ABCC2 p.R393W variant contributes to Dubin-Johnson syndrome by targeting MRP2 to proteasome degradation
Background Dubin-Johnson syndrome (DJS), a rare autosomal recessive liver condition, is caused by biallelic loss-of-function mutations of the ABCC2 gene. This study aimed to investigate genetic variations in the drug efflux transporter ABCC2 (MRP2) gene in patients with DJS and to characterise the e...
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BMJ Publishing Group
2024-02-01
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| Series: | eGastroenterology |
| Online Access: | https://egastroenterology.bmj.com/content/2/1/e100039.full |
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| author | Dan Wang Yi-Ming Chen Luca Valenti Ming-Hua Zheng Ting Zhu Rong-Yue Sun Mian-Mian Zhu Ji-an Sun Hong-Wei Wang Chen-Yu Wu Yu-Jing Gong Chao-Sheng Lu Luisa Ronzoni |
| author_facet | Dan Wang Yi-Ming Chen Luca Valenti Ming-Hua Zheng Ting Zhu Rong-Yue Sun Mian-Mian Zhu Ji-an Sun Hong-Wei Wang Chen-Yu Wu Yu-Jing Gong Chao-Sheng Lu Luisa Ronzoni |
| author_sort | Dan Wang |
| collection | DOAJ |
| description | Background Dubin-Johnson syndrome (DJS), a rare autosomal recessive liver condition, is caused by biallelic loss-of-function mutations of the ABCC2 gene. This study aimed to investigate genetic variations in the drug efflux transporter ABCC2 (MRP2) gene in patients with DJS and to characterise the expression and mechanism of the ABCC2 gene variant.Methods Trio whole exome sequencing was performed in the family to identify the genetic causes. Bioinformatics analysis was performed to assess pathogenicity. In in vitro experiments, site-directed mutagenesis was used to introduce ABCC2 variants in constructs then expressed in HEK293T, HuH-7 and HepG2 cell lines. The expression of total and cell membrane MRP2 was quantified in cells expressing the wild-type or variant forms. Chloroquine and MG132 were used to evaluate the effects of p.R393W on lysosomal and/or proteasomal degradation.Results The twin probands carry DJS-associated variants c.1177C>T (rs777902199) in the ABCC2 gene inherited from the father and the c.3632T>C mutation in the other allele inherited from the mother. The ABCC2 variant, c.1177C>T, results in a p.R393W substitution in MRP2 that is highly conserved among vertebrates, drastically decreasing the expression of mutant protein by promoting proteasomal degradation. Another variant c.3632T>C results in a p.L1211P substitution in MRP2, decreasing the expression of membrane MRP2 but not changing the expression of total protein.Conclusion These results strongly suggest that the p.R393W variant affects the stability of the MRP2 protein and decreases its expression by ubiquitin-mediated proteasomal degradation, and the p.L1211P decreases the expression of membrane MRP2, indicating that these two variants, respectively, cause a loss-of-function of the MRP2 protein and membrane MRP2 ultimately leading to DJS development. |
| format | Article |
| id | doaj-art-ea7c654919aa4da397c41a87b410cf0b |
| institution | Kabale University |
| issn | 2766-0125 2976-7296 |
| language | English |
| publishDate | 2024-02-01 |
| publisher | BMJ Publishing Group |
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| series | eGastroenterology |
| spelling | doaj-art-ea7c654919aa4da397c41a87b410cf0b2024-12-25T15:00:10ZengBMJ Publishing GroupeGastroenterology2766-01252976-72962024-02-012110.1136/egastro-2023-100039ABCC2 p.R393W variant contributes to Dubin-Johnson syndrome by targeting MRP2 to proteasome degradationDan Wang0Yi-Ming Chen1Luca Valenti2Ming-Hua Zheng3Ting Zhu4Rong-Yue Sun5Mian-Mian Zhu6Ji-an Sun7Hong-Wei Wang8Chen-Yu Wu9Yu-Jing Gong10Chao-Sheng Lu11Luisa Ronzoni12The Affiliated Xuzhou Rehabilitation Hospital of Xuzhou Medical University, Xuzhou Medical University, Xuzhou, Jiangsu, China3 Faculty of Medicine, National Yang Ming University, Taipei, Taiwan18 Translational Medicine, Department of Transfusion Medicine and Hematology, Fondazione IRCCS C`a Granda Ospedale Maggiore Policlinico, Department of Pathophysiology and Transplantation, University