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1
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Wu Z, Cardona EA, Cohn JA, Pierce JT. Nonapoptotic role of EGL-1 in exopher production and neuronal health in Caenorhabditis elegans. Proc Natl Acad Sci U S A 2025; 122:e2407909122. [PMID: 39786930 PMCID: PMC11745333 DOI: 10.1073/pnas.2407909122] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/22/2024] [Accepted: 11/20/2024] [Indexed: 01/30/2025] Open
Abstract
While traditionally studied for their proapoptotic functions in activating the caspase, research suggests BH3-only proteins also have other roles such as mitochondrial dynamics regulation. Here, we find that EGL-1, the BH3-only protein in Caenorhabditis elegans, promotes the cell-autonomous production of exophers in adult neurons. Exophers are large, micron-scale vesicles that are ejected from the cell and contain cellular components such as mitochondria. EGL-1 facilitates exopher production potentially through regulation of mitochondrial dynamics. Moreover, an endogenous, low level of EGL-1 expression appears to benefit dendritic health. Our findings provide insights into the role of neuronal BH3-only protein in mitochondrial dynamics, downstream exopher production, and ultimately neuronal health.
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Affiliation(s)
- Zheng Wu
- Department of Neuroscience, Center for Learning and Memory, Waggoner Center for Alcohol & Addiction Research, University of Texas at Austin, Austin, TX78712
| | - Eric A. Cardona
- Department of Neuroscience, Center for Learning and Memory, Waggoner Center for Alcohol & Addiction Research, University of Texas at Austin, Austin, TX78712
| | - Jesse A. Cohn
- Department of Neuroscience, Center for Learning and Memory, Waggoner Center for Alcohol & Addiction Research, University of Texas at Austin, Austin, TX78712
| | - Jonathan T. Pierce
- Department of Neuroscience, Center for Learning and Memory, Waggoner Center for Alcohol & Addiction Research, University of Texas at Austin, Austin, TX78712
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2
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Zhang M, Mouzannar K, Zhang Z, Teraoka Y, Piotrowski J, Ishida Y, Tateno-Mukaidani C, Saito T, Abe-Chayama H, Chayama K, Liang TJ. Hepatitis B virus genotypes A1 and A2 have distinct replication phenotypes due to polymorphisms in the HBx gene. PLoS Pathog 2025; 21:e1012803. [PMID: 39787208 PMCID: PMC11717313 DOI: 10.1371/journal.ppat.1012803] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/02/2024] [Accepted: 12/03/2024] [Indexed: 01/12/2025] Open
Abstract
HBV genotype A has two major subtypes, A1 (commonly in Africa) and A2 (commonly in Europe) with only 4% nucleotide differences. Individuals infected with these two subtypes appear to have different clinical manifestations and virologic features. Whether such a difference results from the virus or host has not been established. Using HBV generated from molecule clones of subtypes A1 and A2 in cell culture (HBVcc), we demonstrate that HBVcc of subtypes A1 and A2 can be passaged in vitro and in vivo and respond equally well to human IFN-α treatment. HBVcc passaged in human liver chimeric mice (HBVmp) infected human hepatocytes more efficiently than that of the original HBVcc. Subtype A2 showed a much higher viral replication level than that of subtype A1. Mechanistic investigations using constructs with chimeric A1/A2 sequences and specific mutations indicated that subtype A2 has an inherently higher replication phenotype due to specific polymorphisms in the HBx gene resulting in amino acid variations. Studies of HBx expression demonstrated that A1 HBx is expressed at a much lower level than that of A2 HBx. Mutagenesis studies identified two HBx amino acid variations responsible for the observed phenotypic difference. Using AlphaFold2, we generated structural models of HBx proteins of A1 and A2. Superposition of the two models reveal that the overall structural motifs are similarly aligned, except for the C-terminal peptides diverging between the A1 and A2 models, possibly explaining their functional difference. In conclusion, using various in vitro and in vivo models, here we show that subtype A2 has an inherently higher replication phenotype due to polymorphisms in HBx that result in possible differences in structure and expression level of the two subtype HBx proteins. This genotypic difference potentially explains the reported clinical differences between the two subtypes as well as providing a previously unrecognized association between viral sequence variations and clinical manifestations of HBV infection in humans.
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Affiliation(s)
- Min Zhang
- Liver Diseases Branch, NIDDK, NIH, Bethesda, Maryland, United States of America
| | - Karim Mouzannar
- Liver Diseases Branch, NIDDK, NIH, Bethesda, Maryland, United States of America
| | - Zhensheng Zhang
- Liver Diseases Branch, NIDDK, NIH, Bethesda, Maryland, United States of America
| | - Yuji Teraoka
- Department of Gastroenterology and Metabolism, Graduate School of Biomedical & Health Science, Hiroshima University, Hiroshima, Japan
| | - Jason Piotrowski
- Liver Diseases Branch, NIDDK, NIH, Bethesda, Maryland, United States of America
| | - Yuji Ishida
- Division of Gastrointestinal and Liver Diseases, Department of Medicine, University of Southern California, Keck School of Medicine, Los Angeles, California, United States of America
- PhoenixBio Co., Ltd., Higashi-Hiroshima, Hiroshima, Japan
| | - Chise Tateno-Mukaidani
- Division of Gastrointestinal and Liver Diseases, Department of Medicine, University of Southern California, Keck School of Medicine, Los Angeles, California, United States of America
- PhoenixBio Co., Ltd., Higashi-Hiroshima, Hiroshima, Japan
| | - Takeshi Saito
- Division of Gastrointestinal and Liver Diseases, Department of Medicine, University of Southern California, Keck School of Medicine, Los Angeles, California, United States of America
| | - Hiromi Abe-Chayama
- Center for Medical Specialist Graduate Education and Research, Hiroshima, Japan
| | - Kazuaki Chayama
- Collaborative Research Laboratory of Medical Innovation, Hiroshima University, Hiroshima, Japan
| | - T. Jake Liang
- Liver Diseases Branch, NIDDK, NIH, Bethesda, Maryland, United States of America
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Bächer J, Allweiss L, Dandri M. SMC5/6-Mediated Transcriptional Regulation of Hepatitis B Virus and Its Therapeutic Potential. Viruses 2024; 16:1667. [PMID: 39599784 PMCID: PMC11598903 DOI: 10.3390/v16111667] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/09/2024] [Revised: 10/16/2024] [Accepted: 10/23/2024] [Indexed: 11/29/2024] Open
Abstract
Cells have developed various mechanisms to counteract viral infections. In an evolutionary arms race, cells mobilize cellular restriction factors to fight off viruses, targeted by viral factors to facilitate their own replication. The hepatitis B virus (HBV) is a small dsDNA virus that causes acute and chronic infections of the liver. Its genome persists in the nuclei of infected hepatocytes as a covalently closed circular DNA (cccDNA) minichromosome, thus building up an episomal persistence reservoir. The chromosomal maintenance complex SMC5/6 acts as a restriction factor hindering cccDNA transcription, whereas the viral regulatory protein HBx targets SMC5/6 for proteasomal degradation, thus relieving transcriptional suppression of the HBV minichromosome. To date, no curative therapies are available for chronic HBV carriers. Knowledge of the factors regulating the cccDNA and the development of therapies involving silencing the minichromosome or specifically interfering with the HBx-SMC5/6 axis holds promise in achieving sustained viral control. Here, we summarize the current knowledge of the mechanism of SMC5/6-mediated HBV restriction. We also give an overview of SMC5/6 cellular functions and how this compares to the restriction of other DNA viruses. We further discuss the therapeutic potential of available and investigational drugs interfering with the HBx-SMC5/6 axis.
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Affiliation(s)
- Johannes Bächer
- I. Department of Internal Medicine, Center for Internal Medicine, University Medical Center Hamburg-Eppendorf, Martinistr. 52, 20246 Hamburg, Germany; (J.B.); (L.A.)
| | - Lena Allweiss
- I. Department of Internal Medicine, Center for Internal Medicine, University Medical Center Hamburg-Eppendorf, Martinistr. 52, 20246 Hamburg, Germany; (J.B.); (L.A.)
- German Center for Infection Research (DZIF), Hamburg-Lübeck-Borstel-Riems Site, Germany
| | - Maura Dandri
- I. Department of Internal Medicine, Center for Internal Medicine, University Medical Center Hamburg-Eppendorf, Martinistr. 52, 20246 Hamburg, Germany; (J.B.); (L.A.)
- German Center for Infection Research (DZIF), Hamburg-Lübeck-Borstel-Riems Site, Germany
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4
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Wu Z, Cardona EA, Pierce JT. Non-apoptotic role of EGL-1 in exopher production and neuronal health in Caenorhabditis elegans. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2024:2024.04.19.590348. [PMID: 38712027 PMCID: PMC11071422 DOI: 10.1101/2024.04.19.590348] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 05/08/2024]
Abstract
While traditionally studied for their pro-apoptotic functions, recent research suggests BH3-only proteins also have non-apoptotic roles. Here, we find that EGL-1, the BH3-only protein in Caenorhabditis elegans, promotes the cell-autonomous production of exophers in adult neurons. Exophers are large, micron-scale vesicles that are ejected from the cell and contain cellular components such as mitochondria. EGL-1 facilitates exopher production potentially through regulation of mitochondrial dynamics. Moreover, an endogenous, low level of EGL-1 expression appears to benefit dendritic health. Our findings provide insights into the mechanistic role of BH3-only protein in mitochondrial dynamics, downstream exopher production, and ultimately neuronal health.
