and accumulates within the cell cytoplasm, in both HepG2 and MDCK cells. Login to comment
117 ABCB1-I541F-GFP Is Also Intracellular. Login to comment
118 The I541F mutation is localized next to the ABC signature (LSSGGQ) in the first nucleotide-binding domain. Login to comment
123 Expression of ABCB4 and ABCB4-I541F in HepG2 and MDCK cells. Login to comment
124 HepG2 cells transiently expressing ABCB4 (A) or ABCB4-I541F (B), and filter-grown MDCK cells stably expressing ABCB4 (C, E) or ABCB4-I541F (D, F) were fixed with methanol/acetone and processed for immunofluorescence using the P3II-26 monoclonal antibody and Alexa 488-conjugated anti-mouse immunoglobulin G. Login to comment
127 Bars, 10 ␮m. activity.20 In MDCK cells, ABCB1-GFP was exclusively expressed at the apical surface (Fig. 3A), as already reported.24 In contrast, ABCB1-I541F-GFP was intracellular, as observed for ABCB4-I541F. Login to comment
132 ABCB1-I541F-GFP accumulated around the nuclei, in membrane compartments that appeared sometimes granular (Fig. 3B). Login to comment
134 ABCB1-I541F-GFP Is Retained in an Endoplasmic Reticulum/Golgi Compartment. Login to comment
135 To precisely determine the sites of intracellular accumulation, different cellular compartments were identified with specific antibodies in MDCK/ABCB1-I541F-GFP cells grown to subconfluence. Login to comment
137 In all of these colocalization experiments, ABCB1-I541F-GFP was observed exclusively in the cytoplasm, especially around the nuclei, whereas little ABCB1-GFP was intracellular (Fig. 4A). Login to comment
138 ABCB1-I541F-GFP colocalized strongly with calnexin and to a lesser extent with protein disulfide isomerase, two endoplasmic reticulum (ER) markers (Fig. 4B). Login to comment
140 In nontransfected cells, the distribution of calnexin was finely granular, whereas in ABCB1-I541F cells, bundles strongly labeled for calnexin colocalized with the fluorescent mutant (Fig. 4B). Login to comment
197 To check whether the temperature rescue of ABCB1-I541F in MDCK cells might be relevant for PFIC3 therapy, we studied the effect of low temperature on ABCB4 expression in HepG2 cells. Login to comment
198 Cells were transiently transfected with either ABCB4 or ABCB4-I541F cDNA. Login to comment
200 Figure 7A shows that ABCB4-I541F was detected in significant amount at the membrane of bile canaliculi in cells grown at 27°C. Login to comment
204 Low temperature rescues functional ABCB1-I541F at the apical membrane. Login to comment
205 (A) MDCK/ABCB1-I541F-GFP cells were grown on filters until confluence at 37°C, then shifted or not to 27°C for 24 hours. Login to comment
211 MDCK/ABCB1-I541F-GFP cells were grown at 37°C then shifted or not to 27°C 24 hours before the experiment. Login to comment
216 ABCB4-I541F migrated as a single 140-kDa polypeptide in cells grown at 37°C. Login to comment
218 These results show that, for ABCB1-I541F in MDCK cells, low temperature is effective on processing and bile canalicular delivery of the ABCB4 mutant in hepatic cells. Login to comment
221 Homozygous I541F ABCB4 mutation was described in a patient with PFIC3 who presented with persistent cholestasis from the age of 1 year and developed severe portal hypertension.4 In the patient`s liver, immunohistochemistry did not show any canalicular staining for ABCB4.4 This suggested that in the liver in vivo, the mutant protein could not reach the canalicular membrane because of a targeting defect or protein degradation. Login to comment
222 In both HepG2 and MDCK cells, we found that ABCB4-I541F accumulated intracellularly and was not detected at the canalicular membrane, as in the previously reported mutated patient. Login to comment
227 The location of the I541F mutation, in the first NBD domain, next to the LSSGQ signature, predicted that ATP-binding or hydrolysis could be affected. Login to comment
235 Many inherited diseases are known to arise because of point mutations within a gene that result in the production of proteins unable to assume a stable conformation within the cell.28,29 Mutations lead to non-native protein folding intermediates that are recognized by specialized chaperones and eventually targeted for destruction by the quality control machinery of the cell.30 The fact that trafficking of I541F-mutant can be rescued by lowering the temperature suggests that this mutation causes a folding defect. Login to comment
237 In the case of I541F, both isoleucine and phenylalanine are hydrophobic amino acids, but isoleucine has a branched chain, whereas phenylalanine has a Fig. 7. Login to comment
238 ABCB4-I541F reaches the bile canalicular membrane at low temperature. Login to comment
239 HepG2 cells were transfected with either ABCB4 or ABCB4-I541F cDNA. Login to comment
247 Our observation that folding of ABCB1-I541F at 27°C allows translocation of calcein indicates that, despite the mutation, the NBD domain is able to adopt a native transport-competent conformation. Login to comment
249 Response to UDCA therapy is variable among PFIC3 patients.4 The patient harboring the I541F mutation was not improved by UDCA therapy and required liver transplantation. Login to comment
252 The observation that the I541F-mutant is transport-competent if it successfully transits to the plasma membrane raises the possibility that strategies to influence protein folding inside cells might prove to have therapeutic value. Login to comment

