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Chang HC, Shapiro JS, Jiang X, Senyei G, Sato T, Geier J, Sawicki KT, Ardehali H. Augmenter of liver regeneration regulates cellular iron homeostasis by modulating mitochondrial transport of ATP-binding cassette B8. eLife 2021; 10:e65158. [PMID: 33835027 PMCID: PMC8055271 DOI: 10.7554/elife.65158] [Citation(s) in RCA: 10] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/25/2020] [Accepted: 04/06/2021] [Indexed: 12/15/2022] Open
Abstract
Chronic loss of Augmenter of Liver Regeneration (ALR) results in mitochondrial myopathy with cataracts; however, the mechanism for this disorder remains unclear. Here, we demonstrate that loss of ALR, a principal component of the MIA40/ALR protein import pathway, results in impaired cytosolic Fe/S cluster biogenesis in mammalian cells. Mechanistically, MIA40/ALR facilitates the mitochondrial import of ATP-binding cassette (ABC)-B8, an inner mitochondrial membrane protein required for cytoplasmic Fe/S cluster maturation, through physical interaction with ABCB8. Downregulation of ALR impairs mitochondrial ABCB8 import, reduces cytoplasmic Fe/S cluster maturation, and increases cellular iron through the iron regulatory protein-iron response element system. Our finding thus provides a mechanistic link between MIA40/ALR import machinery and cytosolic Fe/S cluster maturation through the mitochondrial import of ABCB8, and offers a potential explanation for the pathology seen in patients with ALR mutations.
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Affiliation(s)
- Hsiang-Chun Chang
- Feinberg Cardiovascular and Renal Research Institute, Northwestern University School of MedicineChicagoUnited States
| | - Jason Solomon Shapiro
- Feinberg Cardiovascular and Renal Research Institute, Northwestern University School of MedicineChicagoUnited States
| | - Xinghang Jiang
- Feinberg Cardiovascular and Renal Research Institute, Northwestern University School of MedicineChicagoUnited States
| | - Grant Senyei
- Feinberg Cardiovascular and Renal Research Institute, Northwestern University School of MedicineChicagoUnited States
| | - Teruki Sato
- Feinberg Cardiovascular and Renal Research Institute, Northwestern University School of MedicineChicagoUnited States
| | - Justin Geier
- Feinberg Cardiovascular and Renal Research Institute, Northwestern University School of MedicineChicagoUnited States
| | - Konrad T Sawicki
- Feinberg Cardiovascular and Renal Research Institute, Northwestern University School of MedicineChicagoUnited States
| | - Hossein Ardehali
- Feinberg Cardiovascular and Renal Research Institute, Northwestern University School of MedicineChicagoUnited States
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Augmenter of liver regeneration: Essential for growth and beyond. Cytokine Growth Factor Rev 2018; 45:65-80. [PMID: 30579845 DOI: 10.1016/j.cytogfr.2018.12.003] [Citation(s) in RCA: 21] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/30/2018] [Revised: 12/12/2018] [Accepted: 12/14/2018] [Indexed: 12/11/2022]
Abstract
Liver regeneration is a well-orchestrated process that is triggered by tissue loss due to trauma or surgical resection and by hepatocellular death induced by toxins or viral infections. Due to the central role of the liver for body homeostasis, intensive research was conducted to identify factors that might contribute to hepatic growth and regeneration. Using a model of partial hepatectomy several factors including cytokines and growth factors that regulate this process were discovered. Among them, a protein was identified to specifically support liver regeneration and therefore was named ALR (Augmenter of Liver Regeneration). ALR protein is encoded by GFER (growth factor erv1-like) gene and can be regulated by various stimuli. ALR is expressed in different tissues in three isoforms which are associated with multiple functions: The long forms of ALR were found in the inner-mitochondrial space (IMS) and the cytosol. Mitochondrial ALR (23 kDa) was shown to cooperate with Mia40 to insure adequate protein folding during import into IMS. On the other hand short form ALR, located mainly in the cytosol, was attributed with anti-apoptotic and anti-oxidative properties as well as its inflammation and metabolism modulating effects. Although a considerable amount of work has been devoted to summarizing the knowledge on ALR, an investigation of ALR expression in different organs (location, subcellular localization) as well as delineation between the isoforms and function of ALR is still missing. This review provides a comprehensive evaluation of ALR structure and expression of different ALR isoforms. Furthermore, we highlight the functional role of endogenously expressed and exogenously applied ALR, as well as an analysis of the clinical importance of ALR, with emphasis on liver disease and in vivo models, as well as the consequences of mutations in the GFER gene.
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Gupta P, Venugopal SK. Augmenter of liver regeneration: A key protein in liver regeneration and pathophysiology. Hepatol Res 2018; 48:587-596. [PMID: 29633440 DOI: 10.1111/hepr.13077] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/15/2018] [Revised: 03/10/2018] [Accepted: 03/29/2018] [Indexed: 12/22/2022]
Abstract
Liver is constantly exposed to pathogens, viruses, chemicals, and toxins, and several of them cause injury, leading to the loss of liver mass and sometimes resulting in cirrhosis and cancer. Under physiological conditions, liver can regenerate if the loss of cells is less than the proliferation of hepatocytes. If the loss is more than the proliferation, the radical treatment available is liver transplantation. Due to this reason, the search for an alternative therapeutic agent has been the focus of liver research. Liver regeneration is regulated by several growth factors; one of the key factors is augmenter of liver regeneration (ALR). Involvement of ALR has been reported in crucial processes such as oxidative phosphorylation, maintenance of mitochondria and mitochondrial biogenesis, and regulation of autophagy and cell proliferation. Augmenter of liver regeneration has been observed to be involved in liver regeneration by not only overcoming cell cycle inhibition but by maintaining the stem cell pool as well. These observations have created curiosity regarding the possible role of ALR in maintenance of liver health. Thus, this review brings a concise presentation of the work done in areas exploring the role of ALR in normal liver physiology and in liver health maintenance by fighting liver diseases, such as liver failure, non-alcoholic fatty liver disease/non-alcoholic steatohepatitis, viral infections, cirrhosis, and hepatocellular carcinoma.
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Affiliation(s)
- Parul Gupta
- Faculty of Life Sciences and Biotechnology, South Asian University, New Delhi, India
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Augmenter of liver regeneration potentiates doxorubicin anticancer efficacy by reducing the expression of ABCB1 and ABCG2 in hepatocellular carcinoma. J Transl Med 2017; 97:1400-1411. [PMID: 28825695 DOI: 10.1038/labinvest.2017.72] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/12/2016] [Revised: 05/03/2017] [Accepted: 05/23/2017] [Indexed: 12/15/2022] Open
Abstract
Hepatocellular carcinoma (HCC) is highly chemoresistant and therefore challenges both physicians and patients. Augmenter of liver regeneration (ALR), previously also known as 'hepatic stimulator substance', is reported to inhibit the epithelial-mesenchymal transition (EMT) in HCC, one of the frequent events that occur in cancer metastasis, suggesting that ALR is involved in HCC. In this study, we report for the first time that the transfection of ALR enhances the antitumor effect of chemotherapy with doxorubicin, a typical anticancer drug, on HCC in vitro and in vivo. The efflux of doxorubicin from ALR-transfected HCC cells is efficiently suppressed. This implies the intracellular retention of doxorubicin in tumor cells, which is at least partly attributable to the effective inhibition of ABCB1 and ABCG2 transporter expression in ALR-expressing cells. The downregulation of ALR expression by short hairpin RNA diminishes the antitumor effect of ALR. We further demonstrate that ALR inhibits the AKT/Snail signaling pathway, resulting in the downregulation of ABCB1 and ABCG2 expression. In conclusion, our results suggest that ALR is a potential chemotherapeutic agent against HCC.