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Affiliation(s)
- Zheng Wu
- Department of Neuroscience, Center for Learning and Memory, Waggoner Center for Alcohol & Addiction Research, University of Texas at Austin, Austin, TX
| | - Eric A. Cardona
- Department of Neuroscience, Center for Learning and Memory, Waggoner Center for Alcohol & Addiction Research, University of Texas at Austin, Austin, TX
| | - Jonathan T. Pierce
- Department of Neuroscience, Center for Learning and Memory, Waggoner Center for Alcohol & Addiction Research, University of Texas at Austin, Austin, TX
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Kusunoki H, Sakamoto T, Kobayashi N, Kohno T, Wakamatsu K, Nagata T. Structural Insights into the Interaction between the C-Terminal-Deleted BH3-like Motif Peptide of Hepatitis B Virus X Protein and Bcl-x L. Biochemistry 2024; 63:632-643. [PMID: 38377677 DOI: 10.1021/acs.biochem.3c00709] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/22/2024]
Abstract
Hepatitis B virus X protein (HBx) plays a crucial role in the development of hepatocellular carcinoma (HCC) associated with hepatitis B virus (HBV) infection. The full-length HBx protein interacts with Bcl-xL and is involved in the HBV replication and cell death processes. The three hydrophobic residues Trp120, Leu123, and Ile127 of the HBx BH3-like motif are essential for the Bcl-xL-binding. On the other hand, various lengths of C-terminal-truncated HBx mutants are frequently detected in HCC tissues, and these mutants, rather than the full-length HBx, appear to be responsible for HCC development. Notably, the region spanning residues 1-120 of HBx [HBx(1 and 120)] has been strongly associated with an increased risk of HCC development. However, the mode of interaction between HBx(1-120) and Bcl-xL remains unclear. HBx(1-120) possesses only Trp120 among the three hydrophobic residues essential for the Bcl-xL-binding. To elucidate this interaction mode, we employed a C-terminal-deleted HBx BH3-like motif peptide composed of residues 101-120. Here, we present the NMR complex structure of Bcl-xL and HBx(101-120). Our results demonstrate that HBx(101-120) binds to Bcl-xL in a weaker manner. Considering the high expression of Bcl-xL in HCC cells, this weak interaction, in conjunction with the overexpression of Bcl-xL in HCC cells, may potentially contribute to HCC development through the interaction between C-terminal-truncated HBx and Bcl-xL.
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Affiliation(s)
- Hideki Kusunoki
- Research Center for Biological Products in the Next Generation, National Institute of Infectious Diseases, Musashimurayama, Tokyo 208-0011, Japan
| | - Taiichi Sakamoto
- Department of Life Science, Faculty of Advanced Engineering, Chiba Institute of Technology, Narashino, Chiba 275-0016, Japan
| | - Naohiro Kobayashi
- Laboratory for Advanced NMR Application and Development, RIKEN Center for Biosystems Dynamics Research, Yokohama, Kanagawa 230-0045, Japan
| | - Toshiyuki Kohno
- Department of Medical Informatics, Research and Development Center for Medical Education, Kitasato University School of Medicine, Sagamihara, Kanagawa 252-0374, Japan
| | - Kaori Wakamatsu
- Department of Chemistry and Chemical Biology, Graduate School of Engineering, Gunma University, Kiryu, Gunma 376-8515, Japan
| | - Takashi Nagata
- Institute of Advanced Energy, Kyoto University, Kyoto, Uji 611-0011, Japan
- Graduate School of Energy Science, Kyoto University, Kyoto, Uji 611-0011, Japan
- Integrated Research Center for Carbon Negative Science, Kyoto University, Kyoto, Uji 611-0011, Japan
- Biomass Product Tree Industry-Academia Collaborative Research Laboratory, Kyoto University, Kyoto, Uji 611-0011, Japan
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Meng X, Dai X, Huang J, Han T, Liao X, Cheng K, Sun X, Xie Q, Sun P, Zhou X. The influence of male HBV infection on sperm quality, embryonic development, and assisted reproductive outcomes. Hum Reprod 2024; 39:43-52. [PMID: 37994690 DOI: 10.1093/humrep/dead235] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/27/2022] [Revised: 10/12/2023] [Indexed: 11/24/2023] Open
Abstract
STUDY QUESTION What is the impact of male hepatitis B virus (HBV) infection on sperm quality, embryonic development, and assisted reproductive outcomes? SUMMARY ANSWER Male HBV infection did not affect assisted reproductive outcomes, but HBV is capable of impairing human sperm and embryo formation in the early stages following fertilization. WHAT IS KNOWN ALREADY HBV is found in germ cells and early embryos of patients with HBV. HBV may impair human sperm function via increasing reactive oxygen species. STUDY DESIGN, SIZE, DURATION We conducted a retrospective cohort study of 1581 infertile couples, including 496 male patients clinically confirmed to have hepatitis B infection, and a laboratory study of effects of HBV proteins on early embryos, using human embryonic stem cells (hESCs), human sperm, and golden hamster oocytes. PARTICIPANTS/MATERIALS, SETTING, METHODS In total, 1581 infertile couples (24-40 years of age) who were admitted to a reproductive medicine center to undergo ART for the first time from January 2019 to November 2021 were selected as the study subjects. The case group was composed of 469 couples with hepatitis B surface antigen (HBsAg)-seropositive men and seronegative women (368 for IVF and 101 for ICSI treatment). The negative control group was composed of 1112 couples where both men and women were seronegative for hepatitis B antigen. We divided these couples into three comparison groups (IVF/ICSI, IVF, and ICSI). IVF of human sperm and hamster oocytes was used to evaluate the influence of the HBV HBs protein on formation of 2-cell embryos. Mitochondrial membrane potential (MMP) of hESCs was assayed via a fluorescence intensity system. Immunofluorescence staining of the phosphorylated histone H2A.X was applied to identify DNA damage to hESCs caused by the HBV X (HBx) protein. MAIN RESULTS AND THE ROLE OF CHANCE Sperm concentration, total sperm number, and sperm with normal morphology were decreased in the couples with HBV-infected males in couples who were undergoing IVF/ICSI (male HBV(+) vs control: 469 vs 1112 individuals; sperm number, P < 0.01; normal sperm morphology, P < 0.01), IVF (368 vs 792; sperm number, P < 0.01; normal sperm morphology, P ≤ 0.05), and ICSI (101 vs 306; sperm number, P < 0.01; normal sperm morphology, P < 0.001). There was no significant difference in the number of embryo cleavages, blastocyst formation, biochemical pregnancy rate, clinical pregnancy rate, and live-birth rate between case and control groups. The 2PN fertilization rate in IVF/ICSI (P < 0.01) and ICSI (P < 0.05) couples, and the number of 2PN-fertilized oocytes in IVF (P < 0.001) couples were lower in couples with male HBV infection compared to control couples. HBV HBs protein reduced the MMP of human sperm and decreased 2-cell embryo formation in IVF of human sperm and zona-free-hamster oocyte. A reduction in fluorescence intensity and immunofluorescence staining of phosphorylated histone H2A.X indicated that HBx caused MMP impairment and DNA damage in human early embryonic cells, respectively. LARGE SCALE DATA N/A. LIMITATIONS, REASONS FOR CAUTION HBV can be examined in samples of sperm or discarded IVF early embryos from HBsAg-seropositive men and seronegative women. The hESC model in vitro may not fully mimic the natural embryos in vivo. WIDER IMPLICATIONS OF THE FINDINGS This study furthers our understanding of the influence of male HBV infection on embryonic development. Our results suggest that a semen-washing process may be necessary for male patients with HBV undergoing ART to minimize the potential negative effects of HBV infection on the early embryo. STUDY FUNDING/COMPETING INTEREST(S) This work was funded by grants from the National Natural Science Foundation of China, grant numbers 81870432 and 81570567 to X.Z., 81571994 to P.S., and 81950410640, the Natural Science Foundation of Guangdong Province, China (No. 2023A1515010660 to X.Z.), and the Li Ka Shing Shantou University Foundation (Grant No. L11112008). The authors have no conflicts of interest.
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Affiliation(s)
- Xiangqian Meng
- Chengdu Xinan Gynecology Hospital; Sichuan Jinxin Xinan Women's & Children's Hospital (Bisheng), Chengdu, China
| | - Ximing Dai
- Stem Cell Research Center, Shantou University Medical College, Shantou, China
- Research Center for Reproductive Medicine, Shantou University Medical College, Shantou, China
- Guangdong Provincial Key Laboratory of Infectious Diseases and Molecular Immunopathology, Shantou University Medical College, Shantou, China
| | - Jihua Huang
- Chengdu Xinan Gynecology Hospital; Sichuan Jinxin Xinan Women's & Children's Hospital (Bisheng), Chengdu, China
| | - Tingting Han
- Chengdu Xinan Gynecology Hospital; Sichuan Jinxin Xinan Women's & Children's Hospital (Bisheng), Chengdu, China
| | - Xue Liao
- Chengdu Xinan Gynecology Hospital; Sichuan Jinxin Xinan Women's & Children's Hospital (Bisheng), Chengdu, China
| | - Ke Cheng
- Stem Cell Research Center, Shantou University Medical College, Shantou, China
- Research Center for Reproductive Medicine, Shantou University Medical College, Shantou, China
- Guangdong Provincial Key Laboratory of Infectious Diseases and Molecular Immunopathology, Shantou University Medical College, Shantou, China
| | - Xiaoyue Sun
- Stem Cell Research Center, Shantou University Medical College, Shantou, China
- Research Center for Reproductive Medicine, Shantou University Medical College, Shantou, China
- Guangdong Provincial Key Laboratory of Infectious Diseases and Molecular Immunopathology, Shantou University Medical College, Shantou, China
| | - Qingdong Xie
- Stem Cell Research Center, Shantou University Medical College, Shantou, China
- Research Center for Reproductive Medicine, Shantou University Medical College, Shantou, China
- Guangdong Provincial Key Laboratory of Infectious Diseases and Molecular Immunopathology, Shantou University Medical College, Shantou, China
| | - Pingnan Sun
- Stem Cell Research Center, Shantou University Medical College, Shantou, China
- Research Center for Reproductive Medicine, Shantou University Medical College, Shantou, China
- Guangdong Provincial Key Laboratory of Infectious Diseases and Molecular Immunopathology, Shantou University Medical College, Shantou, China
| | - Xiaoling Zhou
- Stem Cell Research Center, Shantou University Medical College, Shantou, China
- Research Center for Reproductive Medicine, Shantou University Medical College, Shantou, China
- Guangdong Provincial Key Laboratory of Infectious Diseases and Molecular Immunopathology, Shantou University Medical College, Shantou, China
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7
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Ye P, Fang Q, Hu X, Zou W, Huang M, Ke M, Li Y, Liu M, Cai X, Zhang C, Hua N, Al-Sheikh U, Liu X, Yu P, Jiang P, Pan PY, Luo J, Jiang LH, Xu S, Fang EF, Su H, Kang L, Yang W. TRPM2 as a conserved gatekeeper determines the vulnerability of DA neurons by mediating ROS sensing and calcium dyshomeostasis. Prog Neurobiol 2023; 231:102530. [PMID: 37739206 DOI: 10.1016/j.pneurobio.2023.102530] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/11/2023] [Revised: 09/01/2023] [Accepted: 09/17/2023] [Indexed: 09/24/2023]
Abstract
Different dopaminergic (DA) neuronal subgroups exhibit distinct vulnerability to stress, while the underlying mechanisms are elusive. Here we report that the transient receptor potential melastatin 2 (TRPM2) channel is preferentially expressed in vulnerable DA neuronal subgroups, which correlates positively with aging in Parkinson's Disease (PD) patients. Overexpression of human TRPM2 in the DA neurons of C. elegans resulted in selective death of ADE but not CEP neurons in aged worms. Mechanistically, TRPM2 activation mediates FZO-1/CED-9-dependent mitochondrial hyperfusion and mitochondrial permeability transition (MPT), leading to ADE death. In mice, TRPM2 knockout reduced vulnerable substantia nigra pars compacta (SNc) DA neuronal death induced by stress. Moreover, the TRPM2-mediated vulnerable DA neuronal death pathway is conserved from C. elegans to toxin-treated mice model and PD patient iPSC-derived DA neurons. The vulnerable SNc DA neuronal loss is the major symptom and cause of PD, and therefore the TRPM2-mediated pathway serves as a promising therapeutic target against PD.