PMID: 20425482 [PubMed] Santos JL et al: "Cholestatic liver disease in children."

No. Sentence Comment

82 This mechanism was illustrated in one study by tracking the signal produced by the I541F MDR3 mutant in liver and kidney cell lines, which was detected in the cytoplasm because of trapping within the ER and cis-Golgi [21]. Login to comment

PMID: 22184139 [PubMed] Gautherot J et al: "Effects of cellular, chemical, and pharmacological chaperones on the rescue of a trafficking-defective mutant of the ATP-binding cassette transporter proteins ABCB1/ABCB4."

No. Sentence Comment

5 Variations in the ABCB4 gene sequence cause progressivefamilialintrahepaticcholestasistype3.Wehaveshown previously that the I541F mutation, when reproduced either in ABCB1orinABCB4,ledtoretentionintheendoplasmicreticulum (ER)/Golgi. Login to comment
12 Glycerol improved maturation and exit of the mutant from theER.CyclosporinA,acompetitivesubstrateforABCB1,restored maturation, plasma membrane expression, and activity of ABCB1-I541F. Login to comment
13 Cyclosporin A also improved maturation of ABCB4-I541F in Madin-Darby canine kidney cells. Login to comment
14 In HepG2 cells transfected with ABCB4-I541F cDNA, cyclosporin A allowed a significant amount of the mutant protein to reach the membrane of bile canaliculi. Login to comment
44 We have previously investigated effects of the I541F ABCB4 mutation that has been identified in a progressive familial intrahepatic cholestasis type 3 patient. Login to comment
47 Nevertheless, intracellular traffic was restored when cells were grown at 27 °C, and the rescued ABCB1-I541F mutant was functional. Login to comment
48 The aim of this work was to test other means to provide functional rescue for the I541F mutation. Login to comment
49 We have taken the ABCB1-I541F mutant as a model to explore different strategies. Login to comment
65 The generation of MDCK cells stably expressing GFP-ABCB1, GFP-ABCB1-I541F, ABCB4 and ABCBA-I541F has been previously described (10). Login to comment
102 RESULTS The I541F Mutant Is More Susceptible to Protease Degradation-We previously showed that the I541F mutant was retained in the ER and that retention could be rescued by low temperature, suggesting that the mutant had a folding defect (10). Login to comment
104 We therefore studied whether the ABCB1-I541F was more susceptible to in situ proteolysis, using isolated microsomes from MDCK cells stably transfected with either GFP-ABCB1-WT or GFP-ABCB1-I541F. Login to comment
122 Immunoprecipitation experiments showed that calnexin coimmunoprecipitated with both ABCB1-WT and ABCB1-I541F (Fig. 2B). Login to comment
156 In these conditions, the expression of ABCB1-I541F was notably increased but remained immature, indicating that it did not exit the ER (Fig. 5B). Login to comment
157 Effect of Chemical Chaperones on ABCB1-I541F Expression-Different compounds referred to as chemical chaperones have been shown to rescue folding of mutant proteins. Login to comment
159 ABCB1-I541F expressing cells were treated with thapsigargin or 4-PB at concentrations up to 100 ␮M for 24 h. Login to comment
160 The Western blot analysis pattern of ABCB1-I541F did not change in treated cells (Fig. 6A). Login to comment
163 On the other hand, treatment with 7.5% glycerol increased the mature form of ABCB1-I541F on Western blot analyses (Fig. 6C). Login to comment
197 A substantial amount of matured ABCB4-I541F was detected in cells treated with cyclosporin A at a concentration of 2 ␮M (Fig. 8, A and B). Login to comment
199 To confirm that a similar rescue would also occur in hepatic cells, HepG2 cells were transfected with the ABCB4-I541F plasmid. Login to comment
203 In control cells, ABCB4-I541F was detected exclusively in the cytoplasm and was not detected at the membrane of bile canaliculi. Login to comment
207 We found that the ABCB1-I541F mutant was more susceptible to the in situ action of proteolytic enzymes, especially in the first moiety of the molecule, where the mutation is located. Login to comment
209 In keeping with this conclusion, it was shown previously that ABCB4 was undetectable by immunohistochemistry in the liver tissue from a patient bearing the ABCB4-I541F mutation (16). Login to comment
213 Effect of chemical chaperones on the expression pattern of ABCB1-I541F. Login to comment
214 A, MDCK cells stably transfected with GFP-tagged ABCB1-WT or ABCB1-I541F were treated with vehicle (-), 4-PB, or thapsigargin (Thap) at the indicated concentrations for 24 h. Login to comment
216 B, filter-grown MDCK cells stably transfected with GFP-tagged ABCB1-I541F were treated with vehicle (control), 100 ␮M thapsigargin, or 1 mM 4-phenylbutyrate for 24 h. Login to comment
219 D, MDCK cells stably transfected with GFP-tagged ABCB1-I541F were treated with 7.5% glycerol for 24 h. Login to comment
223 Scale bar ϭ 10 ␮m. E, the colocalization of GFP-tagged ABCB1-I541F with GM130 was quantified using the ImageJ 1.41 measure colocalization function on multiple confocal sections of at least 30 cells in three independent experiments. Login to comment
256 In our system, treatment with thapsigargin and 4-PB had no effect on the I541F mutant. Login to comment
257 Only glycerol did improve the maturation of the ABCB1-I541F mutant, which was able to exit the ER. Login to comment
262 Finally, rescue of the I541F mutant was obtained with a specific substrate. Login to comment