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Shen Y, Liu Q, Lou S, Luo Y, Sun H, Zeng H, Deng J. Decreased expression of the augmenter of liver regeneration results in growth inhibition and increased chemosensitivity of acute T lymphoblastic leukemia cells. Oncol Rep 2017; 38:3130-3136. [PMID: 29048676 DOI: 10.3892/or.2017.5984] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/23/2017] [Accepted: 08/17/2017] [Indexed: 11/06/2022] Open
Abstract
Augmenter of liver regeneration (ALR) plays crucial roles in cell survival and growth. Previous studies have demonstrated that ALR exerts a protective effect on toxic agent‑induced cell death in acute T lymphoblastic leukemia cells and ALR knockdown can sensitize cancer cells to radiation. However, the biological functions of ALR against drug resistance in T-cell acute lymphoblastic leukemia are mostly unknown. In the present study, we investigated the effect of small interfering RNA (siRNA)-induced ALR silencing on cell proliferation and sensitivity to vincristine (VCR) of Jurkat cells. We found that ALR siRNA effectively decreased the ALR expression, then inhibited cell growth and increased sensitivity to VCR in Jurkat cells. Flow cytometry assay revealed that the downregulation of ALR expression promoted cell apoptosis and regulated cell cycle distribution. Following incubation with VCR, apoptosis-related proteins, such as pro-PARP, pro-caspase 8, pro-caspase 3 and Bcl-2 were downregulated in the siRNA/ALR group. Pretreatment with siRNA/ALR in combination with VCR resulted in prolonged G2/M arrest, accompanied by downregulation of cdc25c and cdc2 expression and dissociation of cyclin B1. In conclusion, the results of this study demonstrated that targeted inhibition of the ALR expression in Jurkat cells triggered cell growth inhibition and sensitized cells to VCR via promoting apoptosis and regulating the cell cycle.
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Affiliation(s)
- Yan Shen
- Department of Hematology, The Second Affiliated Hospital, Chongqing Medical University, Chongqing 400010, P.R. China
| | - Qi Liu
- Institute for Viral Hepatitis, Key Laboratory of Molecular Biology for Infectious Diseases, The Second Affiliated Hospital, Chongqing Medical University, Chongqing 400010, P.R. China
| | - Shifeng Lou
- Department of Hematology, The Second Affiliated Hospital, Chongqing Medical University, Chongqing 400010, P.R. China
| | - Yun Luo
- Department of Hematology, The Second Affiliated Hospital, Chongqing Medical University, Chongqing 400010, P.R. China
| | - Hang Sun
- Institute for Viral Hepatitis, Key Laboratory of Molecular Biology for Infectious Diseases, The Second Affiliated Hospital, Chongqing Medical University, Chongqing 400010, P.R. China
| | - Hanqing Zeng
- Department of Hematology, The Second Affiliated Hospital, Chongqing Medical University, Chongqing 400010, P.R. China
| | - Jianchuan Deng
- Department of Hematology, The Second Affiliated Hospital, Chongqing Medical University, Chongqing 400010, P.R. China
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Weiss TS, Lupke M, Ibrahim S, Buechler C, Lorenz J, Ruemmele P, Hofmann U, Melter M, Dayoub R. Attenuated lipotoxicity and apoptosis is linked to exogenous and endogenous augmenter of liver regeneration by different pathways. PLoS One 2017; 12:e0184282. [PMID: 28877220 PMCID: PMC5587239 DOI: 10.1371/journal.pone.0184282] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/02/2017] [Accepted: 08/21/2017] [Indexed: 02/06/2023] Open
Abstract
Nonalcoholic fatty liver disease (NAFLD) covers a spectrum from simple steatosis to nonalcoholic steatohepatitis (NASH) and cirrhosis. Free fatty acids (FFA) induce steatosis and lipo-toxicity and correlate with severity of NAFLD. In this study we aimed to investigate the role of exogenous and endogenous ALR (augmenter of liver regeneration) for FFA induced ER (endoplasmatic reticulum) -stress and lipoapoptosis. Primary human hepatocytes or hepatoma cells either treated with recombinant human ALR (rhALR, 15kDa) or expressing short form ALR (sfALR, 15kDa) were incubated with palmitic acid (PA) and analyzed for lipo-toxicity, -apoptosis, activation of ER-stress response pathways, triacylglycerides (TAG), mRNA and protein expression of lipid metabolizing genes. Both, exogenous rhALR and cytosolic sfALR reduced PA induced caspase 3 activity and Bax protein expression and therefore lipotoxicity. Endogenous sfALR but not rhALR treatment lowered TAG levels, diminished activation of ER-stress mediators C-Jun N-terminal kinase (JNK), X-box binding protein-1 (XBP1) and proapoptotic transcription factor C/EBP-homologous protein (CHOP), and reduced death receptor 5 protein expression. Cellular ALR exerts its lipid lowering and anti-apoptotic actions by enhancing FABP1, which binds toxic FFA, increasing mitochondrial β-oxidation by elevating the mitochondrial FFA transporter CPT1α, and decreasing ELOVL6, which delivers toxic FFA metabolites. We found reduced hepatic mRNA levels of ALR in a high fat diet mouse model, and of ALR and FOXA2, a transcription factor inducing ALR expression, in human steatotic as well as NASH liver samples, which may explain increased lipid deposition and reduced β-oxidation in NASH patients. Present study shows that exogenous and endogenous ALR reduce PA induced lipoapoptosis. Furthermore, cytosolic sfALR changes mRNA and protein expression of genes regulating lipid metabolism, reduces ER-stress finally impeding progression of NASH.