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Affiliation(s)
- Peiwu Ye
- Department of Biophysics, Institute of Neuroscience, NHC and CAMS Key Laboratory of Medical Neurobiology, Zhejiang University School of Medicine, Hangzhou 310058, China
| | - Qiuyuan Fang
- Department of Biophysics, Institute of Neuroscience, NHC and CAMS Key Laboratory of Medical Neurobiology, Zhejiang University School of Medicine, Hangzhou 310058, China
| | - Xupang Hu
- Second Clinical Medical College, Affiliated Secondary Hospital, Zhejiang Chinese Medical University, Hangzhou, Zhejiang 310011, China
| | - Wenjuan Zou
- Department of Neurobiology and Department of Neurosurgery of the First Affiliated Hospital, Zhejiang University School of Medicine, Zhejiang 310053, China
| | - Miaodan Huang
- Institute of Chinese Medical Sciences, University of Macau, Macau, China
| | - Minjing Ke
- Institute of Chinese Medical Sciences, University of Macau, Macau, China
| | - Yunhao Li
- Institute of Chinese Medical Sciences, University of Macau, Macau, China
| | - Min Liu
- Department of Biophysics, Institute of Neuroscience, NHC and CAMS Key Laboratory of Medical Neurobiology, Zhejiang University School of Medicine, Hangzhou 310058, China
| | - Xiaobo Cai
- Department of Biophysics, Institute of Neuroscience, NHC and CAMS Key Laboratory of Medical Neurobiology, Zhejiang University School of Medicine, Hangzhou 310058, China
| | - Congyi Zhang
- Department of Biophysics, Institute of Neuroscience, NHC and CAMS Key Laboratory of Medical Neurobiology, Zhejiang University School of Medicine, Hangzhou 310058, China
| | - Ning Hua
- Department of Biophysics, Institute of Neuroscience, NHC and CAMS Key Laboratory of Medical Neurobiology, Zhejiang University School of Medicine, Hangzhou 310058, China
| | - Umar Al-Sheikh
- Department of Neurobiology and Department of Neurosurgery of the First Affiliated Hospital, Zhejiang University School of Medicine, Zhejiang 310053, China
| | - Xingyu Liu
- Department of Biophysics, Institute of Neuroscience, NHC and CAMS Key Laboratory of Medical Neurobiology, Zhejiang University School of Medicine, Hangzhou 310058, China
| | - Peilin Yu
- Department of Toxicology, School of Public Health, Zhejiang University, Hangzhou, Zhejiang 310058, China
| | - Peiran Jiang
- School of Brain Science and Brain Medicine, Zhejiang University, Hangzhou 310058, China
| | - Ping-Yue Pan
- Department of Neuroscience and Cell Biology, Rutgers University Robert Wood Johnson Medical School, 675 Hoes Lane, Piscataway, NJ 08854, USA
| | - Jianhong Luo
- School of Brain Science and Brain Medicine, Zhejiang University, Hangzhou 310058, China
| | - Lin-Hua Jiang
- School of Biomedical Sciences, University of Leeds, Leeds LS2 9JT, UK; Sino-UK Laboratory of Brain Function and Injury of Henan Province, Department of Physiology and Neurobiology, Xinxiang Medical University, Xinxiang 453000, China; University of Leeds, Leeds LS2 9JT, UK
| | - Suhong Xu
- Center for Stem Cell and Regenerative Medicine and Department of Cardiology of The Second Affiliated Hospital, Zhejiang University School of Medicine, 310058, Hangzhou, China
| | - Evandro F Fang
- Department of Clinical Molecular Biology, University of Oslo and Akershus University Hospital, Lørenskog, Norway
| | - Huanxing Su
- Institute of Chinese Medical Sciences, University of Macau, Macau, China.
| | - Lijun Kang
- Second Clinical Medical College, Affiliated Secondary Hospital, Zhejiang Chinese Medical University, Hangzhou, Zhejiang 310011, China; School of Brain Science and Brain Medicine, Zhejiang University, Hangzhou 310058, China.
| | - Wei Yang
- Department of Biophysics, Institute of Neuroscience, NHC and CAMS Key Laboratory of Medical Neurobiology, Zhejiang University School of Medicine, Hangzhou 310058, China.
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8
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Saeed U, Piracha ZZ, Alrokayan S, Hussain T, Almajhdi FN, Waheed Y. Immunoinformatics and Evaluation of Peptide Vaccines Derived from Global Hepatitis B Viral HBx and HBc Proteins Critical for Covalently Closed Circular DNA Integrity. Microorganisms 2023; 11:2826. [PMID: 38137971 PMCID: PMC10745757 DOI: 10.3390/microorganisms11122826] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/18/2023] [Revised: 11/07/2023] [Accepted: 11/17/2023] [Indexed: 12/24/2023] Open
Abstract
The Hepatitis B virus (HBV) HBx and HBc proteins play a crucial role in associating with covalently closed circular DNA (cccDNA), the primary factor contributing to intrahepatic viral persistence and a major obstacle in achieving a cure for HBV. The cccDNA serves as a reservoir for viral persistence. Targeting the viral HBc and HBx proteins' interaction with cccDNA could potentially limit HBV replication. In this study, we present epitopes identified from global consensus sequences of HBx and HBc proteins that have the potential to serve as targets for the development of effective vaccine candidates. Furthermore, conserved residues identified through this analysis can be utilized in designing novel, site-specific anti-HBV agents capable of targeting all major genotypes of HBV. Our approach involved designing global consensus sequences for HBx and HBc proteins, enabling the analysis of variable regions and highly conserved motifs. These identified motifs and regions offer potent sites for the development of peptide vaccines, the design of site-specific RNA interference, and the creation of anti-HBV inhibitors. The epitopes derived from global consensus sequences of HBx and HBc proteins emerge as promising targets for the development of effective vaccine candidates. Additionally, the conserved residues identified provide valuable insights for the development of innovative, site-specific anti-HBV agents capable of targeting all major genotypes of HBV from A to J.
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Affiliation(s)
- Umar Saeed
- Clinical and Biomedical Research Center (CBRC) and Multidisciplinary Laboratory (MDL), Foundation University Islamabad, Islamabad 44000, Pakistan;
- Department of Microbiology, Ajou University School of Medicine, Suwon 443-749, Republic of Korea;
| | - Zahra Zahid Piracha
- Department of Microbiology, Ajou University School of Medicine, Suwon 443-749, Republic of Korea;
- International Center of Medical Sciences Research (ICMSR), Islamabad 44000, Pakistan
| | - Salman Alrokayan
- Research Chair for Biomedical Application of Nanomaterials, Biochemistry Department, College of Sciences, King Saud University, Riyadh 11362, Saudi Arabia;
| | - Tajamul Hussain
- Research Chair for Biomedical Application of Nanomaterials, Biochemistry Department, College of Sciences, King Saud University, Riyadh 11362, Saudi Arabia;
- Center of Excellence in Biotechnology Research, College of Applied Medical Sciences, King Saud University, P.O. Box 10219, Riyadh 11451, Saudi Arabia
| | - Fahad N. Almajhdi
- Botany and Microbiology Department, College of Science, King Saud University, P.O. Box 2455, Riyadh 11451, Saudi Arabia;
| | - Yasir Waheed
- Office of Research, Innovation, and Commercialization, Shaheed Zulfiqar Ali Bhutto Medical University, Islamabad 44000, Pakistan;
- Gilbert and Rose-Marie Chagoury School of Medicine, Lebanese American University, Byblos 1401, Lebanon
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9
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Shoraka S, Hosseinian SM, Hasibi A, Ghaemi A, Mohebbi SR. The role of hepatitis B virus genome variations in HBV-related HCC: effects on host signaling pathways. Front Microbiol 2023; 14:1213145. [PMID: 37588887 PMCID: PMC10426804 DOI: 10.3389/fmicb.2023.1213145] [Citation(s) in RCA: 2] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/27/2023] [Accepted: 07/12/2023] [Indexed: 08/18/2023] Open
Abstract
Hepatocellular carcinoma (HCC) is a significant global health issue, with a high prevalence in many regions. There are variations in the etiology of HCC in different regions, but most cases are due to long-term infection with viral hepatitis. Hepatitis B virus (HBV) is responsible for more than 50% of virus-related HCC, which highlights the importance of HBV in pathogenesis of the disease. The development and progression of HBV-related HCC is a complex multistep process that can involve host, viral, and environmental factors. Several studies have suggested that some HBV genome mutations as well as HBV proteins can dysregulate cell signaling pathways involved in the development of HCC. Furthermore, it seems that the pathogenicity, progression of liver diseases, response to treatment and also viral replication are different among HBV mutants. Understanding the relationship between HBV genome variations and host signaling pathway alteration will improve our understanding of the molecular pathogenesis of HBV-related HCC. Furthermore, investigating commonly dysregulated pathways in HBV-related HCC is necessary to discover more specific therapeutic targets and develop more effective strategies for HCC treatment. The objective of this review is to address the role of HBV in the HCC progression and primarily focus on the impacts of HBV genome variations on HCC-related signaling pathways.
