Hepatitis B virus enhancer 1 activates preS1 and preS2 promoters of integrated HBV DNA impairing HBsAg secretion
Zhiqiang Gu,
Qianqian Jiang,
Abudurexiti Abulaiti,
Xiaojie Chen,
Mingwei Li,
Na Gao,
Guiwen Guan,
Ting Zhang,
Danli Yang,
Jingyuan Xi,
Guangxin Yu,
Shuhong Liu,
Zhijun Zhu,
Zhiliang Gao,
Jingmin Zhao,
Hongxin Huang,
Xiangmei Chen,
Fengmin Lu
Affiliations
Zhiqiang Gu
Department of Microbiology & Infectious Disease Center, School of Basic Medical Sciences, Peking University, Beijing, China
Qianqian Jiang
Peking University People’s Hospital, Peking University Hepatology Institute, Beijing Key Laboratory of Hepatitis C and Immunotherapy for Liver Diseases, Beijing International Cooperation Base for Science and Technology on NAFLD Diagnosis, Beijing, China
Abudurexiti Abulaiti
Department of Microbiology & Infectious Disease Center, School of Basic Medical Sciences, Peking University, Beijing, China
Xiaojie Chen
Liver Transplantation Center, National Clinical Research Center for Digestive Diseases, Beijing Friendship Hospital, Capital Medical University, Beijing, China; Clinical Research Center for Pediatric Liver Transplantation of Capital Medical University, Beijing, China
Mingwei Li
Research Center for Clinical Medical Sciences, The Fourth Hospital of Shijiazhuang, Shijiazhuang, Hebei, China
Na Gao
Department of Infectious Diseases, Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, Guangdong, China
Guiwen Guan
Department of Microbiology & Infectious Disease Center, School of Basic Medical Sciences, Peking University, Beijing, China
Ting Zhang
Department of Microbiology & Infectious Disease Center, School of Basic Medical Sciences, Peking University, Beijing, China
Danli Yang
Department of Microbiology & Infectious Disease Center, School of Basic Medical Sciences, Peking University, Beijing, China
Jingyuan Xi
Department of Clinical Laboratory Center, Beijing Children′s Hospital, Capital Medical University, National Center for Children′s Health, Beijing, China
Guangxin Yu
Department of Microbiology & Infectious Disease Center, School of Basic Medical Sciences, Peking University, Beijing, China
Shuhong Liu
Department of Pathology and Hepatology, The Fifth Medical Center of PLA General Hospital, Beijing, China
Zhijun Zhu
Liver Transplantation Center, National Clinical Research Center for Digestive Diseases, Beijing Friendship Hospital, Capital Medical University, Beijing, China; Clinical Research Center for Pediatric Liver Transplantation of Capital Medical University, Beijing, China
Zhiliang Gao
Department of Infectious Diseases, Third Affiliated Hospital of Sun Yat-Sen University, Guangzhou, Guangdong, China; Guangdong Key Laboratory of Liver Disease Research, The Third Affiliated Hospital of Sun Yat-sen University, Guangzhou, Guangdong, China; Key Laboratory of Tropical Disease Control (Sun Yat-Sen University), Ministry of Education, Guangzhou, Guangdong, China
Jingmin Zhao
Department of Pathology and Hepatology, The Fifth Medical Center of PLA General Hospital, Beijing, China
Hongxin Huang
Department of Pathogen Biology and Biosecurity, Zhongshan School of Medicine, Sun Yat-sen University, Guangzhou, Guangdong, China; Corresponding authors. Addresses: Department of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University, Beijing 100191, China; Beijing Key Laboratory of Hepatitis C and Immunotherapy for Liver Diseases, Beijing International Cooperation Base for Science and Technology on NAFLD Diagnosis, Peking University Hepatology Institute, Peking University People's Hospital, Beijing 100044, China (F. Lu); Department of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University, Beijing 100191, China (X. Chen); Department of Pathogen Biology and Biosecurity, Zhongshan School of Medicine, Sun Yat-sen University, Guangdong 510080, China (H. Huang).