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Affiliation(s)
- Thomas S. Weiss
- Children’s University Hospital, University of Regensburg, Regensburg, Germany
- Center for Liver Cell Research, University of Regensburg Hospital, Regensburg, Germany
- * E-mail:
| | - Madeleine Lupke
- Children’s University Hospital, University of Regensburg, Regensburg, Germany
| | - Sara Ibrahim
- Children’s University Hospital, University of Regensburg, Regensburg, Germany
| | - Christa Buechler
- Department of Internal Medicine, University of Regensburg Hospital, Regensburg, Germany
| | - Julia Lorenz
- Children’s University Hospital, University of Regensburg, Regensburg, Germany
| | - Petra Ruemmele
- Institute of Pathology, University Hospital Erlangen, Friedrich-Alexander University Erlangen-Nuernberg, Erlangen, Germany
| | - Ute Hofmann
- Dr. Margarete Fischer-Bosch Institute of Clinical Pharmacology and University of Tübingen, Stuttgart, Germany
| | - Michael Melter
- Children’s University Hospital, University of Regensburg, Regensburg, Germany
| | - Rania Dayoub
- Children’s University Hospital, University of Regensburg, Regensburg, Germany
- Department of Biochemistry and Microbiology, Faculty of Pharmacy, Damascus University, Damascus, Syria
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Nambot S, Gavrilov D, Thevenon J, Bruel A, Bainbridge M, Rio M, Goizet C, Rötig A, Jaeken J, Niu N, Xia F, Vital A, Houcinat N, Mochel F, Kuentz P, Lehalle D, Duffourd Y, Rivière J, Thauvin-Robinet C, Beaudet A, Faivre L. Further delineation of a rare recessive encephalomyopathy linked to mutations in GFER thanks to data sharing of whole exome sequencing data. Clin Genet 2017; 92:188-198. [DOI: 10.1111/cge.12985] [Citation(s) in RCA: 19] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/21/2016] [Revised: 12/24/2016] [Accepted: 01/25/2017] [Indexed: 02/06/2023]
Affiliation(s)
- S. Nambot
- Centre de Génétique et Centre de référence «Anomalies du Développement et Syndromes Malformatifs», Hôpital d'Enfants; Centre Hospitalier Universitaire de Dijon; Dijon France
- Laboratoire de Génétique Moléculaire, Plateau Technique de Biologie; Centre Hospitalier Universitaire de Dijon; Dijon France
| | - D. Gavrilov
- Biochemical Genetics Laboratory, Department of Laboratory Medicine and Pathology; Mayo Clinic College of Medicine; Rochester Minnesota
- Department of Genetics and Genomics; Mayo Clinic College of Medicine; Rochester Minnesota
| | - J. Thevenon
- Centre de Génétique et Centre de référence «Anomalies du Développement et Syndromes Malformatifs», Hôpital d'Enfants; Centre Hospitalier Universitaire de Dijon; Dijon France
- Fédération Hospitalo-Universitaire Médecine Translationnelle et Anomalies du Développement (FHU TRANSLAD); Centre Hospitalier Universitaire de Dijon et Université de Bourgogne-Franche Comté; Dijon France
- Génétique des Anomalies du Développement; Université de Bourgogne; Dijon France
| | - A.L. Bruel
- Laboratoire de Génétique Moléculaire, Plateau Technique de Biologie; Centre Hospitalier Universitaire de Dijon; Dijon France
- Génétique des Anomalies du Développement; Université de Bourgogne; Dijon France
| | - M. Bainbridge
- Human Genome Sequencing Center; Baylor College of Medicine; Houston Texas
| | - M. Rio
- Service de Génétique Médicale; Hôpital Necker Enfants Malades; Paris France
| | - C. Goizet
- Service de Génétique Médicale; Centre Hospitalier Universitaire de Bordeaux-GH Pellegrin; Bordeaux France
| | - A. Rötig
- Laboratoire de Génétique Moléculaire, Institut de Recherche Necker Enfants Malades; Hôpital Necker Enfants Malades; Paris France
| | - J. Jaeken
- Center for Metabolic Diseases; University Hospital Gasthuisberg; Leuven Belgium
| | - N. Niu
- Department of Molecular and Human Genetics; Baylor College of Medicine; Houston Texas
| | - F. Xia
- Department of Molecular and Human Genetics; Baylor College of Medicine; Houston Texas
| | - A. Vital
- Service de Pathologie, Pôle Biologie et Pathologie; Centre Hospitalier Universitaire de Bordeaux-GH Pellegrin; Bordeaux France
| | - N. Houcinat
- Centre de Génétique et Centre de référence «Anomalies du Développement et Syndromes Malformatifs», Hôpital d'Enfants; Centre Hospitalier Universitaire de Dijon; Dijon France
| | - F. Mochel
- Service de Génétique médicale; Centre Hospitalier Universitaire La Pitié Salpêtrière-Charles Foix; Paris France
| | - P. Kuentz
- Laboratoire de Génétique Moléculaire, Plateau Technique de Biologie; Centre Hospitalier Universitaire de Dijon; Dijon France
| | - D. Lehalle
- Centre de Génétique et Centre de référence «Anomalies du Développement et Syndromes Malformatifs», Hôpital d'Enfants; Centre Hospitalier Universitaire de Dijon; Dijon France
| | - Y. Duffourd
- Fédération Hospitalo-Universitaire Médecine Translationnelle et Anomalies du Développement (FHU TRANSLAD); Centre Hospitalier Universitaire de Dijon et Université de Bourgogne-Franche Comté; Dijon France
- Génétique des Anomalies du Développement; Université de Bourgogne; Dijon France
| | - J.B. Rivière
- Laboratoire de Génétique Moléculaire, Plateau Technique de Biologie; Centre Hospitalier Universitaire de Dijon; Dijon France
- Fédération Hospitalo-Universitaire Médecine Translationnelle et Anomalies du Développement (FHU TRANSLAD); Centre Hospitalier Universitaire de Dijon et Université de Bourgogne-Franche Comté; Dijon France
- Génétique des Anomalies du Développement; Université de Bourgogne; Dijon France
| | - C. Thauvin-Robinet
- Centre de Génétique et Centre de référence «Anomalies du Développement et Syndromes Malformatifs», Hôpital d'Enfants; Centre Hospitalier Universitaire de Dijon; Dijon France
- Fédération Hospitalo-Universitaire Médecine Translationnelle et Anomalies du Développement (FHU TRANSLAD); Centre Hospitalier Universitaire de Dijon et Université de Bourgogne-Franche Comté; Dijon France
- Génétique des Anomalies du Développement; Université de Bourgogne; Dijon France
| | - A.L. Beaudet
- Department of Molecular and Human Genetics; Baylor College of Medicine; Houston Texas
| | - L. Faivre
- Centre de Génétique et Centre de référence «Anomalies du Développement et Syndromes Malformatifs», Hôpital d'Enfants; Centre Hospitalier Universitaire de Dijon; Dijon France
- Fédération Hospitalo-Universitaire Médecine Translationnelle et Anomalies du Développement (FHU TRANSLAD); Centre Hospitalier Universitaire de Dijon et Université de Bourgogne-Franche Comté; Dijon France
- Génétique des Anomalies du Développement; Université de Bourgogne; Dijon France
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Yu HY, Zhu MH, Xiang DR, Li J, Sheng JF. High expression of 23 kDa protein of augmenter of liver regeneration (ALR) in human hepatocellular carcinoma. Onco Targets Ther 2014; 7:887-93. [PMID: 24940072 PMCID: PMC4051792 DOI: 10.2147/ott.s61531] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/20/2022] Open
Abstract
Background Augmenter of liver regeneration (ALR) is an important polypeptide that participates in the process of liver regeneration. Two forms of ALR proteins are expressed in hepatocytes. Previous data have shown that ALR is essential for cell survival and has potential antimetastatic properties in hepatocellular carcinoma (HCC). Aims The study aimed to evaluate the expression levels of two forms of ALR proteins in HCC and their possible significance in HCC development. Methods Balb/c mouse monoclonal antibody against ALR protein was prepared in order to detect the ALR protein in HCC by Western blotting and immunohistochemistry. ALR mRNA expression levels were measured by real-time polymerase chain reaction in HCC tissues and compared to paracancerous liver tissues in 22 HCC patients. Results ALR mRNA expression in HCC liver tissues (1.51×106 copies/μL) was higher than in paracancerous tissues (1.04×104 copies/μL). ALR protein expression was also enhanced in HCC liver tissues. The enhanced ALR protein was shown to be 23 kDa by Western blotting. Immunohistochemical analysis showed that the 23 kDa ALR protein mainly existed in the hepatocyte cytosol. Conclusion The 23 kDa ALR protein was highly expressed in HCC and may play an important role in hepatocarcinogenesis.