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Affiliation(s)
- Shahrzad Shoraka
- Basic and Molecular Epidemiology of Gastrointestinal Disorders Research Center, Research Institute for Gastroenterology and Liver Diseases, Shahid Beheshti University of Medical Sciences, Tehran, Iran
- Department of Microbiology and Microbial Biotechnology, Faculty of Life Sciences and Biotechnology, Shahid Beheshti University, Tehran, Iran
| | - Seyed Mahdi Hosseinian
- Gastroenterology and Liver Diseases Research Center, Research Institute for Gastroenterology and Liver Diseases, Shahid Beheshti University of Medical Sciences, Tehran, Iran
| | - Ayda Hasibi
- Foodborne and Waterborne Diseases Research Center, Research Institute for Gastroenterology and Liver Diseases, Shahid Beheshti University of Medical Sciences, Tehran, Iran
| | - Amir Ghaemi
- Department of Virology, Pasteur Institute of Iran, Tehran, Iran
| | - Seyed Reza Mohebbi
- Gastroenterology and Liver Diseases Research Center, Research Institute for Gastroenterology and Liver Diseases, Shahid Beheshti University of Medical Sciences, Tehran, Iran
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10
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Guo X, Feng Y, Zhao X, Qiao S, Ma Z, Li Z, Zheng H, Xiao S. Coronavirus Porcine Epidemic Diarrhea Virus Utilizes Chemokine Interleukin-8 to Facilitate Viral Replication by Regulating Ca 2+ Flux. J Virol 2023; 97:e0029223. [PMID: 37133374 PMCID: PMC10231212 DOI: 10.1128/jvi.00292-23] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/23/2023] [Accepted: 04/10/2023] [Indexed: 05/04/2023] Open
Abstract
Chemokine production by epithelial cells is crucial for neutrophil recruitment to sites of inflammation during viral infection. However, the effect of chemokine on epithelia and how chemokine is involved in coronavirus infection remains to be fully understood. Here, we identified an inducible chemokine interleukin-8 (CXCL8/IL-8), which could promote coronavirus porcine epidemic diarrhea virus (PEDV) infection in African green monkey kidney epithelial cells (Vero) and Lilly Laboratories cell-porcine kidney 1 epithelial cells (LLC-PK1). IL-8 deletion restrained cytosolic calcium (Ca2+), whereas IL-8 stimulation improved cytosolic Ca2+. The consumption of Ca2+ restricted PEDV infection. PEDV internalization and budding were obvious reductions when cytosolic Ca2+ was abolished in the presence of Ca2+ chelators. Further study revealed that the upregulated cytosolic Ca2+ redistributes intracellular Ca2+. Finally, we identified that G protein-coupled receptor (GPCR)-phospholipase C (PLC)-inositol trisphosphate receptor (IP3R)-store-operated Ca2+ (SOC) signaling was crucial for enhancive cytosolic Ca2+ and PEDV infection. To our knowledge, this study is the first to uncover the function of chemokine IL-8 during coronavirus PEDV infection in epithelia. PEDV induces IL-8 expression to elevate cytosolic Ca2+, promoting its infection. Our findings reveal a novel role of IL-8 in PEDV infection and suggest that targeting IL-8 could be a new approach to controlling PEDV infection. IMPORTANCE Coronavirus porcine epidemic diarrhea virus (PEDV) is a highly contagious enteric coronavirus that caused severe economic losses worldwide, and more effort is needed to develop economical and efficient vaccines to control or eliminate this disease. The chemokine interleukin-8 (CXCL8/IL-8) is indispensable for the activation and trafficking of inflammatory mediators and tumor progression and metastasis. This study evaluated the effect of IL-8 on PEDV infection in epithelia. We found that IL-8 expression improved cytosolic Ca2+ in epithelia, facilitating PEDV rapid internalization and egress. G protein-coupled receptor (GPCR)-phospholipase C (PLC)-inositol trisphosphate receptor (IP3R)-SOC signaling was activated by IL-8, releasing the intracellular Ca2+ stores from endoplasmic reticulum (ER). These findings provide a better understanding of the role of IL-8 in PEDV-induced immune responses, which will help develop small-molecule drugs for coronavirus cure.
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Affiliation(s)
- Xuyang Guo
- College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, China
| | - Yingtong Feng
- College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, China
| | - Xiaojing Zhao
- College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, China
| | - Shuang Qiao
- College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, China
| | - Zhiqian Ma
- State Key Laboratory for Animal Disease Control and Prevention, College of Veterinary Medicine, Lanzhou University, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou, Gansu, China
| | - Zhiwei Li
- College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, China
| | - Haixue Zheng
- State Key Laboratory for Animal Disease Control and Prevention, College of Veterinary Medicine, Lanzhou University, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou, Gansu, China
| | - Shuqi Xiao
- College of Veterinary Medicine, Northwest A&F University, Yangling, Shaanxi, China
- State Key Laboratory for Animal Disease Control and Prevention, College of Veterinary Medicine, Lanzhou University, Lanzhou Veterinary Research Institute, Chinese Academy of Agricultural Sciences, Lanzhou, Gansu, China
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11
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Wang Y, Wu W, Gong J. Live or death in cells: from micronutrition metabolism to cell fate. Front Cell Dev Biol 2023; 11:1185989. [PMID: 37250891 PMCID: PMC10213646 DOI: 10.3389/fcell.2023.1185989] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/14/2023] [Accepted: 04/28/2023] [Indexed: 05/31/2023] Open
Abstract
Micronutrients and cell death have a strong relationship and both are essential for human to maintain good body health. Dysregulation of any micronutrients causes metabolic or chronic diseases, including obesity, cardiometabolic condition, neurodegeneration, and cancer. The nematode Caenorhabditis elegans is an ideal genetic organism for researching the mechanisms of micronutrients in metabolism, healthspan, and lifespan. For example, C. elegans is a haem auxotroph, and the research of this special haem trafficking pathway contributes important reference to mammal study. Also, C. elegans characteristics including anatomy simply, clear cell lineage, well-defined genetics, and easily differentiated cell forms make it a powerful tool for studying the mechanisms of cell death including apoptosis, necrosis, autophagy, and ferroptosis. Here, we describe the understanding of micronutrient metabolism currently and also sort out the fundamental mechanisms of different kinds of cell death. A thorough understanding of these physiological processes not only builds a foundation for developing better treatments for various micronutrient disorders but also provides key insights into human health and aging.
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Affiliation(s)
- Yuting Wang
- Key Laboratory of Molecular Biophysics of MOE, College of Life Science and Technology, Huazhong University of Science and Technology, Wuhan, Hubei, China
| | - Wei Wu
- State Key Laboratory for Diagnosis and Treatment of Infectious Diseases, National Clinical Research Center for Infectious Diseases, The First Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou, China
| | - Jianke Gong
- Key Laboratory of Molecular Biophysics of MOE, College of Life Science and Technology, Huazhong University of Science and Technology, Wuhan, Hubei, China
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12
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Kar A, Samanta A, Mukherjee S, Barik S, Biswas A. The HBV web: An insight into molecular interactomes between the hepatitis B virus and its host en route to hepatocellular carcinoma. J Med Virol 2023; 95:e28436. [PMID: 36573429 DOI: 10.1002/jmv.28436] [Citation(s) in RCA: 18] [Impact Index Per Article: 9.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/26/2022] [Revised: 11/26/2022] [Accepted: 12/23/2022] [Indexed: 12/28/2022]
Abstract
Hepatitis B virus (HBV) is a major aetiology associated with the development and progression of hepatocellular carcinoma (HCC), the most common primary liver malignancy. Over the past few decades, direct and indirect mechanisms have been identified in the pathogenesis of HBV-associated HCC which include altered signaling pathways, genome integration, mutation-induced genomic instability, chromosomal deletions and rearrangements. Intertwining of the HBV counterparts with the host cellular factors, though well established, needs to be systemized to understand the dynamics of host-HBV crosstalk and its consequences on HCC progression. Existence of a vast array of protein-protein and protein-nucleic acid interaction databases has led to the uncoiling of the compendia of genes/gene products associated with these interactions. This review covers the existing knowledge about the HBV-host interplay and brings it down under one canopy emphasizing on the HBV-host interactomics; and thereby highlights new strategies for therapeutic advancements against HBV-induced HCC.