of Milan, Milan, Italy31 Institute of Hepatology, Wenzhou Medical University, Wenzhou, Zhejiang, China1 Department of Pediatrics, The First Affiliated Hospital Medical University, Wenzhou, Zhejiang, China1 Department of Pediatrics, The First Affiliated Hospital Medical University, Wenzhou, Zhejiang, China1 Department of Pediatrics, The First Affiliated Hospital Medical University, Wenzhou, Zhejiang, China1 Department of Pediatrics, The First Affiliated Hospital Medical University, Wenzhou, Zhejiang, China3 Key Laboratory of Diagnosis and Treatment of Severe Hepato‐Pancreatic Diseases, The First Affiliated Hospital Medical University, Wenzhou, Zhejiang, China1 Department of Pediatrics, The First Affiliated Hospital Medical University, Wenzhou, Zhejiang, China1 Department of Pediatrics, The First Affiliated Hospital Medical University, Wenzhou, Zhejiang, China1 Department of Pediatrics, The First Affiliated Hospital Medical University, Wenzhou, Zhejiang, China4 Fondazione IRCCS Ca’ Granda Ospedale Maggiore Policlinico, Milan, ItalyBackground Dubin-Johnson syndrome (DJS), a rare autosomal recessive liver condition, is caused by biallelic loss-of-function mutations of the ABCC2 gene. This study aimed to investigate genetic variations in the drug efflux transporter ABCC2 (MRP2) gene in patients with DJS and to characterise the expression and mechanism of the ABCC2 gene variant.Methods Trio whole exome sequencing was performed in the family to identify the genetic causes. Bioinformatics analysis was performed to assess pathogenicity. In in vitro experiments, site-directed mutagenesis was used to introduce ABCC2 variants in constructs then expressed in HEK293T, HuH-7 and HepG2 cell lines. The expression of total and cell membrane MRP2 was quantified in cells expressing the wild-type or variant forms. Chloroquine and MG132 were used to evaluate the effects of p.R393W on lysosomal and/or proteasomal degradation.Results The twin probands carry DJS-associated variants c.1177C>T (rs777902199) in the ABCC2 gene inherited from the father and the c.3632T>C mutation in the other allele inherited from the mother. The ABCC2 variant, c.1177C>T, results in a p.R393W substitution in MRP2 that is highly conserved among vertebrates, drastically decreasing the expression of mutant protein by promoting proteasomal degradation. Another variant c.3632T>C results in a p.L1211P substitution in MRP2, decreasing the expression of membrane MRP2 but not changing the expression of total protein.Conclusion These results strongly suggest that the p.R393W variant affects the stability of the MRP2 protein and decreases its expression by ubiquitin-mediated proteasomal degradation, and the p.L1211P decreases the expression of membrane MRP2, indicating that these two variants, respectively, cause a loss-of-function of the MRP2 protein and membrane MRP2 ultimately leading to DJS development.https://egastroenterology.bmj.com/content/2/1/e100039.full |
| spellingShingle | Dan Wang Yi-Ming Chen Luca Valenti Ming-Hua Zheng Ting Zhu Rong-Yue Sun Mian-Mian Zhu Ji-an Sun Hong-Wei Wang Chen-Yu Wu Yu-Jing Gong Chao-Sheng Lu Luisa Ronzoni ABCC2 p.R393W variant contributes to Dubin-Johnson syndrome by targeting MRP2 to proteasome degradation eGastroenterology |
| title | ABCC2 p.R393W variant contributes to Dubin-Johnson syndrome by targeting MRP2 to proteasome degradation |
| title_full | ABCC2 p.R393W variant contributes to Dubin-Johnson syndrome by targeting MRP2 to proteasome degradation |
| title_fullStr | ABCC2 p.R393W variant contributes to Dubin-Johnson syndrome by targeting MRP2 to proteasome degradation |
| title_full_unstemmed | ABCC2 p.R393W variant contributes to Dubin-Johnson syndrome by targeting MRP2 to proteasome degradation |
| title_short | ABCC2 p.R393W variant contributes to Dubin-Johnson syndrome by targeting MRP2 to proteasome degradation |
| title_sort | abcc2 p r393w variant contributes to dubin johnson syndrome by targeting mrp2 to proteasome degradation |
| url | https://egastroenterology.bmj.com/content/2/1/e100039.full |
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