PMID: 16622704 [PubMed] Oude Elferink RP et al: "Function and pathophysiological importance of ABCB4 (MDR3 P-glycoprotein)."

No. Sentence Comment

141 Canalicular lipid transport defects can cause gallstone formation Cholesterol supersaturation of bile, which occurs in a large proportion of humans, leads to the formation of cholesterol Walker B; L556R 571del Truncation PFIC3 LPAC ICP 27 splice Truncation 132 del Truncation TM 2; W138R TM 12; 981 del Truncation Linker; Q636X Truncation TM 11; R957X Truncation TM 6; S346I E395G Walker B; I541F TM 12; G983S Walker A; V425M Walker A; T424A Walker B; D564G TM 7; F711S 180 del truncation 336 delT truncation Exon 22-23 del truncation F165I T175A TM 5; M301T TM 5; S320F 336 insT truncation Walker A; 432 insA truncation E528D L591Q W658stop 757 insT R788E A934T P1161S TM 5; S320F TM 8; G762ER144X Walker B; A546D Walker B; G535AALL 96 del Truncation Walker B; L556R 571del Truncation PFIC3 LPAC ICP 27 splice Truncation 132 del Truncation TM 2; W138R TM 12; 981 del Truncation Linker; Q636X Truncation TM 11; R957X Truncation TM 6; S346I E395G Walker B; I541F TM 12; G983S Walker A; V425M Walker A; T424A Walker B; D564G TM 7; F711S 180 del truncation 336 delT truncation Exon 22-23 del truncation F165I T175A TM 5; M301T TM 5; S320F 336 insT truncation Walker A; 432 insA truncation E528D L591Q W658stop 757 insT R788E A934T P1161S TM 5; S320F TM 8; G762ER144X Walker B; A546D Walker B; G535AALL 96 del Truncation Fig. 3 Summary of the known mutations and their localization in the protein, as identified in patients with PFIC type 3, LPAC syndrome (intrahepatic gallstone formation), and intrahepatic cholestasis of pregnancy (ICP). Login to comment

PMID: 11313315 [PubMed] Jacquemin E et al: "The wide spectrum of multidrug resistance 3 deficiency: from neonatal cholestasis to cirrhosis of adulthood."