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Affiliation(s)
- Hai-Ying Yu
- State Key Laboratory of Infectious Disease and Department of Infectious Disease, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, People's Republic of China
| | - Man-Hua Zhu
- State Key Laboratory of Infectious Disease and Department of Infectious Disease, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, People's Republic of China
| | - Dai-Rong Xiang
- State Key Laboratory of Infectious Disease and Department of Infectious Disease, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, People's Republic of China
| | - Jun Li
- State Key Laboratory of Infectious Disease and Department of Infectious Disease, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, People's Republic of China
| | - Ji-Fang Sheng
- State Key Laboratory of Infectious Disease and Department of Infectious Disease, First Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, People's Republic of China
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The mitochondrial disulfide relay system: roles in oxidative protein folding and beyond. Int J Cell Biol 2013; 2013:742923. [PMID: 24348563 PMCID: PMC3848088 DOI: 10.1155/2013/742923] [Citation(s) in RCA: 35] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/09/2013] [Accepted: 10/01/2013] [Indexed: 12/31/2022] Open
Abstract
Disulfide bond formation drives protein import of most proteins of the mitochondrial intermembrane space (IMS). The main components of this disulfide relay machinery are the oxidoreductase Mia40 and the sulfhydryl oxidase Erv1/ALR. Their precise functions have been elucidated in molecular detail for the yeast and human enzymes in vitro and in intact cells. However, we still lack knowledge on how Mia40 and Erv1/ALR impact cellular and organism physiology and whether they have functions beyond their role in disulfide bond formation. Here we summarize the principles of oxidation-dependent protein import mediated by the mitochondrial disulfide relay. We proceed by discussing recently described functions of Mia40 in the hypoxia response and of ALR in influencing mitochondrial morphology and its importance for tissue development and embryogenesis. We also include a discussion of the still mysterious function of Erv1/ALR in liver regeneration.
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Shen Y, Liu Q, Sun H, Li X, Wang N, Guo H. Protective effect of augmenter of liver regeneration on vincristine-induced cell death in Jurkat T leukemia cells. Int Immunopharmacol 2013; 17:162-167. [PMID: 23810409 DOI: 10.1016/j.intimp.2013.05.030] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/14/2013] [Revised: 05/21/2013] [Accepted: 05/28/2013] [Indexed: 11/30/2022]
Abstract
Augmenter of liver regeneration (ALR) is a crucial factor in the process of proliferation of hepatocytes. Recently, it has been demonstrated that ALR plays an important role of anti-apoptosis in several cell lines, but the biological effects of ALR in acute T lymphoblastic leukemia have remained unclear. In this study, we investigated the effect of ALR on Jurkat T leukemia cell growth and survival. We found that ALR was up-regulated in Jurkat cells and could reduce the sensitivity of Jurkat cells to vincristine, but had a minimal effect on proliferation of Jurkat cells. Results from analysis of flow cytometry showed ALR attenuated apoptotic cells and inhibited G2/M-arrest in vincristine-treated Jurkat cells. Following incubation with ALR, an increase in pro-caspase8, pro-caspase3, pro-PARP and Bcl-2 levels was observed in vincristine-treated Jurkat cells. In summary, the results of this study demonstrate that ALR protects Jurkat T leukemia cells from vincristine-induced cell death via regulation of apoptotic signaling pathways and cell cycle.
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Affiliation(s)
- Yan Shen
- Chongqing Medical University, Chongqing, China
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Decreased expression of the augmenter of liver regeneration results in increased apoptosis and oxidative damage in human-derived glioma cells. Cell Death Dis 2012; 3:e289. [PMID: 22476097 PMCID: PMC3358005 DOI: 10.1038/cddis.2012.25] [Citation(s) in RCA: 29] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/08/2023]
Abstract
The mammalian growth factor erv1-like (GFER) gene encodes a sulfhydryl oxidase enzyme, named Augmenter of Liver Regeneration (ALR). Recently it has been demonstrated that ALR supports cell proliferation acting as an anti-apoptotic factor. This effect is determined by ALR ability to support the anti-apoptotic gene expression and to preserve cellular normoxic conditions. We recently demonstrated that the addition of recombinant ALR (rALR) in the culture medium of H2O2-treated neuroblastoma cells reduces the lethal effects induced by the hydrogen peroxide. Similar data have been reported in the regenerating liver tissue from partially hepatectomized rats treated with rALR. The purpose of the present study was to evaluate the effect of the GFER inhibition, via the degradation of the complementary mRNA by the specific siRNA, on the behaviour of the apoptosis (apoptotic gene and caspase expression and apoptotic cell number) and of the oxidative stress-induced parameters (reactive oxygen species (ROS), clusterin expression and mitochondrial integrity) in T98G glioma cells. The results revealed a reduction of (i) ALR, (ii) clusterin and (iii) bcl-2 and an increase of (iv) caspase-9, activated caspase-3, ROS, apoptotic cell number and mitochondrial degeneration. These data confirm the anti-apoptotic role of ALR and its anti-oxidative properties, and shed some light on the molecular pathways through which ALR modulates its biological effects.
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12
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Li Y, Farooq M, Sheng D, Chandramouli C, Lan T, Mahajan NK, Kini RM, Hong Y, Lisowsky T, Ge R. Augmenter of liver regeneration (alr) promotes liver outgrowth during zebrafish hepatogenesis. PLoS One 2012; 7:e30835. [PMID: 22292055 PMCID: PMC3266923 DOI: 10.1371/journal.pone.0030835] [Citation(s) in RCA: 28] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/20/2011] [Accepted: 12/29/2011] [Indexed: 02/06/2023] Open
Abstract
Augmenter of Liver Regeneration (ALR) is a sulfhydryl oxidase carrying out fundamental functions facilitating protein disulfide bond formation. In mammals, it also functions as a hepatotrophic growth factor that specifically stimulates hepatocyte proliferation and promotes liver regeneration after liver damage or partial hepatectomy. Whether ALR also plays a role during vertebrate hepatogenesis is unknown. In this work, we investigated the function of alr in liver organogenesis in zebrafish model. We showed that alr is expressed in liver throughout hepatogenesis. Knockdown of alr through morpholino antisense oligonucleotide (MO) leads to suppression of liver outgrowth while overexpression of alr promotes liver growth. The small-liver phenotype in alr morphants results from a reduction of hepatocyte proliferation without affecting apoptosis. When expressed in cultured cells, zebrafish Alr exists as dimer and is localized in mitochondria as well as cytosol but not in nucleus or secreted outside of the cell. Similar to mammalian ALR, zebrafish Alr is a flavin-linked sulfhydryl oxidase and mutation of the conserved cysteine in the CxxC motif abolishes its enzymatic activity. Interestingly, overexpression of either wild type Alr or enzyme-inactive Alr(C131S) mutant promoted liver growth and rescued the liver growth defect of alr morphants. Nevertheless, alr(C131S) is less efficacious in both functions. Meantime, high doses of alr MOs lead to widespread developmental defects and early embryonic death in an alr sequence-dependent manner. These results suggest that alr promotes zebrafish liver outgrowth using mechanisms that are dependent as well as independent of its sulfhydryl oxidase activity. This is the first demonstration of a developmental role of alr in vertebrate. It exemplifies that a low-level sulfhydryl oxidase activity of Alr is essential for embryonic development and cellular survival. The dose-dependent and partial suppression of alr expression through MO-mediated knockdown allows the identification of its late developmental role in vertebrate liver organogenesis.