Xiangmei Chen
Department of Microbiology & Infectious Disease Center, School of Basic Medical Sciences, Peking University, Beijing, China; Corresponding authors. Addresses: Department of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University, Beijing 100191, China; Beijing Key Laboratory of Hepatitis C and Immunotherapy for Liver Diseases, Beijing International Cooperation Base for Science and Technology on NAFLD Diagnosis, Peking University Hepatology Institute, Peking University People's Hospital, Beijing 100044, China (F. Lu); Department of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University, Beijing 100191, China (X. Chen); Department of Pathogen Biology and Biosecurity, Zhongshan School of Medicine, Sun Yat-sen University, Guangdong 510080, China (H. Huang).
Fengmin Lu
Department of Microbiology & Infectious Disease Center, School of Basic Medical Sciences, Peking University, Beijing, China; Peking University People’s Hospital, Peking University Hepatology Institute, Beijing Key Laboratory of Hepatitis C and Immunotherapy for Liver Diseases, Beijing International Cooperation Base for Science and Technology on NAFLD Diagnosis, Beijing, China; Corresponding authors. Addresses: Department of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University, Beijing 100191, China; Beijing Key Laboratory of Hepatitis C and Immunotherapy for Liver Diseases, Beijing International Cooperation Base for Science and Technology on NAFLD Diagnosis, Peking University Hepatology Institute, Peking University People's Hospital, Beijing 100044, China (F. Lu); Department of Microbiology and Infectious Disease Center, School of Basic Medical Sciences, Peking University, Beijing 100191, China (X. Chen); Department of Pathogen Biology and Biosecurity, Zhongshan School of Medicine, Sun Yat-sen University, Guangdong 510080, China (H. Huang).
Background & Aims: The expression of HBsAg from integrated HBV DNA limits the achievement of functional cure for chronic hepatitis B. Thus, characterising the unique expression and secretion of HBsAg derived from integrated HBV DNA is of clinical significance. Methods: A total of 563 treatment-naive patients and 62 functionally cured patients were enrolled, and HBsAg and HBcAg immunohistochemistry of their liver biopsy tissues was conducted followed by semi-quantitative analysis. Then, based on stratified analysis of HBeAg-positive and -negative patients, long-read RNA sequencing analysis, as well as an in vitro HBV integration model, we explored the HBsAg secretion characteristics of integrated HBV DNA and underlying mechanisms. Results: In contrast to the significantly lower serum HBsAg levels, no significant decrease of intrahepatic HBsAg protein was observed in HBeAg-negative patients, as compared with HBeAg-positive patients. The results of long-read RNA sequencing of liver tissues from patients with chronic HBV infection and in vitro studies using integrated HBV DNA mimicking dslDNA plasmid revealed that, the lower HBsAg secretion efficiency seen in HBeAg-negative patients might be attributed to an increased proportion of preS1 mRNA derived from integrated HBV DNA instead of covalently closed circular DNA. The latter resulted in an increased L-HBsAg proportion and impaired HBsAg secretion. Enhancer 1 (EnhI) in integrated HBV DNA could retarget preS1 (SP1) and preS2 (SP2) promoters to disrupt their transcriptional activity balance. Conclusions: The secretion of HBsAg originating from integrated HBV DNA was impaired. Mechanistically, functional deficiency of core promoter leads to retargeting of EnhI and thus uneven activation of the SP1 over the SP2 promoter, resulting in an increase in the proportion of L-HBsAg. Impact and implications: Integrated hepatitis B virus (HBV) DNA can serve as an important reservoir for HBV surface antigen (HBsAg) expression, and this limits the achievement of a functional cure. This study revealed that secretion efficiency is lower for HBsAg derived from integrated HBV DNA than HBsAg derived from covalently closed circular DNA, as determined by the unique sequence features of integrated HBV DNA. This study can broaden our understanding of the role of HBV integration and shed new light on antiviral strategies to facilitate a functional cure. We believe our results are of great general interest to a broad audience, including patients and patient organisations, the medical community, academia, the life science industry and the public.