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Affiliation(s)
- Arpita Kar
- Department of Signal Transduction and Biogenic Amines, Chittaranjan National Cancer Institute, Kolkata, West Bengal, India
| | - Abhisekh Samanta
- Department of Signal Transduction and Biogenic Amines, Chittaranjan National Cancer Institute, Kolkata, West Bengal, India
| | - Soumyadeep Mukherjee
- Department of In Vitro Carcinogenesis and Cellular Chemotherapy, Chittaranjan National Cancer Institute, Kolkata, West Bengal, India
| | - Subhasis Barik
- Department of In Vitro Carcinogenesis and Cellular Chemotherapy, Chittaranjan National Cancer Institute, Kolkata, West Bengal, India
| | - Avik Biswas
- Department of Signal Transduction and Biogenic Amines, Chittaranjan National Cancer Institute, Kolkata, West Bengal, India
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13
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Fu Y, Fang F, Guo H, Xiao X, Hu Y, Zeng Y, Chen T, Wu S, Lin N, Huang J, Jiang L, Ou Q, Liu C. Compartmentalisation of Hepatitis B virus X gene evolution in hepatocellular carcinoma microenvironment and the genotype-phenotype correlation of tumorigenicity in HBV-related patients with hepatocellular carcinoma. Emerg Microbes Infect 2022; 11:2486-2501. [PMID: 36102940 PMCID: PMC9621239 DOI: 10.1080/22221751.2022.2125344] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/25/2022] [Revised: 08/26/2022] [Accepted: 09/12/2022] [Indexed: 02/08/2023]
Abstract
Hepatitis B virus (HBV) exists as quasispecies (QS). However, the evolutionary characteristics of haplotypes of HBV X gene in the hepatocellular carcinoma (HCC) microenvironment remain unclear. Mutations across X gene are essential for the tumorigenicity of HBV X protein (HBx). However, the functional phenotypes of many mutant HBx remain unknown. This study aims to compare the characteristics of X gene evolution between tumour and non-tumour tissues in HCC patients and investigate the tumorigenic phenotype of HBx harbouring mutation T81P/S101P/L123S. This study included 24 HCC patients. Molecular cloning of X gene was performed to analyse characteristics of haplotypes in liver tissues. HCC cell lines stably expressing wild-type or mutant HBx and subcutaneous tumour xenograft mouse model were used to assess HBx-T81P/S101P/L123S tumorigenicity. The mean heterogeneity of HBV QS across X gene in tumour tissues was lower than that in non-tumour tissues. A location bias was observed in X gene clones with genotype C or D in tumour tissues compared to those with genotype B. Mutations in genotype-C or - D clones were mainly clustered in the dimerization region and aa110-aa140 within the transactivation region. A novel mutation combination at residues 81, 101 and 123 was identified in tumour tissues. Further, HBx-T81P/S101P/L123S promotes cell proliferation and increases genomic instability, which was mediated by MYC. This study elucidates the compartmentalized evolution patterns of HBV X gene between intra tumour and non-tumour tissues in HCC patients and provides a new mechanism underlying HBV-driven hepatocarcinogenesis, suggesting a potential viral marker for monitoring HCC.
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Affiliation(s)
- Ya Fu
- Department of Laboratory Medicine, The First Affiliated Hospital, Fujian Medical University, Fuzhou, People’s Republic of China
- Clinical Laboratory Diagnostics, The First Clinical College, Fujian Medical University, Fuzhou, People’s Republic of China
- Fujian Key Laboratory of Laboratory Medicine, The First Affiliated Hospital, Fujian Medical University, Fuzhou, People’s Republic of China
- Gene Diagnosis Research Center, Fujian Medical University, Fuzhou, People’s Republic of China
| | - Fengling Fang
- Department of Laboratory Medicine, The First Affiliated Hospital, Fujian Medical University, Fuzhou, People’s Republic of China
- Clinical Laboratory Diagnostics, The First Clinical College, Fujian Medical University, Fuzhou, People’s Republic of China
- Fujian Key Laboratory of Laboratory Medicine, The First Affiliated Hospital, Fujian Medical University, Fuzhou, People’s Republic of China
- Gene Diagnosis Research Center, Fujian Medical University, Fuzhou, People’s Republic of China
| | - Hongyan Guo
- Department of Laboratory Medicine, The First Affiliated Hospital, Fujian Medical University, Fuzhou, People’s Republic of China
- Clinical Laboratory Diagnostics, The First Clinical College, Fujian Medical University, Fuzhou, People’s Republic of China
- Fujian Key Laboratory of Laboratory Medicine, The First Affiliated Hospital, Fujian Medical University, Fuzhou, People’s Republic of China
- Gene Diagnosis Research Center, Fujian Medical University, Fuzhou, People’s Republic of China
| | - Xialin Xiao
- Department of Laboratory Medicine, The First Affiliated Hospital, Fujian Medical University, Fuzhou, People’s Republic of China
- Clinical Laboratory Diagnostics, The First Clinical College, Fujian Medical University, Fuzhou, People’s Republic of China
- Fujian Key Laboratory of Laboratory Medicine, The First Affiliated Hospital, Fujian Medical University, Fuzhou, People’s Republic of China
- Gene Diagnosis Research Center, Fujian Medical University, Fuzhou, People’s Republic of China
| | - Yuhai Hu
- Department of Hepatobiliary Surgery, The First Affiliated Hospital, Fujian Medical University, Fuzhou, People’s Republic of China
| | - Yongbin Zeng
- Department of Laboratory Medicine, The First Affiliated Hospital, Fujian Medical University, Fuzhou, People’s Republic of China
- Clinical Laboratory Diagnostics, The First Clinical College, Fujian Medical University, Fuzhou, People’s Republic of China
- Fujian Key Laboratory of Laboratory Medicine, The First Affiliated Hospital, Fujian Medical University, Fuzhou, People’s Republic of China
- Gene Diagnosis Research Center, Fujian Medical University, Fuzhou, People’s Republic of China
| | - Tianbin Chen
- Department of Laboratory Medicine, The First Affiliated Hospital, Fujian Medical University, Fuzhou, People’s Republic of China
- Clinical Laboratory Diagnostics, The First Clinical College, Fujian Medical University, Fuzhou, People’s Republic of China
- Fujian Key Laboratory of Laboratory Medicine, The First Affiliated Hospital, Fujian Medical University, Fuzhou, People’s Republic of China
- Gene Diagnosis Research Center, Fujian Medical University, Fuzhou, People’s Republic of China
| | - Songhang Wu
- Department of Laboratory Medicine, The First Affiliated Hospital, Fujian Medical University, Fuzhou, People’s Republic of China
- Clinical Laboratory Diagnostics, The First Clinical College, Fujian Medical University, Fuzhou, People’s Republic of China
- Fujian Key Laboratory of Laboratory Medicine, The First Affiliated Hospital, Fujian Medical University, Fuzhou, People’s Republic of China
- Gene Diagnosis Research Center, Fujian Medical University, Fuzhou, People’s Republic of China
| | - Ni Lin
- Department of Laboratory Medicine, The First Affiliated Hospital, Fujian Medical University, Fuzhou, People’s Republic of China
- Clinical Laboratory Diagnostics, The First Clinical College, Fujian Medical University, Fuzhou, People’s Republic of China
- Fujian Key Laboratory of Laboratory Medicine, The First Affiliated Hospital, Fujian Medical University, Fuzhou, People’s Republic of China
- Gene Diagnosis Research Center, Fujian Medical University, Fuzhou, People’s Republic of China
| | - Jinlan Huang
- Department of Laboratory Medicine, The First Affiliated Hospital, Fujian Medical University, Fuzhou, People’s Republic of China
- Clinical Laboratory Diagnostics, The First Clinical College, Fujian Medical University, Fuzhou, People’s Republic of China
- Fujian Key Laboratory of Laboratory Medicine, The First Affiliated Hospital, Fujian Medical University, Fuzhou, People’s Republic of China
- Gene Diagnosis Research Center, Fujian Medical University, Fuzhou, People’s Republic of China
| | - Ling Jiang
- Department of Laboratory Medicine, The First Affiliated Hospital, Fujian Medical University, Fuzhou, People’s Republic of China
- Clinical Laboratory Diagnostics, The First Clinical College, Fujian Medical University, Fuzhou, People’s Republic of China
- Fujian Key Laboratory of Laboratory Medicine, The First Affiliated Hospital, Fujian Medical University, Fuzhou, People’s Republic of China
- Gene Diagnosis Research Center, Fujian Medical University, Fuzhou, People’s Republic of China
| | - Qishui Ou
- Department of Laboratory Medicine, The First Affiliated Hospital, Fujian Medical University, Fuzhou, People’s Republic of China
- Clinical Laboratory Diagnostics, The First Clinical College, Fujian Medical University, Fuzhou, People’s Republic of China
- Fujian Key Laboratory of Laboratory Medicine, The First Affiliated Hospital, Fujian Medical University, Fuzhou, People’s Republic of China
- Gene Diagnosis Research Center, Fujian Medical University, Fuzhou, People’s Republic of China
- Qishui Ou Department of Laboratory Medicine, The First Affiliated Hospital, Clinical Laboratory Diagnostics, The First Clinical College, Fujian Key Laboratory of Laboratory Medicine, The First Affiliated Hospital, Gene Diagnosis Research Center, Fujian Medical University, Fuzhou, People’s Republic of China
| | - Can Liu
- Department of Laboratory Medicine, The First Affiliated Hospital, Fujian Medical University, Fuzhou, People’s Republic of China
- Clinical Laboratory Diagnostics, The First Clinical College, Fujian Medical University, Fuzhou, People’s Republic of China
- Fujian Key Laboratory of Laboratory Medicine, The First Affiliated Hospital, Fujian Medical University, Fuzhou, People’s Republic of China
- Gene Diagnosis Research Center, Fujian Medical University, Fuzhou, People’s Republic of China
- Can Liu Department of Laboratory Medicine, The First Affiliated Hospital, Clinical Laboratory Diagnostics, The First Clinical College, Fujian Key Laboratory of Laboratory Medicine, The First Affiliated Hospital, Gene Diagnosis Research Center, Fujian Medical University, Fuzhou, People’s Republic of China
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14
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Kusunoki H, Hamaguchi I, Kobayashi N, Nagata T. Chemical shift assignments of a fusion protein comprising the C-terminal-deleted hepatitis B virus X protein BH3-like motif peptide and Bcl-x L. BIOMOLECULAR NMR ASSIGNMENTS 2022; 16:357-361. [PMID: 36044106 DOI: 10.1007/s12104-022-10104-4] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 07/29/2022] [Accepted: 08/23/2022] [Indexed: 06/15/2023]
Abstract
Chronic hepatitis B virus (HBV) infection is a major risk factor for the development of liver diseases including fibrosis, cirrhosis, and hepatocellular carcinoma (HCC). HBV has the multifunctional protein, HBV X protein (HBx, 154 residues), which plays key roles in HBV replication and liver disease development. Interaction of HBx through its BH3-like motif with the anti-apoptotic protein Bcl-xL leads to HBV replication and induction of apoptosis, resulting in HCC development. Our previous nuclear magnetic resonance (NMR) study revealed that the HBx BH3-like motif peptide (residues 101-136) binds to the common BH3-binding groove of Bcl-xL. Importantly, a C-terminal-truncated HBx, e.g., residues 1-120 of HBx, is strongly associated with the increased risk of HBV-related HCC development. However, the interaction mode between the C-terminal-truncated HBx and Bcl-xL remains unclear. To elucidate this interaction mode, the C-terminal-deleted HBx BH3-like motif peptide (residues 101-120) was used as a model peptide in this study. To facilitate the NMR analysis, we prepared a fusion protein of HBx (101-120) and Bcl-xL connected with five repeats of the glycine-serine dipeptide as a linker. Here, we report the 1H, 13C, and 15N resonance assignments of the fusion protein. This is the first step for the elucidation of the pathogenesis of liver diseases caused by the interaction between the C-terminal-truncated HBx and Bcl-xL.