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Affiliation(s)
- Yan Li
- Department of Biological Sciences, National University of Singapore, Singapore, Singapore
| | - Muhammad Farooq
- Department of Biological Sciences, National University of Singapore, Singapore, Singapore
- Department of Zoology, College of Science, King Saud University, Riyadh, Kingdom of Saudi Arabia
| | - Donglai Sheng
- Department of Biological Sciences, National University of Singapore, Singapore, Singapore
| | - Chanchal Chandramouli
- Department of Biological Sciences, National University of Singapore, Singapore, Singapore
| | - Tian Lan
- Department of Biological Sciences, National University of Singapore, Singapore, Singapore
| | - Nilesh K. Mahajan
- Department of Biological Sciences, National University of Singapore, Singapore, Singapore
| | - R. Manjunatha Kini
- Department of Biological Sciences, National University of Singapore, Singapore, Singapore
- Department of Biochemistry and Molecular Biology, Medical College of Virginia, Virginia Commonwealth University, Richmond, Virginia, United States of America
| | - Yunhan Hong
- Department of Biological Sciences, National University of Singapore, Singapore, Singapore
| | | | - Ruowen Ge
- Department of Biological Sciences, National University of Singapore, Singapore, Singapore
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Molecular recognition and substrate mimicry drive the electron-transfer process between MIA40 and ALR. Proc Natl Acad Sci U S A 2011; 108:4811-6. [PMID: 21383138 DOI: 10.1073/pnas.1014542108] [Citation(s) in RCA: 87] [Impact Index Per Article: 6.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/19/2022] Open
Abstract
Oxidative protein folding in the mitochondrial intermembrane space requires the transfer of a disulfide bond from MIA40 to the substrate. During this process MIA40 is reduced and regenerated to a functional state through the interaction with the flavin-dependent sulfhydryl oxidase ALR. Here we present the mechanistic basis of ALR-MIA40 interaction at atomic resolution by biochemical and structural analyses of the mitochondrial ALR isoform and its covalent mixed disulfide intermediate with MIA40. This ALR isoform contains a folded FAD-binding domain at the C-terminus and an unstructured, flexible N-terminal domain, weakly and transiently interacting one with the other. A specific region of the N-terminal domain guides the interaction with the MIA40 substrate binding cleft (mimicking the interaction of the substrate itself), without being involved in the import of ALR. The hydrophobicity-driven binding of this region ensures precise protein-protein recognition needed for an efficient electron transfer process.
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14
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Dong LY, Sun G, Jiang L, Shao L, Hu Y, Jiang Y, Wang Y, An W. Epidermal growth factor down-regulates the expression of human hepatic stimulator substance via CCAAT/enhancer-binding protein β in HepG2 cells. Biochem J 2010; 431:277-287. [PMID: 20690902 DOI: 10.1042/bj20100671] [Citation(s) in RCA: 13] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/05/2025]
Abstract
hHSS (human hepatic stimulator substance), acting as a hepatotrophic growth factor, promotes liver regeneration. However, the regulatory mechanisms for hHSS transcription are still poorly understood. In the present study, we investigated transcription of hHSS triggered by EGF (epidermal growth factor) and the role of C/EBPβ (CCAAT/enhancer-binding protein β) as a potential core factor responsible for hHSS transcription in HepG2 cells. The results show that EGF suppresses hHSS mRNA expression at early time points. Using a promoter deletion assay, we identified a proximal region (-358/-212) that is required for EGF suppression. Overexpression of C/EBPβ enhances EGF suppression of hHSS, and mutation of the C/EBPβ-binding site at -292/-279 or siRNA (short interfering RNA) interference abolishes EGF suppression. Furthermore, using an electrophoretic mobility-shift assay and chromatin immunoprecipitation analysis, we found that C/EBPβ specifically binds to the -292/-279 site that is responsible for EGF inhibition. Moreover, using a knockin (overexpression) and knockdown strategy (siRNA), we confirmed that C/EBPβ is a key factor responsible for inhibition of hHSS mRNA expression. Pre-treatment with an inhibitor of JNK (c-Jun N-terminal kinase) or down-regulation of JNK1 with specific siRNA reverses EGF-inhibited hHSS expression. Our results provide a crucial regulatory mechanism for EGF in hHSS transcription within the promoter proximal region.
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Affiliation(s)
- Ling-Yue Dong
- Department of Cell Biology, Municipal Laboratory of Liver Protection, Regulation and Regeneration, Capital Medical University, Beijing, China
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15
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Daithankar VN, Schaefer SA, Dong M, Bahnson BJ, Thorpe C. Structure of the human sulfhydryl oxidase augmenter of liver regeneration and characterization of a human mutation causing an autosomal recessive myopathy . Biochemistry 2010; 49:6737-45. [PMID: 20593814 DOI: 10.1021/bi100912m] [Citation(s) in RCA: 44] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/11/2022]
Abstract
The sulfhydryl oxidase augmenter of liver regeneration (ALR) binds FAD in a helix-rich domain that presents a CxxC disulfide proximal to the isoalloxazine ring of the flavin. Head-to-tail interchain disulfide bonds link subunits within the homodimer of both the short, cytokine-like, form of ALR (sfALR), and a longer form (lfALR) which resides in the mitochondrial intermembrane space (IMS). lfALR has an 80-residue N-terminal extension with an additional CxxC motif required for the reoxidation of reduced Mia40 during oxidative protein folding within the IMS. Recently, Di Fonzo et al. [Di Fonzo, A., Ronchi, D., Lodi, T., Fassone, E., Tigano, M., Lamperti, C., Corti, S., Bordoni, A., Fortunato, F., Nizzardo, M., Napoli, L., Donadoni, C., Salani, S., Saladino, F., Moggio, M., Bresolin, N., Ferrero, I., and Comi, G. P. (2009) Am. J. Hum. Genet. 84, 594-604] described an R194H mutation of human ALR that led to cataract, progressive muscle hypotonia, and hearing loss in three children. The current work presents a structural and enzymological characterization of the human R194H mutant in lf- and sfALR. A crystal structure of human sfALR was determined by molecular replacement using the rat sfALR structure. R194 is located at the subunit interface of sfALR, close to the intersubunit disulfide bridges. The R194 guanidino moiety participates in three H-bonds: two main-chain carbonyl oxygen atoms (from R194 itself and from C95 of the intersubunit disulfide of the other protomer) and with the 2'-OH of the FAD ribose. The R194H mutation has minimal effect on the enzyme activity using model and physiological substrates of short and long ALR forms. However, the mutation adversely affects the stability of both ALR forms: e.g., by decreasing the melting temperature by about 10 degrees C, by increasing the rate of dissociation of FAD from the holoenzyme by about 45-fold, and by strongly enhancing the susceptibility of sfALR to partial proteolysis and to reduction of its intersubunit disulfide bridges by glutathione. Finally, a comparison of the TROSY-HSQC 2D NMR spectra of wild-type sfALR and its R194H mutant reveals a significant increase in conformational flexibility in the mutant protein. In sum, these in vitro data document the major impact of the seemingly conservative R194H mutation on the stability of dimeric ALR and complement the in vivo observations of Di Fonzo et al.
