Jian Ding, Hui-Yu She, Ye Cheng, Hong-Yuan Sun, Jia-Yan Feng, Teng Liu, Yi-Ling Qiu, Bing-Xuan Wei, Jin Zhang, Yu Su, Yun-Qian Li, Jun-Jie Zhang, Si-Yuan Chen, Ting Wang, Yue Yu, Sven C D van IJzendoorn, Jian-She Wang, Qing-He Xing
{"title":"一种涉及usp53调控的BSEP贩运的新机制是低ggt肝内胆汁淤积。","authors":"Jian Ding, Hui-Yu She, Ye Cheng, Hong-Yuan Sun, Jia-Yan Feng, Teng Liu, Yi-Ling Qiu, Bing-Xuan Wei, Jin Zhang, Yu Su, Yun-Qian Li, Jun-Jie Zhang, Si-Yuan Chen, Ting Wang, Yue Yu, Sven C D van IJzendoorn, Jian-She Wang, Qing-He Xing","doi":"10.1097/HEP.0000000000001501","DOIUrl":null,"url":null,"abstract":"<p><strong>Background and aims: </strong>USP53 variants cause low-GGT progressive familial intrahepatic cholestasis (PFIC). The mechanism is not well understood. USP53, which encodes a ubiquitin-specific protease, has been proposed to be involved in blood-bile barrier impairment. However, Usp53 knockout mice did not show blood-bile barrier impairment. The aim of this study was to investigate the molecular mechanism underlying USP53 -PFIC.</p><p><strong>Approach and results: </strong>Immunohistochemistry of patient tissue, confocal immunofluorescence microscopy and surface protein biotinylation were performed to investigate the localization of proteins of interest. Small-interference RNA and CRISPR-Cas9 technology were used to downregulate or knock out genes, respectively. Site-directed mutagenesis was performed to generate gene variants for expression in cells. Co-immunoprecipitation experiments were performed to investigate (variant) protein-protein interactions. Live cell imaging and fluorescence recovery after photobleaching were performed to investigate protein dynamics. The mislocalization of the bile salt export pump (BSEP) was demonstrated in hepatocytes of a USP53-associated PFIC patient and USP53 -KO cells. Loss of USP53 caused BSEP accumulation in MYO5B-positive and RAB11A-positive recycling endosomes and impaired BSEP trafficking to the plasma membrane. USP53 colocalized with MYO5B and interacted with its IQ domain. The recurrent MYO5B-PFIC-associated p.(Arg824Cys) variant, located in the IQ domain, failed to interact with USP53. Loss of USP53 expression resulted in increased ubiquitination of MYO5B and interfered with the endosomal recruitment of MYO5B.</p><p><strong>Conclusions: </strong>Loss of USP53 interaction with MYO5B and its p.(Arg824Cys) variant impaired BSEP trafficking in USP53 -associated and MYO5B -associated low-GGT intrahepatic cholestasis. These results provide a novel mechanism that underlies USP53 -PFIC and implicates USP53 in the pathogenesis of MYO5B -PFIC.</p>","PeriodicalId":177,"journal":{"name":"Hepatology","volume":" ","pages":""},"PeriodicalIF":15.8000,"publicationDate":"2025-08-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":"{\"title\":\"A novel mechanism involving USP53-regulated BSEP trafficking underlies low-GGT intrahepatic cholestasis.\",\"authors\":\"Jian Ding, Hui-Yu She, Ye Cheng, Hong-Yuan Sun, Jia-Yan Feng, Teng Liu, Yi-Ling Qiu, Bing-Xuan Wei, Jin Zhang, Yu Su, Yun-Qian Li, Jun-Jie Zhang, Si-Yuan Chen, Ting Wang, Yue Yu, Sven C D van IJzendoorn, Jian-She Wang, Qing-He Xing\",\"doi\":\"10.1097/HEP.0000000000001501\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><strong>Background and aims: </strong>USP53 variants cause low-GGT progressive familial intrahepatic cholestasis (PFIC). The mechanism is not well understood. USP53, which encodes a ubiquitin-specific protease, has been proposed to be involved in blood-bile barrier impairment. However, Usp53 knockout mice did not show blood-bile barrier impairment. The aim of this study was to investigate the molecular mechanism underlying USP53 -PFIC.</p><p><strong>Approach and results: </strong>Immunohistochemistry of patient tissue, confocal immunofluorescence microscopy and surface protein biotinylation were performed to investigate the localization of proteins of interest. Small-interference RNA and CRISPR-Cas9 technology were used to downregulate or knock out genes, respectively. Site-directed mutagenesis was performed to generate gene variants for expression in cells. Co-immunoprecipitation experiments were performed to investigate (variant) protein-protein interactions. Live cell imaging and fluorescence recovery after photobleaching were performed to investigate protein dynamics. The mislocalization of the bile salt export pump (BSEP) was demonstrated in hepatocytes of a USP53-associated PFIC patient and USP53 -KO cells. Loss of USP53 caused BSEP accumulation in MYO5B-positive and RAB11A-positive recycling endosomes and impaired BSEP trafficking to the plasma membrane. USP53 colocalized with MYO5B and interacted with its IQ domain. The recurrent MYO5B-PFIC-associated p.