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Affiliation(s)
- Hideki Kusunoki
- Research Center for Biological Products in the Next Generation, National Institute of Infectious Diseases, Musashimurayama, Tokyo, 208-0011, Japan.
| | - Isao Hamaguchi
- Research Center for Biological Products in the Next Generation, National Institute of Infectious Diseases, Musashimurayama, Tokyo, 208-0011, Japan
| | - Naohiro Kobayashi
- Laboratory for Advanced NMR Application and Development, RIKEN Center for Biosystems Dynamics Research, Yokohama, Kanagawa, 230-0045, Japan
| | - Takashi Nagata
- Institute of Advanced Energy, Kyoto University, Uji, Kyoto, 611-0011, Japan.
- Graduate School of Energy Science, Kyoto University, Uji, Kyoto, 611-0011, Japan.
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15
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Zhao R, Zhu Z, Geng R, Jiang X, Li W, Ou G. Inhibition of histone H3-H4 chaperone pathways rescues C. elegans sterility by H2B loss. PLoS Genet 2022; 18:e1010223. [PMID: 35679337 PMCID: PMC9216614 DOI: 10.1371/journal.pgen.1010223] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/18/2022] [Revised: 06/22/2022] [Accepted: 04/28/2022] [Indexed: 12/14/2022] Open
Abstract
Oncohistone mutations are crucial drivers for tumorigenesis, but how a living organism governs the loss-of-function oncohistone remains unclear. We generated a histone H2B triple knockout (3KO) strain in Caenorhabditis elegans, which decreased the embryonic H2B, disrupted cell divisions, and caused animal sterility. By performing genetic suppressor screens, we uncovered that mutations defective in the histone H3-H4 chaperone UNC-85 restored H2B 3KO fertility by decreasing chromatin H3-H4 levels. RNA interference of other H3-H4 chaperones or H3 or H4 histones also rescued H2B 3KO sterility. We showed that blocking H3-H4 chaperones recovered cell division in C. elegans carrying the oncohistone H2BE74K mutation that distorts the H2B-H4 interface and induces nucleosome instability. Our results indicate that reducing chromatin H3-H4 rescues the dysfunctional H2B in vivo and suggest that inhibiting H3-H4 chaperones may provide an effective therapeutic strategy for treating cancers resulting from loss-of-function H2B oncohistone.
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Affiliation(s)
- Ruixue Zhao
- Tsinghua-Peking Center for Life Sciences, Beijing Frontier Research Center for Biological Structure, McGovern Institute for Brain Research, School of Life Sciences and MOE Key Laboratory for Protein Science, Tsinghua University, Beijing, China
| | - Zhiwen Zhu
- Tsinghua-Peking Center for Life Sciences, Beijing Frontier Research Center for Biological Structure, McGovern Institute for Brain Research, School of Life Sciences and MOE Key Laboratory for Protein Science, Tsinghua University, Beijing, China
| | - Ruxu Geng
- Tsinghua-Peking Center for Life Sciences, Beijing Frontier Research Center for Biological Structure, McGovern Institute for Brain Research, School of Life Sciences and MOE Key Laboratory for Protein Science, Tsinghua University, Beijing, China
| | - Xuguang Jiang
- Tsinghua-Peking Center for Life Sciences, Beijing Frontier Research Center for Biological Structure, McGovern Institute for Brain Research, School of Life Sciences and MOE Key Laboratory for Protein Science, Tsinghua University, Beijing, China
| | - Wei Li
- School of Medicine, Tsinghua University, Beijing, China
| | - Guangshuo Ou
- Tsinghua-Peking Center for Life Sciences, Beijing Frontier Research Center for Biological Structure, McGovern Institute for Brain Research, School of Life Sciences and MOE Key Laboratory for Protein Science, Tsinghua University, Beijing, China
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16
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Zhang C, Xie Y, Lai R, Wu J, Guo Z. Nonsynonymous C1653T Mutation of Hepatitis B Virus X Gene Enhances Malignancy of Hepatocellular Carcinoma Cells. J Hepatocell Carcinoma 2022; 9:367-377. [PMID: 35535232 PMCID: PMC9078866 DOI: 10.2147/jhc.s348690] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/10/2021] [Accepted: 04/26/2022] [Indexed: 11/23/2022] Open
Abstract
Purpose Methods Results Conclusion
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Affiliation(s)
- Cuifang Zhang
- Department of Rheumatology and Immunology, The Fourth Hospital of Hebei Medical University, Shijiazhuang, People’s Republic of China
- Department of Oncology, The Pingshan County People’s Hospital, Shijiazhuang, People’s Republic of China
| | - Ying Xie
- Hebei Key Laboratory of Laboratory Animal Science, Hebei Medical University, Shijiazhuang, People’s Republic of China
| | - Ruixue Lai
- Department of Rheumatology and Immunology, The Fourth Hospital of Hebei Medical University, Shijiazhuang, People’s Republic of China
| | - Jianhua Wu
- Animal Center, The Fourth Hospital of Hebei Medical University, Shijiazhuang, People’s Republic of China
| | - Zhanjun Guo
- Department of Rheumatology and Immunology, The Fourth Hospital of Hebei Medical University, Shijiazhuang, People’s Republic of China
- Correspondence: Zhanjun Guo, Department of Rheumatology and Immunology, The Fourth Hospital of Hebei Medical University, 12 Jiankang Road, Shijiazhuang, 050011, People’s Republic of China, Tel + 86 311 8609 5734, Fax + 86 311 8609 5237, Email
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Canonical and Divergent N-Terminal HBx Isoform Proteins Unveiled: Characteristics and Roles during HBV Replication. Biomedicines 2021; 9:biomedicines9111701. [PMID: 34829930 PMCID: PMC8616016 DOI: 10.3390/biomedicines9111701] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/21/2021] [Revised: 11/11/2021] [Accepted: 11/11/2021] [Indexed: 11/16/2022] Open
Abstract
Hepatitis B virus (HBV) X protein (HBx) is a viral regulatory and multifunctional protein. It is well-known that the canonical HBx reading frame bears two phylogenetically conserved internal in-frame translational initiation codons at Met2 and Met3, thus possibly generating divergent N-terminal smaller isoforms during translation. Here, we demonstrate that the three distinct HBx isoforms are generated from the ectopically expressed HBV HBx gene, named XF (full-length), XM (medium-length), and XS (short-length); they display different subcellular localizations when expressed individually in cultured hepatoma cells. Particularly, the smallest HBx isoform, XS, displayed a predominantly cytoplasmic localization. To study HBx proteins during viral replication, we performed site-directed mutagenesis to target the individual or combinatorial expression of the HBx isoforms within the HBV viral backbone (full viral genome). Our results indicate that of all HBx isoforms, only the smallest HBx isoform, XS, can restore WT levels of HBV replication, and bind to the viral mini chromosome, thereby establishing an active chromatin state, highlighting its crucial activities during HBV replication. Intriguingly, we found that sequences of HBV HBx genotype H are devoid of the conserved Met3 position, and therefore HBV genotype H infection is naturally silent for the expression of the HBx XS isoform. Finally, we found that the HBx XM (medium-length) isoform shares significant sequence similarity with the N-terminus domain of the COMMD8 protein, a member of the copper metabolism MURR1 domain-containing (COMMD) protein family. This novel finding might facilitate studies on the phylogenetic origin of the HBV X protein. The identification and functional characterization of its isoforms will shift the paradigm by changing the concept of HBx from being a unique, canonical, and multifunctional protein toward the occurrence of different HBx isoforms, carrying out different overlapping functions at different subcellular localizations during HBV genome replication. Significantly, our current work unveils new crucial HBV targets to study for potential antiviral research, and human virus pathogenesis.
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Kong F, Zhang F, Liu X, Qin S, Yang X, Kong D, Pan X, You H, Zheng K, Tang R. Calcium signaling in hepatitis B virus infection and its potential as a therapeutic target. Cell Commun Signal 2021; 19:82. [PMID: 34362380 PMCID: PMC8349099 DOI: 10.1186/s12964-021-00762-7] [Citation(s) in RCA: 19] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/12/2021] [Accepted: 06/25/2021] [Indexed: 12/15/2022] Open
Abstract
As a ubiquitous second messenger, calcium (Ca2+) can interact with numerous cellular proteins to regulate multiple physiological processes and participate in a variety of diseases, including hepatitis B virus (HBV) infection, which is a major cause of hepatitis, fibrosis, cirrhosis, and hepatocellular carcinoma. In recent years, several studies have demonstrated that depends on the distinct Ca2+ channels on the plasma membrane, endoplasmic reticulum, as well as mitochondria, HBV can elevate cytosolic Ca2+ levels. Moreover, within HBV-infected cells, the activation of intracellular Ca2+ signaling contributes to viral replication via multiple molecular mechanisms. Besides, the available evidence indicates that targeting Ca2+ signaling by suitable pharmaceuticals is a potent approach for the treatment of HBV infection. In the present review, we summarized the molecular mechanisms related to the elevation of Ca2+ signaling induced by HBV to modulate viral propagation and the recent advances in Ca2+ signaling as a potential therapeutic target for HBV infection. Video Abstract.