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Affiliation(s)
- Vidyadhar N Daithankar
- Department of Chemistry and Biochemistry, University of Delaware, Newark, Delaware 19716, USA
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16
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Dayoub R, Groitl P, Dobner T, Bosserhoff AK, Schlitt HJ, Weiss TS. Foxa2 (HNF-3beta) regulates expression of hepatotrophic factor ALR in liver cells. Biochem Biophys Res Commun 2010; 395:465-70. [PMID: 20382118 DOI: 10.1016/j.bbrc.2010.04.023] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/01/2010] [Accepted: 04/03/2010] [Indexed: 12/11/2022]
Abstract
Liver regeneration is a multistep and well-orchestrated process which is initiated by injuries such as tissue loss, infectious or toxic insults. Augmenter of liver regeneration (ALR) is a hepatotrophic growth factor which has been shown to stimulate hepatic regeneration after partial hepatectomy and therefore seems to be regulated during the regenerative process in the liver. Our aim was to analyze how ALR is regulated in hepatic tissues and which transcription factors might regulate its tissue-specific expression. Promoter studies of ALR (-733/+527 bp) revealed potential regulatory elements for various transcription factors like Foxa2, IL-6 RE-BP and C/EBPbeta. Analysis of the promoter activity by performing luciferase assays revealed that co-transfection with Foxa2 significantly induced the activity of ALR promoter in HepG2 cells. EMSA and Supershift analysis using anti-Foxa2 antibody confirmed the specific binding of Foxa2 to ALR promoter and this binding was inducible when the cells were simultaneously stimulated with IL-6. The increased binding after activation with IL-6 and/or Foxa2 was confirmed by elevated ALR protein levels using Western blot technique. In addition, we could not detect any binding of C/EBPbeta and IL-6 RE-BP to the promoter of ALR. In conclusion, these results indicate that ALR is regulated by Foxa2, and this regulation may be amplified by IL-6.
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Affiliation(s)
- Rania Dayoub
- Center for Liver Cell Research, University Medical Center Regensburg, Germany
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17
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Ishii J, Fukuda N, Tanaka T, Ogino C, Kondo A. Protein-protein interactions and selection: yeast-based approaches that exploit guanine nucleotide-binding protein signaling. FEBS J 2010; 277:1982-95. [DOI: 10.1111/j.1742-4658.2010.07625.x] [Citation(s) in RCA: 26] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/30/2022]
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Daithankar VN, Farrell SR, Thorpe C. Augmenter of liver regeneration: substrate specificity of a flavin-dependent oxidoreductase from the mitochondrial intermembrane space. Biochemistry 2009; 48:4828-37. [PMID: 19397338 DOI: 10.1021/bi900347v] [Citation(s) in RCA: 59] [Impact Index Per Article: 3.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/01/2023]
Abstract
Augmenter of liver regeneration (ALR) is both a growth factor and a sulfhydryl oxidase that binds FAD in an unusual helix-rich domain containing a redox-active CxxC disulfide proximal to the flavin ring. In addition to the cytokine form of ALR (sfALR) that circulates in serum, a longer form, lfALR, is believed to participate in oxidative trapping of reduced proteins entering the mitochondrial intermembrane space (IMS). This longer form has an 80-residue N-terminal extension containing an additional, distal, CxxC motif. This work presents the first enzymological characterization of human lfALR. The N-terminal region conveys no catalytic advantage toward the oxidation of the model substrate dithiothreitol (DTT). In addition, a C71A or C74A mutation of the distal disulfide does not increase the turnover number toward DTT. Unlike Erv1p, the yeast homologue of lfALR, static spectrophotometric experiments with the human oxidase provide no evidence of communication between distal and proximal disulfides. An N-terminal His-tagged version of human Mia40, a resident oxidoreductase of the IMS and a putative physiological reductant of lfALR, was subcloned and expressed in Escherichia coli BL21 DE3 cells. Mia40, as isolated, shows a visible spectrum characteristic of an Fe-S center and contains 0.56 +/- 0.02 atom of iron per subunit. Treatment of Mia40 with guanidine hydrochloride and triscarboxyethylphosphine hydrochloride during purification removed this chromophore. The resulting protein, with a reduced CxC motif, was a good substrate of lfALR. However, neither sfALR nor lfALR mutants lacking the distal disulfide could oxidize reduced Mia40 efficiently. Thus, catalysis involves a flow of reducing equivalents from the reduced CxC motif of Mia40 to distal and then proximal CxxC motifs of lfALR to the flavin ring and, finally, to cytochrome c or molecular oxygen.
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Affiliation(s)
- Vidyadhar N Daithankar
- Department of Chemistry and Biochemistry, University of Delaware, Newark, Delaware 19716, USA
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19
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Di Fonzo A, Ronchi D, Lodi T, Fassone E, Tigano M, Lamperti C, Corti S, Bordoni A, Fortunato F, Nizzardo M, Napoli L, Donadoni C, Salani S, Saladino F, Moggio M, Bresolin N, Ferrero I, Comi GP. The mitochondrial disulfide relay system protein GFER is mutated in autosomal-recessive myopathy with cataract and combined respiratory-chain deficiency. Am J Hum Genet 2009; 84:594-604. [PMID: 19409522 DOI: 10.1016/j.ajhg.2009.04.004] [Citation(s) in RCA: 109] [Impact Index Per Article: 6.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/02/2009] [Revised: 03/13/2009] [Accepted: 04/08/2009] [Indexed: 11/28/2022] Open
Abstract
A disulfide relay system (DRS) was recently identified in the yeast mitochondrial intermembrane space (IMS) that consists of two essential components: the sulfhydryl oxidase Erv1 and the redox-regulated import receptor Mia40. The DRS drives the import of cysteine-rich proteins into the IMS via an oxidative folding mechanism. Erv1p is reoxidized within this system, transferring its electrons to molecular oxygen through interactions with cytochrome c and cytochrome c oxidase (COX), thereby linking the DRS to the respiratory chain. The role of the human Erv1 ortholog, GFER, in the DRS has been poorly explored. Using homozygosity mapping, we discovered that a mutation in the GFER gene causes an infantile mitochondrial disorder. Three children born to healthy consanguineous parents presented with progressive myopathy and partial combined respiratory-chain deficiency, congenital cataract, sensorineural hearing loss, and developmental delay. The consequences of the mutation at the level of the patient's muscle tissue and fibroblasts were 1) a reduction in complex I, II, and IV activity; 2) a lower cysteine-rich protein content; 3) abnormal ultrastructural morphology of the mitochondria, with enlargement of the IMS space; and 4) accelerated time-dependent accumulation of multiple mtDNA deletions. Moreover, the Saccharomyces cerevisiae erv1(R182H) mutant strain reproduced the complex IV activity defect and exhibited genetic instability of the mtDNA and mitochondrial morphological defects. These findings shed light on the mechanisms of mitochondrial biogenesis, establish the role of GFER in the human DRS, and promote an understanding of the pathogenesis of a new mitochondrial disease.