No. Sentence Comment

126 MDR3 Mutations, Immunohistochemistry, and Biliary Phospholipids in Patients With High GGT PFIC (PFIC3) Patients Exon Nucleotide mutation Amino acid change Protein consequence Family MDR3 liver Phospholipids (%) bile Homozygous Deletion/Insertion M Y M I (sister of M Y) 2 111 A Ͼ G, splice donor site, exon/intron boundary 2/3 27 Frame shift in NH2 terminus, then truncation ND Absent ND 1.6 B Ka 6 426-432 del 132 Frame shift in TMD 2, 29 novel amino acids then truncation Mother ϩ/- Father ϩ/- 1 sibling ϩ/ϩ Absent ND B Sa 14 1744 del T 571 Frame shift, 15 novel amino acids then truncation Mother ϩ/- (ICP) Father ϩ/- 3 siblings ϩ/ϩ Absent ND P G 24 2975-2984 del Compound heterozygous 981 Frame shift in TMD 12, 3 novel amino acids then truncation Mother ϩ/ϩ Father ϩ/- ND 14.9 Bo S Bo N (sister of Bo S) 16 Nonsense 1938 C Ͼ T Q636X Linker region Truncation Mother ϩ/- Father ϩ/- 2 siblings ϩ/- Absent 2 ND B Aa 23 2901 C Ͼ T R957X TMD 11 Truncation Mother ϩ/- (ICP) Father ϩ/- 4 siblings ϩ/- or ϩ/ϩ Absent ND S F 10 Missense 1069 G Ͼ T S346I Serine to isoleucine in TMD6 Mother ϩ/- Father ϩ/- Faint 1 B H 11 1216 A Ͼ G E395G Glutamate to glycine between TMD 6 and first Walker A motif ND ND ND N I 14 1653 A Ͼ T I541F Isoleucine to phenylalanine in first Walker B motif Mother ϩ/- Father ϩ/- Absent ND M M 14 1699 T Ͼ G L556R Leucine to arginine in first Walker B motif ND ND ND P G 24 2979 G Ͼ A Compound heterozygous G983S Glycine to serine in TMD 12 Mother ϩ/- Father ϩ/ϩ ND 14.9 P A 6 Heterozygous Missense 444 T Ͼ C W138R Trytophan to arginine in TMD 2 Mother ϩ/- Father ϩ/ϩ ND 6.7 M Aa 12 1302 A Ͼ G T424A Threonine to alanine in first Walker A motif ND gallbladder lithiasis in father Faint 6.3 P K 12 1307 G Ͼ A V425M Valine to methionine in first Walker A motif ND Normal ND G A 14 1723 A Ͼ G D564G Aspartate to glycine in first Walker B motif ND ND ND G M 17 2132 T Ͼ C F711S Phenylalanine to serine in TMD 7 ND ND ND L M 16 Polymorphism 1986 A Ͼ G Homozygous R652G Arginine to glycine in linker region Mother ϩ/- (suspicion of ICP) Father ϩ/- ND ND Ga M " Heterozygous " " Mother ϩ/- Father ϩ/ϩ 1 sibling ϩ/- ND 9.7 L H L F (sister of L H) " Heterozygous " " ND ND ND VH C " Heterozygous " " Mother ϩ/ϩ Father ϩ/- ND ND NOTE. Login to comment

PMID: 23261441 [PubMed] Jang GH et al: "Functional characterization of genetic variations in the MDR3 promoter."

No. Sentence Comment

19 For example, a nonsynonymous mutation, p.I541F, found in a PFIC3 patient showed decreased transport activity owing to reduced membrane expression [8]. Login to comment

PMID: 24953525 [PubMed] Falguieres T et al: "ABCB4: Insights from pathobiology into therapy."

No. Sentence Comment

83 Wild-type ABCB4 (A) or I541F missense mutant (B, C) were expressed in HepG2 cells. Login to comment
105 The I541F mutation led to improper folding, retention in the endoplasmic reticulum and premature degradation [59,60] (Fig. 2). Login to comment
127 We recently showed that the ER-retained I541F missense mutant of ABCB4, identified in patients with PFIC3, was rescued by cyclosporin A treatment, while 4-phenylbutyrate had no effect [60] (Fig. 2). Login to comment

PMID: 24381502 [PubMed] Kim TH et al: "Functional Characterization of ABCB4 Mutations Found in Low Phospholipid-Associated Cholelithiasis (LPAC)."

No. Sentence Comment

20 For example, a ABCB4 mutation found in PFIC3 patients, I541F, was shown to decrease transport activity through reduction of membrane MDR3 expression [10,11]. Login to comment
123 One of the ABCB4 mutations, I541F, was found in PFIC3 patients and was shown to be a trafficking-defective mutation [10,11]. Login to comment
125 2 mutants, F165I and S320F, examined in this study also showed decreased transport activity and protein expression, although the extent of reduction was less than that of I541F. Login to comment