(Arg824Cys) variant, located in the IQ domain, failed to interact with USP53. Loss of USP53 expression resulted in increased ubiquitination of MYO5B and interfered with the endosomal recruitment of MYO5B.</p><p><strong>Conclusions: </strong>Loss of USP53 interaction with MYO5B and its p.(Arg824Cys) variant impaired BSEP trafficking in USP53 -associated and MYO5B -associated low-GGT intrahepatic cholestasis. These results provide a novel mechanism that underlies USP53 -PFIC and implicates USP53 in the pathogenesis of MYO5B -PFIC.</p>\",\"PeriodicalId\":177,\"journal\":{\"name\":\"Hepatology\",\"volume\":\" \",\"pages\":\"\"},\"PeriodicalIF\":15.8000,\"publicationDate\":\"2025-08-19\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Hepatology\",\"FirstCategoryId\":\"3\",\"ListUrlMain\":\"https://doi.org/10.1097/HEP.0000000000001501\",\"RegionNum\":1,\"RegionCategory\":\"医学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"\",\"PubModel\":\"\",\"JCR\":\"Q1\",\"JCRName\":\"GASTROENTEROLOGY & HEPATOLOGY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Hepatology","FirstCategoryId":"3","ListUrlMain":"https://doi.org/10.1097/HEP.0000000000001501","RegionNum":1,"RegionCategory":"医学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q1","JCRName":"GASTROENTEROLOGY & HEPATOLOGY","Score":null,"Total":0}
Background and aims: USP53 variants cause low-GGT progressive familial intrahepatic cholestasis (PFIC). The mechanism is not well understood. USP53, which encodes a ubiquitin-specific protease, has been proposed to be involved in blood-bile barrier impairment. However, Usp53 knockout mice did not show blood-bile barrier impairment. The aim of this study was to investigate the molecular mechanism underlying USP53 -PFIC.
Approach and results: Immunohistochemistry of patient tissue, confocal immunofluorescence microscopy and surface protein biotinylation were performed to investigate the localization of proteins of interest. Small-interference RNA and CRISPR-Cas9 technology were used to downregulate or knock out genes, respectively. Site-directed mutagenesis was performed to generate gene variants for expression in cells. Co-immunoprecipitation experiments were performed to investigate (variant) protein-protein interactions. Live cell imaging and fluorescence recovery after photobleaching were performed to investigate protein dynamics. The mislocalization of the bile salt export pump (BSEP) was demonstrated in hepatocytes of a USP53-associated PFIC patient and USP53 -KO cells. Loss of USP53 caused BSEP accumulation in MYO5B-positive and RAB11A-positive recycling endosomes and impaired BSEP trafficking to the plasma membrane. USP53 colocalized with MYO5B and interacted with its IQ domain. The recurrent MYO5B-PFIC-associated p.(Arg824Cys) variant, located in the IQ domain, failed to interact with USP53. Loss of USP53 expression resulted in increased ubiquitination of MYO5B and interfered with the endosomal recruitment of MYO5B.
Conclusions: Loss of USP53 interaction with MYO5B and its p.(Arg824Cys) variant impaired BSEP trafficking in USP53 -associated and MYO5B -associated low-GGT intrahepatic cholestasis. These results provide a novel mechanism that underlies USP53 -PFIC and implicates USP53 in the pathogenesis of MYO5B -PFIC.
期刊介绍：
HEPATOLOGY is recognized as the leading publication in the field of liver disease. It features original, peer-reviewed articles covering various aspects of liver structure, function, and disease. The journal's distinguished Editorial Board carefully selects the best articles each month, focusing on topics including immunology, chronic hepatitis, viral hepatitis, cirrhosis, genetic and metabolic liver diseases, liver cancer, and drug metabolism.
求助内容：
应助结果提醒方式：
京公网安备 11010802042870号