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Affiliation(s)
- Alessio Di Fonzo
- Dino Ferrari Centre, Department of Neurological Sciences, University of Milan, 20122 Milan, Italy
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Chu ZL, Wang J, Yang YS, Tang LJ, Chen H, Yang XM, Wang SY. Identification of interaction between hepatopoietin 205 and cytochrome C. Shijie Huaren Xiaohua Zazhi 2008; 16:1281-1286. [DOI: 10.11569/wcjd.v16.i12.1281] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
AIM: To identify the interaction between hepatopoietin 205 (HPO205) and cytochrome C (Cytc).
METHODS: The coding genes of HPO205 and Cytc, amplified by polymerase chain reaction, were cloned into pDBLeu and pPC86 vector respectively. The interaction was confirmed by co-transformation with the recombinant plasmids into MaV203 of yeast two-hybrid system (Y2H), and verified by GST-Pull down assay simultaneously.
RESULTS: The coding genes of HPO205 and Cytc were successfully cloned into relevant vectors, and the obtained vectors were named as pDBLeu-GRER, pDBLeu-CYCS, pPC86-GFER and pPC86-CYCS. After Y2H identification, we found that co-transformation of pDBLeu-GFER pPC86-CYCS activated reporter genes Ura and His, but co-transformation of pDBLeu-CYCS and pPC86-GFER activated no reporter genes. GST-Pull down assay showed that HPO205 was deposited by GST-CYCS, but not by GST, verifying the interaction between HPO205 and Cytc.
CONCLUSION: The interaction between HPO205 and Cytc suggests that HPO205 participates in the biology processes of electron transfer or (and) apoptosis via Cytc.
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21
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Fass D. The Erv family of sulfhydryl oxidases. BIOCHIMICA ET BIOPHYSICA ACTA-MOLECULAR CELL RESEARCH 2007; 1783:557-66. [PMID: 18155671 DOI: 10.1016/j.bbamcr.2007.11.009] [Citation(s) in RCA: 84] [Impact Index Per Article: 4.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Received: 09/21/2007] [Revised: 11/18/2007] [Accepted: 11/20/2007] [Indexed: 10/22/2022]
Abstract
The Erv flavoenzymes contain a compact module that catalyzes the pairing of cysteine thiols into disulfide bonds. High-resolution structures of plant, animal, and fungal Erv enzymes that function in different contexts and intracellular compartments have been determined. Structural features can be correlated with biochemical properties, revealing how core sulfhydryl oxidase activity has been tailored to various functional niches. The introduction of disulfides into cysteine-containing substrates by Erv sulfhydryl oxidases is compared with the mechanisms used by NADPH-driven disulfide reductases and thioredoxin-like oxidoreductases to reduce and transfer disulfides, respectively.
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Affiliation(s)
- Deborah Fass
- Department of Structural Biology, Weizmann Institute of Science, Rehovot 76100, Israel.
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22
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Gatzidou E, Kouraklis G, Theocharis S. Insights on augmenter of liver regeneration cloning and function. World J Gastroenterol 2006; 12:4951-8. [PMID: 16937489 PMCID: PMC4087396 DOI: 10.3748/wjg.v12.i31.4951] [Citation(s) in RCA: 32] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/19/2005] [Revised: 06/12/2006] [Accepted: 06/18/2006] [Indexed: 02/06/2023] Open
Abstract
Hepatic stimulator substance (HSS) has been referred to as a liver-specific but species non-specific growth factor. Gradient purification and sequence analysis of HSS protein indicated that it contained the augmenter of liver regeneration (ALR), also known as hepatopoietin (HPO). ALR, acting as a hepatotrophic growth factor, specifically stimulated proliferation of cultured hepatocytes as well as hepatoma cells in vitro, promoted liver regeneration and recovery of damaged hepatocytes and rescued acute hepatic failure in vivo. ALR belongs to the new Erv1/Alr protein family, members of which are found in lower and higher eukaryotes from yeast to man and even in some double-stranded DNA viruses. The present review article focuses on the molecular biology of ALR, examining the ALR gene and its expression from yeast to man and the biological function of ALR protein. ALR protein seems to be non-liver-specific as was previously believed, increasing the necessity to extend research on mammalian ALR protein in different tissues, organs and developmental stages in conditions of normal and abnormal cellular growth.
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Affiliation(s)
- Elisavet Gatzidou
- Department of Forensic Medicine and Toxicology, University of Athens, Medical School, GR11527, Athens, Greece
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Dong LY, Wang XN, Song ZG, Guo D, Zhao YY, An W. Identification of human hepatic stimulator substance gene promoter and demonstration of dual regulation of AP1/AP4 cis-acting element in different cell lines. Int J Biochem Cell Biol 2006; 39:181-96. [PMID: 16978907 DOI: 10.1016/j.biocel.2006.07.007] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/16/2006] [Revised: 07/14/2006] [Accepted: 07/31/2006] [Indexed: 11/18/2022]
Abstract
Human hepatic stimulator substance (hHSS) is a newly identified growth-promoting factor in the liver. HSS is capable of stimulating hepatic regeneration in partial hepatectomized rats, thus, promoting growth of hepatic tumor cells. To understand and elucidate the transcriptional regulation of hHSS gene, the 4890bp of 5'-flanking region of the gene have been isolated and sequenced. The transcriptional start site, located at 248nt upstream from the ATG starting codon, was identified by 5'-rapid amplification cDNA end (5'-RACE). The classical promoter sequences, such as TATA box or GAATT were not identified in the promoter region, instead a GC-rich segment was formed (>70%) by expanding to a longer than 400bp, and immediately upstream from the ATG start codon. The transient transfection assays, using promoter deletion constructs, showed that hHSS promoter was efficiently capable in driving the reporter expression not only in HepG2 cells, but also in Cos7 cells. A region spanning nucleotides in the range of -447 to -358bp revealed a negative regulation on promoter activity in HepG2 cells, but with positive regulation in Cos7 and Hela cells. The promoter activity was obviously influenced by AP1/AP4 (-375/-369nt) mutation in these three cell lines. EMSAs showed that the site was recognized by AP1 in HepG2 cell, and only by an AP4 protein in Cos7 cells. The c-Jun bound to the promoter was further verified by supershift in HepG2 cells and human liver tissue. Chromatin immuno-precipitation (ChIP) demonstrated that there was a direct association of c-Jun with hHSS promoter in HepG2 cells. The c-Jun strongly suppressed hHSS promoter activity in transient expression analyses in HepG2 cells. Mutations in the AP1 binding sites rescued suppression caused by c-Jun, suggesting this was a direct regulation of the hHSS promoter. In contrast, there was no significant effect in c-Jun over-expressed Cos7 and Hela cells. The tissue-specific function of c-Jun in hHSS promoter activity may in part help explain the differences in biology function of hHSS between liver and non-liver cells.
