Caizhi Liu1, Bing Zhou2, Meiyao Meng3, Wenjun Zhao3, Dongmei Wang3, Youwen Yuan4, Ying Zheng3, Jin Qiu3, Yu Li3, Guoqiang Li3, Xuelian Xiong2, Hua Bian2, Huijie Zhang4, Hua Wang5, Xinran Ma6, Cheng Hu7, Lingyan Xu8, Yan Lu9. 1. Joint Center for Translational Medicine, Fengxian District Central Hospital, Fengxian District, Shanghai, China; Shanghai Key Laboratory of Regulatory Biology, Institute of Biomedical Sciences and School of Life Sciences, East China Normal University, Shanghai, China. 2. The Key Laboratory of Metabolism and Molecular Medicine of the Ministry of Education, Department of Endocrinology and Metabolism, Zhongshan Hospital, Fudan University, Shanghai, China. 3. Shanghai Key Laboratory of Regulatory Biology, Institute of Biomedical Sciences and School of Life Sciences, East China Normal University, Shanghai, China. 4. Department of Endocrinology and Metabolism, Nanfang Hospital, Southern Medical University, Guangzhou, China. 5. Department of Oncology, The First Affiliated Hospital, Institute for Liver Diseases of Anhui Medical University, Hefei, China. 6. Joint Center for Translational Medicine, Fengxian District Central Hospital, Fengxian District, Shanghai, China; Shanghai Key Laboratory of Regulatory Biology, Institute of Biomedical Sciences and School of Life Sciences, East China Normal University, Shanghai, China. Electronic address: xrma@bio.ecnu.edu.cn. 7. Joint Center for Translational Medicine, Fengxian District Central Hospital, Fengxian District, Shanghai, China; Shanghai Diabetes Institute, Shanghai Key Laboratory of Diabetes Mellitus, Shanghai Clinical Centre for Diabetes, Shanghai Jiao Tong University Affiliated Sixth People's Hospital, Shanghai, China. Electronic address: alfredhc@sjtu.edu.cn. 8. Shanghai Key Laboratory of Regulatory Biology, Institute of Biomedical Sciences and School of Life Sciences, East China Normal University, Shanghai, China. Electronic address: lyxu@bio.ecnu.edu.cn. 9. The Key Laboratory of Metabolism and Molecular Medicine of the Ministry of Education, Department of Endocrinology and Metabolism, Zhongshan Hospital, Fudan University, Shanghai, China. Electronic address: lu.yan2@zs-hospital.sh.cn.
Abstract
BACKGROUND & AIMS: Chronic endoplasmic reticulum (ER) stress in the liver has been shown to play a causative role in non-alcoholic fatty liver disease (NAFLD) progression, yet the underlying molecular mechanisms remain to be elucidated. Forkhead box A3 (FOXA3), a member of the FOX family, plays critical roles in metabolic homeostasis, although its possible functions in ER stress and fatty liver progression are unknown. METHODS: Adenoviral delivery, siRNA delivery, and genetic knockout mice were used to crease FOXA3 gain- or loss-of-function models. Tunicamycin (TM) and a high-fat diet (HFD) were used to induce acute or chronic ER stress in mice. Chromatin immunoprecipiation (ChIP)-seq, luciferase assay, and adenoviral-mediated downstream gene manipulations were performed to reveal the transcriptional axis involved. Key axis protein levels in livers from healthy donors and patients with NAFLD were assessed via immunohistochemical staining. RESULTS: FOXA3 transcription is specifically induced by XBP1s upon ER stress. FOXA3 exacerbates the excessive lipid accumulation caused by the acute ER-inducer TM, whereas FOXA3 deficiency in hepatocytes and mice alleviates it. Importantly, FOXA3 deficiency in mice reduced diet-induced chronic ER stress, fatty liver, and insulin resistance. In addition, FOXA3 suppression via siRNA or adeno-associated virus delivery ameliorated the fatty liver phenotype in HFD-fed and db/db mice. Mechanistically, ChIP-Seq analysis revealed that FOXA3 directly regulates Period1 (Per1) transcription, which in turn promotes the expression of lipogenic genes, including Srebp1c, thus enhancing lipid synthesis. Of pathophysiological significance, FOXA3, PER1, and SREBP1c levels were increased in livers of obese mice and patients with NAFLD. CONCLUSION: The present study identified FOXA3 as the bridging molecule that links ER stress and NAFLD progression. Our results highlighted the role of the XBP1s-FOXA3-PER1/Srebp1c transcriptional axis in the development of NAFLD and identified FOXA3 as a potential therapeutic target for fatty liver disease. LAY SUMMARY: The molecular mechanisms linking endoplasmic reticulum stress to non-alcoholic fatty liver disease (NAFLD) progression remain undefined. Herein, via in vitro and in vivo analysis, we identified Forkhead box A3 (FOXA3) as a key bridging molecule. Of pathophysiological significance, FOXA3 protein levels were increased in livers of obese mice and patients with NAFLD, indicating that FOXA3 could be a potential therapeutic target in fatty liver disease.
BACKGROUND & AIMS: Chronic endoplasmic reticulum (ER) stress in the liver has been shown to play a causative role in non-alcoholic fatty liver disease (NAFLD) progression, yet the underlying molecular mechanisms remain to be elucidated. Forkhead box A3 (FOXA3), a member of the FOX family, plays critical roles in metabolic homeostasis, although its possible functions in ER stress and fatty liver progression are unknown. METHODS: Adenoviral delivery, siRNA delivery, and genetic knockout mice were used to crease FOXA3 gain- or loss-of-function models. Tunicamycin (TM) and a high-fat diet (HFD) were used to induce acute or chronic ER stress in mice. Chromatin immunoprecipiation (ChIP)-seq, luciferase assay, and adenoviral-mediated downstream gene manipulations were performed to reveal the transcriptional axis involved. Key axis protein levels in livers from healthy donors and patients with NAFLD were assessed via immunohistochemical staining. RESULTS: FOXA3 transcription is specifically induced by XBP1s upon ER stress. FOXA3 exacerbates the excessive lipid accumulation caused by the acute ER-inducer TM, whereas FOXA3 deficiency in hepatocytes and mice alleviates it. Importantly, FOXA3 deficiency in mice reduced diet-induced chronic ER stress, fatty liver, and insulin resistance. In addition, FOXA3 suppression via siRNA or adeno-associated virus delivery ameliorated the fatty liver phenotype in HFD-fed and db/db mice. Mechanistically, ChIP-Seq analysis revealed that FOXA3 directly regulates Period1 (Per1) transcription, which in turn promotes the expression of lipogenic genes, including Srebp1c, thus enhancing lipid synthesis. Of pathophysiological significance, FOXA3, PER1, and SREBP1c levels were increased in livers of obese mice and patients with NAFLD. CONCLUSION: The present study identified FOXA3 as the bridging molecule that links ER stress and NAFLD progression. Our results highlighted the role of the XBP1s-FOXA3-PER1/Srebp1c transcriptional axis in the development of NAFLD and identified FOXA3 as a potential therapeutic target for fatty liver disease. LAY SUMMARY: The molecular mechanisms linking endoplasmic reticulum stress to non-alcoholic fatty liver disease (NAFLD) progression remain undefined. Herein, via in vitro and in vivo analysis, we identified Forkhead box A3 (FOXA3) as a key bridging molecule. Of pathophysiological significance, FOXA3 protein levels were increased in livers of obese mice and patients with NAFLD, indicating that FOXA3 could be a potential therapeutic target in fatty liver disease.