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Affiliation(s)
- Fanyun Kong
- Jiangsu Key Laboratory of Immunity and Metabolism, Department of Pathogenic Biology and Immunology, Xuzhou Medical University, Xuzhou, 221004 Jiangsu China
| | - Fulong Zhang
- Imaging Department, The Second Affiliated Hospital of Shandong First Medical University, Taian, Shandong China
| | - Xiangye Liu
- Jiangsu Key Laboratory of Immunity and Metabolism, Department of Pathogenic Biology and Immunology, Xuzhou Medical University, Xuzhou, 221004 Jiangsu China
| | - Suping Qin
- Jiangsu Key Laboratory of Immunity and Metabolism, Department of Pathogenic Biology and Immunology, Xuzhou Medical University, Xuzhou, 221004 Jiangsu China
| | - Xiaoying Yang
- Jiangsu Key Laboratory of Immunity and Metabolism, Department of Pathogenic Biology and Immunology, Xuzhou Medical University, Xuzhou, 221004 Jiangsu China
| | - Delong Kong
- Jiangsu Key Laboratory of Immunity and Metabolism, Department of Pathogenic Biology and Immunology, Xuzhou Medical University, Xuzhou, 221004 Jiangsu China
| | - Xiucheng Pan
- Department of Infectious Diseases, The Affiliated Hospital of Xuzhou Medical University, Xuzhou, Jiangsu China
| | - Hongjuan You
- Jiangsu Key Laboratory of Immunity and Metabolism, Department of Pathogenic Biology and Immunology, Xuzhou Medical University, Xuzhou, 221004 Jiangsu China
| | - Kuiyang Zheng
- Jiangsu Key Laboratory of Immunity and Metabolism, Department of Pathogenic Biology and Immunology, Xuzhou Medical University, Xuzhou, 221004 Jiangsu China
- National Demonstration Center for Experimental Basic Medical Sciences Education, Xuzhou Medical University, Xuzhou, Jiangsu China
| | - Renxian Tang
- Jiangsu Key Laboratory of Immunity and Metabolism, Department of Pathogenic Biology and Immunology, Xuzhou Medical University, Xuzhou, 221004 Jiangsu China
- National Demonstration Center for Experimental Basic Medical Sciences Education, Xuzhou Medical University, Xuzhou, Jiangsu China
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19
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Zhou Q, Yan L, Xu B, Wang X, Sun X, Han N, Tang H, Huang F. Screening of the HBx transactivation domain interacting proteins and the function of interactor Pin1 in HBV replication. Sci Rep 2021; 11:14176. [PMID: 34238995 PMCID: PMC8266847 DOI: 10.1038/s41598-021-93584-z] [Citation(s) in RCA: 9] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/28/2020] [Accepted: 06/28/2021] [Indexed: 02/05/2023] Open
Abstract
Hepatitis B virus (HBV) X protein (HBx) has been determined to play a crucial role in the replication and transcription of HBV, and its biological functions mainly depend on the interaction with other host proteins. This study aims at screening the proteins that bind to the key functional domain of HBx by integrated proteomics. Proteins that specifically bind to the transactivation domain of HBx were selected by comparing interactors of full-length HBx and HBx-D5 truncation determined by glutathione-S-transferase (GST) pull-down assay combined with mass spectrometry (MS). The function of HBx interactor Pin1 in HBV replication was further investigated by in vitro experiments. In this study, a total of 189 proteins were identified from HepG2 cells that specifically bind to the transactivation domain of HBx by GST pull-down and subsequent MS. After gene ontology (GO) analysis, Pin1 was selected as the protein with the highest score in the largest cluster functioning in protein binding, and also classified into the cluster of proteins with the function of structural molecule activity, which is of great potential to be involved in HBV life cycle. The interaction between Pin1 and HBx has been further confirmed by Ni2+-NTA pulldown assay, co-immunoprecipitation, and immunofluorescence microscopy. HBsAg and HBeAg levels significantly decreased in Pin1 expression inhibited HepG2.2.15 cells. Besides, the inhibition of Pin1 expression in HepG2 cells impeded the restored replication of HBx-deficient HBV repaired by ectopic HBx expression. In conclusion, our study identified Pin1 as an interactor binds to the transactivation domain of HBx, and suggested the potential association between Pin1 and the function of HBx in HBV replication.
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Affiliation(s)
- Qiaoxia Zhou
- Department of Forensic Pathology, West China School of Preclinical and Forensic Medicine, Sichuan University, No. 17 Third Renmin Road North, Chengdu, 610041, People's Republic of China
| | - Libo Yan
- Center of Infectious Diseases, West China Hospital of Sichuan University, No.37 Guo Xue Xiang, Chengdu, 610041, People's Republic of China.,Division of Infectious Diseases, State Key Laboratory of Biotherapy, Sichuan University, Chengdu, 610041, People's Republic of China
| | - Baofu Xu
- Department of Forensic Pathology, West China School of Preclinical and Forensic Medicine, Sichuan University, No. 17 Third Renmin Road North, Chengdu, 610041, People's Republic of China.,Xinxiang Key Laboratory of Forensic Science Evidence, School of Forensic Medicine, Xinxiang Medical University, Xinxiang, 453003, People's Republic of China
| | - Xue'er Wang
- Department of Forensic Pathology, West China School of Preclinical and Forensic Medicine, Sichuan University, No. 17 Third Renmin Road North, Chengdu, 610041, People's Republic of China
| | - Xuehong Sun
- Department of Forensic Pathology, West China School of Preclinical and Forensic Medicine, Sichuan University, No. 17 Third Renmin Road North, Chengdu, 610041, People's Republic of China
| | - Ning Han
- Center of Infectious Diseases, West China Hospital of Sichuan University, No.37 Guo Xue Xiang, Chengdu, 610041, People's Republic of China.,Division of Infectious Diseases, State Key Laboratory of Biotherapy, Sichuan University, Chengdu, 610041, People's Republic of China
| | - Hong Tang
- Center of Infectious Diseases, West China Hospital of Sichuan University, No.37 Guo Xue Xiang, Chengdu, 610041, People's Republic of China. .,Division of Infectious Diseases, State Key Laboratory of Biotherapy, Sichuan University, Chengdu, 610041, People's Republic of China.
| | - Feijun Huang
- Department of Forensic Pathology, West China School of Preclinical and Forensic Medicine, Sichuan University, No. 17 Third Renmin Road North, Chengdu, 610041, People's Republic of China.
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20
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Maaroufi H, Potvin M, Cusson M, Levesque RC. Novel antimicrobial anionic cecropins from the spruce budworm feature a poly-L-aspartic acid C-terminus. Proteins 2021; 89:1205-1215. [PMID: 33973678 DOI: 10.1002/prot.26142] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/16/2020] [Revised: 04/28/2021] [Accepted: 05/06/2021] [Indexed: 01/10/2023]
Abstract
Cecropins form a family of amphipathic α-helical cationic peptides with broad-spectrum antibacterial properties and potent anticancer activity. The emergence of bacteria and cancer cells showing resistance to cationic antimicrobial peptides (CAMPs) has fostered a search for new, more selective and more effective alternatives to CAMPs. With this goal in mind, we looked for cecropin homologs in the genome and transcriptome of the spruce budworm, Choristoneura fumiferana. Not only did we find paralogs of the conventional cationic cecropins (Cfcec+ ), our screening also led to the identification of previously uncharacterized anionic cecropins (Cfcec- ), featuring a poly-l-aspartic acid C-terminus. Comparative peptide analysis indicated that the C-terminal helix of Cfcec- is amphipathic, unlike that of Cfcec+ , which is hydrophobic. Interestingly, molecular dynamics simulations pointed to the lower conformational flexibility of Cfcec- peptides, relative to that of Cfcec+ . Phylogenetic analysis suggests that the evolution of distinct Cfcec+ and Cfcec- peptides may have resulted from an ancient duplication event within the Lepidoptera. Finally, we found that both anionic and cationic cecropins contain a BH3-like motif (G-[KQR]-[HKQNR]-[IV]-[KQR]) that could interact with Bcl-2, a protein involved in apoptosis; this observation is congruent with previous reports indicating that cecropins induce apoptosis. Altogether, our observations suggest that cecropins may provide templates for the development of new anticancer drugs. We also estimated the antibacterial activity of Cfcec-2 and a ∆Cfce-2 peptide as AMPs by testing directly their ability in inhibiting bacterial growth in a disk diffusion assay and their potential for development of novel therapeutics.
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Affiliation(s)
- Halim Maaroufi
- Institut de biologie intégrative et des systèmes (IBIS), Université Laval, Quebec City, Canada
| | - Marianne Potvin
- Institut de biologie intégrative et des systèmes (IBIS), Université Laval, Quebec City, Canada
| | - Michel Cusson
- Natural Resources Canada, Canadian Forest Service, Laurentian Forestry Centre, Quebec City, Canada
| | - Roger C Levesque
- Institut de biologie intégrative et des systèmes (IBIS) and Faculté de médecine, Université Laval, Quebec City, Canada
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21
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Cui J, Zhang N, Liu Y, Zhang L, Gao C, Liu S. Microarray gene expression profiling provides insights into functions of TIPE2 in HBV-related apoptosis. Mol Immunol 2021; 131:137-143. [PMID: 33419563 DOI: 10.1016/j.molimm.2020.12.031] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/15/2020] [Revised: 11/06/2020] [Accepted: 12/24/2020] [Indexed: 10/22/2022]
Abstract
Tumor necrosis factor-α-induced protein-8 like-2 (TNFAIP8L2, TIPE2), a member of TNFAIP8 family, functions as a regulator in inflammation. Our previous studies showed that TIPE2 can negatively regulate HBV-specific CD8+ T lymphocyte functions. But the effect of TIPE2 on the apoptosis of HBV-infected hepatocytes which is very important for eliminating viruses remains unclear. Using gene expression microarray analysis, we find that TIPE2 deficiency can regulate the expression of apoptotic genes in liver tissues from HBV hydrodynamic injection (HI) mouse model. TIPE2 protein was detected in TUNEL staining positive hepatocytes in HBV-infected C57 mice. Interestingly, the TIPE2 expressed hepatocytes were just the HBV infected cells. Furthermore, TIPE2 upregulates the mRNA levels of FasL, Bim and TNFRsF1b which promote cells death, when TIPE2 was transfected into HepG2 cells in vitro. As a result, TIPE2 overexpression cells showed a higher number of apoptotic cells and increased level of cleavage caspase3 compared to controls. Those results indicate that TIPE2 participates in HBV infection by regulating apoptosis of virus-infected hepatocytes.