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Affiliation(s)
- Ling-Yue Dong
- Department of Cell Biology and Municipal Laboratory for Liver Protection and Regulation of Regeneration, Capital Medical University, 10 You An Men Wai Xi Tou Tiao, Beijing 100069, China
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Pawlowski R, Jura J. ALR and Liver Regeneration. Mol Cell Biochem 2006; 288:159-69. [PMID: 16691313 DOI: 10.1007/s11010-006-9133-7] [Citation(s) in RCA: 36] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/04/2005] [Accepted: 01/10/2006] [Indexed: 12/18/2022]
Abstract
Liver possesses the capacity to restore its tissue mass and attain optimal volume in response to physical, infectious and toxic injury. The extraordinary ability of liver to regenerate is the effect of cross-talk between growth factors, cytokines, matrix components and many other factors. In this review we present recent findings and existing information about mechanisms that regulate liver growth, paying attention to augmenter of liver regeneration.
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Affiliation(s)
- Rafał Pawlowski
- Department of Cell Biochemistry, Faculty of Biotechnology, Jagiellonian University, Gronostajowa 7, 30-387, Krakow, Poland
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25
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Thasler WE, Schlott T, Thelen P, Hellerbrand C, Bataille F, Lichtenauer M, Schlitt HJ, Jauch KW, Weiss TS. Expression of augmenter of liver regeneration (ALR) in human liver cirrhosis and carcinoma. Histopathology 2005; 47:57-66. [PMID: 15982324 DOI: 10.1111/j.1365-2559.2005.02172.x] [Citation(s) in RCA: 53] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/29/2022]
Abstract
AIMS To determine the expression of a protein termed augmenter of liver regeneration (ALR), recently found to have a specific and beneficial effect on the process of liver regeneration in normal and diseased human liver. METHODS AND RESULTS ALR expression in normal and cirrhotic human livers with various underlying diseases as well as in tissue samples of hepatocellular carcinoma (HCC) and cholangiocellular carcinoma (CCC) was analysed by immunohistochemistry and quantitative reverse transciptase-polymerase chain reaction (RT-PCR). Expression analysis of ALR in total liver protein extracts by Western blotting showed mainly dimeric ALR protein. Immunohistochemically, cytosolic and perinuclear immunosignals were found in hepatocytes and cholangiocytes in normal, cirrhotic or cancerous liver tissue and only weak signals in some endothelial cells in normal livers. Quantitative mRNA analysis revealed significantly increased ALR expression in cirrhosis compared with normal liver tissue. In HCC and CCC ALR mRNA expression was also significantly enhanced compared with normal liver tissue, but expression levels did not differ from the matching non-neoplastic tissue in the same patient. CONCLUSIONS The findings suggest an important role for ALR in hepatocellular regeneration in liver cirrhosis as well as in hepatocarcinogenesis and therefore its potential value in the clinical diagnosis of hepatic cirrhosis and cancer.
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Affiliation(s)
- W E Thasler
- Department of Surgery, Ludwig Maximillians University of Munich Hospital Grosshadern, Munich, Germany
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Wang Y, Lu C, Wei H, Wang N, Chen X, Zhang L, Zhai Y, Zhu Y, Lu Y, He F. Hepatopoietin interacts directly with COP9 signalosome and regulates AP-1 activity. FEBS Lett 2004; 572:85-91. [PMID: 15304329 DOI: 10.1016/j.febslet.2004.07.012] [Citation(s) in RCA: 18] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/26/2004] [Accepted: 07/01/2004] [Indexed: 01/18/2023]
Abstract
Hepatopoietin (HPO)/augmenter of liver regeneration (ALR) is a specific hepatotrophic growth factor, which plays a key role in liver regeneration. Our previous study indicated that HPO executes its function by an inter-reactive network of the autocrine, paracrine and endocrine pathways. Recently, we have demonstrated that intracellular HPO interacts with Jun activation domain-binding protein 1 (JAB1) and leads to potentiation of activating protein-1 (AP-1) activity in a MAPK independent fashion. JAB1 is the fifth subunit of the COP9 signalosome (CSN), which is first identified as a suppressor of plant morphogenesis. A protein complex kinase activity associated with the CSN has been reported but not identified yet. In this report, we investigated further the association of HPO with the whole CSN. HPO exists in a complex with the eight-component CSN, both when purified from glycerol gradient centrifugation and when reciprocal immunoprecipitated from the lysates of transfected COS-7 cells. Intracellular HPO colocalizes with endogenous CSN in nucleus of hepatic cells. In addition, intracellular function of HPO that increases the phosphorylation of c-Jun leading to potentiate the AP-1 activity is inhibited by curcumin, a potent inhibitor of CSN-associated kinase. Taken together, these results elucidate a novel relationship of intracellular growth factor, HPO with large protein complex, CSN, which suggests a possible linkage between CSN and liver regeneration.
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Affiliation(s)
- Yan Wang
- Department of Genomics and Proteomics, Beijing Institute of Radiation Medicine, Chinese National Human Genome Center, 27 Taiping Road, Beijing 100850, PR China
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Abstract
The advent of proteomics techniques has been enthusiastically accepted in most areas of biology and medicine. In neuroscience, a host of applications was proposed ranging from neurotoxicology, neurometabolism, determination of the proteome of the individual brain areas in health and disease, to name a few. Only recently, the limitations of the method have been shown, hampering the rapid spreading of the technology, which in principle consists of two-dimensional gel electrophoresis with in-gel protein digestion of protein spots and identification by mass-spectrometrical approaches or microsequencing. The identification, including quantification using specific software, of brain protein classes, like enzymes, cytoskeleton proteins, heat shock proteins/chaperones, proteins of the transcription and translation machinery, synaptosomal proteins, antioxidant proteins, is a clear domain of proteomics. Furthermore, the concomitant detection of several hundred proteins on a gel allows the demonstration of an expressional pattern, rather generated by a reliable, protein-chemical method than by immunoreactivity, proposed by protein-arrays. An additional advantage is that hitherto unknown proteins, so far only proposed from their nucleic acid structure, designated as hypothetical proteins, can be identified as brain proteins. As to shortcomings and disadvantages of the method we would point to the major problem, the failure to separate hydrophobic proteins. There is so far no way to analyse the vast majority of these proteins in gels. Several other analytical problems need to be overcome, but once the latter problem can be solved, there is nothing to stop the method for a large scale analysis of membrane proteins in neuroscience.
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Affiliation(s)
- Gert Lubec
- Department of Pediatrics, University of Vienna, Währinger Gürtel 18, A 1090, Vienna, Austria.
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