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Affiliation(s)
- Jian Cui
- Department of Immunology, Shandong University School of Basic Medical Science, Ji'nan, China; Key Laboratory of Infection and Immunity of Shandong Province, Shandong University School of Basic Medical Science, Jinan, China
| | - Na Zhang
- Department of Immunology, Shandong University School of Basic Medical Science, Ji'nan, China
| | - Ying Liu
- Department of Immunology, Shandong University School of Basic Medical Science, Ji'nan, China
| | - Lei Zhang
- Department of Immunology, Shandong University School of Basic Medical Science, Ji'nan, China
| | - Chengjiang Gao
- Department of Immunology, Shandong University School of Basic Medical Science, Ji'nan, China; Key Laboratory of Infection and Immunity of Shandong Province, Shandong University School of Basic Medical Science, Jinan, China
| | - Suxia Liu
- Department of Immunology, Shandong University School of Basic Medical Science, Ji'nan, China; Key Laboratory of Infection and Immunity of Shandong Province, Shandong University School of Basic Medical Science, Jinan, China.
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22
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Han N, Yan L, Wang X, Sun X, Huang F, Tang H. An updated literature review: how HBV X protein regulates the propagation of the HBV. Future Virol 2020. [DOI: 10.2217/fvl-2020-0251] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/08/2023]
Abstract
Chronic HBV infection constitutes a burden on human beings and is closely associated with hepatocellular carcinoma. The propagation of the HBV is determined by many factors, and the HBV X protein (HBx) could have a significant influence on this. HBx is a regulatory protein that can directly or indirectly interact with many cellular proteins to affect both the propagation of the HBV and the activity of the host cells. In this review, we summarized the possible mechanisms by which HBx regulates HBV replication at transcriptional and post-transcriptional levels in various experimental systems.
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Affiliation(s)
- Ning Han
- Center of Infectious Diseases, West China Hospital of Sichuan University, Chengdu, PR China
| | - Libo Yan
- Center of Infectious Diseases, West China Hospital of Sichuan University, Chengdu, PR China
| | - Xueer Wang
- Department of Forensic Pathology, West China School of Basic Science & Forensic Medicine, Sichuan University, Chengdu, Sichuan, PR China
| | - Xuehong Sun
- Department of Forensic Pathology, West China School of Basic Science & Forensic Medicine, Sichuan University, Chengdu, Sichuan, PR China
| | - Feijun Huang
- Department of Forensic Pathology, West China School of Basic Science & Forensic Medicine, Sichuan University, Chengdu, Sichuan, PR China
| | - Hong Tang
- Center of Infectious Diseases, West China Hospital of Sichuan University, Chengdu, PR China
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23
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Kusunoki H, Tanaka T, Kohno T, Kimura H, Hosoda K, Wakamatsu K, Hamaguchi I. NMR characterization of the interaction between Bcl-x L and the BH3-like motif of hepatitis B virus X protein. Biochem Biophys Res Commun 2019; 518:445-450. [PMID: 31439373 DOI: 10.1016/j.bbrc.2019.08.036] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/05/2019] [Accepted: 08/07/2019] [Indexed: 01/10/2023]
Abstract
Hepatitis B virus X protein (HBx) possesses a BH3-like motif that directly interacts with the anti-apoptotic proteins, Bcl-2 and Bcl-xL. Here we report the interaction between the HBx BH3-like motif and Bcl-xL, as revealed by nuclear magnetic resonance spectroscopy. Our results showed that this motif binds to the common BH3-binding hydrophobic groove on the surface of Bcl-xL, with a binding affinity of 89 μM. Furthermore, we examined the role of the tryptophan residue (Trp120) in this motif in Bcl-xL binding using three mutants. The W120A mutant showed weaker binding affinity (294 μM) to Bcl-xL, whereas the W120L and W120F mutants exhibited almost equivalent binding affinity to the wild-type. These results indicate that the bulky hydrophobic residues are important for Bcl-xL binding. The findings will be helpful in understanding the apoptosis networks between viral proteins and host factors.
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Affiliation(s)
- Hideki Kusunoki
- Department of Safety Research on Blood and Biological Products, National Institute of Infectious Diseases, 4-7-1 Gakuen, Musashi-Murayama, Tokyo, 208-0011, Japan.
| | - Toshiyuki Tanaka
- Graduate School of Life and Environmental Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki, 305-8572, Japan; Life Science Center, Tsukuba Advanced Research Alliance, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki, 305-8577, Japan
| | - Toshiyuki Kohno
- Department of Medical Informatics, Research and Development Center for Medical Education, Kitasato University School of Medicine, 1-15-1 Kitasato, Minami-ku, Sagamihara, Kanagawa, 252-0374, Japan
| | - Hirokazu Kimura
- Department of Health Science, Gunma Paz University Graduate School of Health Science, 1-7-1 Tonyamachi, Takasaki, Gunma, 370-0006, Japan
| | - Kazuo Hosoda
- Department of Molecular Science, Graduate School of Science and Technology, Gunma University, 1-5-1 Tenjin-cho, Kiryu, Gunma, 376-8515, Japan
| | - Kaori Wakamatsu
- Department of Molecular Science, Graduate School of Science and Technology, Gunma University, 1-5-1 Tenjin-cho, Kiryu, Gunma, 376-8515, Japan
| | - Isao Hamaguchi
- Department of Safety Research on Blood and Biological Products, National Institute of Infectious Diseases, 4-7-1 Gakuen, Musashi-Murayama, Tokyo, 208-0011, Japan.
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24
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Zhang TY, Chen HY, Cao JL, Xiong HL, Mo XB, Li TL, Kang XZ, Zhao JH, Yin B, Zhao X, Huang CH, Yuan Q, Xue D, Xia NS, Yuan YA. Structural and functional analyses of hepatitis B virus X protein BH3-like domain and Bcl-xL interaction. Nat Commun 2019; 10:3192. [PMID: 31324803 PMCID: PMC6642116 DOI: 10.1038/s41467-019-11173-1] [Citation(s) in RCA: 33] [Impact Index Per Article: 5.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/27/2017] [Accepted: 06/25/2019] [Indexed: 12/12/2022] Open
Abstract
Hepatitis B virus (HBV) X protein, HBx, interacts with anti-apoptotic Bcl-2 and Bcl-xL proteins through its BH3-like motif to promote HBV replication and cytotoxicity. Here we report the crystal structure of HBx BH3-like motif in complex with Bcl-xL where the BH3-like motif adopts a short α-helix to snuggle into a hydrophobic pocket in Bcl-xL via its noncanonical Trp120 residue and conserved Leu123 residue. This binding pocket is ~2 Å away from the canonical BH3-only binding pocket in structures of Bcl-xL with proapoptotic BH3-only proteins. Mutations altering Trp120 and Leu123 in HBx impair its binding to Bcl-xL in vitro and HBV replication in vivo, confirming the importance of this motif to HBV. A HBx BH3-like peptide, HBx-aa113-135, restores HBV replication from a HBx-null HBV replicon, while a shorter peptide, HBx-aa118-127, inhibits HBV replication. These results provide crucial structural and functional insights into drug designs for inhibiting HBV replication and treating HBV patients.
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Affiliation(s)
- Tian-Ying Zhang
- State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, School of Public Health and School of Life Sciences, Xiamen University, 361102, Xiamen, China.,National Institute of Diagnostics and Vaccine Development in Infectious Diseases, School of Public Health and School of Life Sciences, Xiamen University, 361102, Xiamen, China
| | - Hong-Ying Chen
- National University of Singapore (Suzhou) Research Institute, 377 Lin Quan Street, Suzhou Industrial Park, 215123, Jiangsu, China
| | - Jia-Li Cao
- State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, School of Public Health and School of Life Sciences, Xiamen University, 361102, Xiamen, China.,National Institute of Diagnostics and Vaccine Development in Infectious Diseases, School of Public Health and School of Life Sciences, Xiamen University, 361102, Xiamen, China
| | - Hua-Long Xiong
- State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, School of Public Health and School of Life Sciences, Xiamen University, 361102, Xiamen, China.,National Institute of Diagnostics and Vaccine Development in Infectious Diseases, School of Public Health and School of Life Sciences, Xiamen University, 361102, Xiamen, China
| | - Xiao-Bing Mo
- National University of Singapore (Suzhou) Research Institute, 377 Lin Quan Street, Suzhou Industrial Park, 215123, Jiangsu, China
| | - Tian-Liang Li
- Department of Molecular, Cellular and Developmental Biology, University of Colorado, Boulder, CO, 80309, USA
| | - Xiao-Zhen Kang
- State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, School of Public Health and School of Life Sciences, Xiamen University, 361102, Xiamen, China.,National Institute of Diagnostics and Vaccine Development in Infectious Diseases, School of Public Health and School of Life Sciences, Xiamen University, 361102, Xiamen, China
| | - Jing-Hua Zhao
- State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, School of Public Health and School of Life Sciences, Xiamen University, 361102, Xiamen, China.,National Institute of Diagnostics and Vaccine Development in Infectious Diseases, School of Public Health and School of Life Sciences, Xiamen University, 361102, Xiamen, China
| | - Bo Yin
- National University of Singapore (Suzhou) Research Institute, 377 Lin Quan Street, Suzhou Industrial Park, 215123, Jiangsu, China
| | - Xiang Zhao
- School of Life Sciences and Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, Tsinghua University, 100084, Beijing, China
| | - Cheng-Hao Huang
- State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, School of Public Health and School of Life Sciences, Xiamen University, 361102, Xiamen, China.,National Institute of Diagnostics and Vaccine Development in Infectious Diseases, School of Public Health and School of Life Sciences, Xiamen University, 361102, Xiamen, China
| | - Quan Yuan
- State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, School of Public Health and School of Life Sciences, Xiamen University, 361102, Xiamen, China. .,National Institute of Diagnostics and Vaccine Development in Infectious Diseases, School of Public Health and School of Life Sciences, Xiamen University, 361102, Xiamen, China.
| | - Ding Xue
- Department of Molecular, Cellular and Developmental Biology, University of Colorado, Boulder, CO, 80309, USA. .,School of Life Sciences and Collaborative Innovation Center for Diagnosis and Treatment of Infectious Diseases, Tsinghua University, 100084, Beijing, China.
| | - Ning-Shao Xia
- State Key Laboratory of Molecular Vaccinology and Molecular Diagnostics, School of Public Health and School of Life Sciences, Xiamen University, 361102, Xiamen, China. .,National Institute of Diagnostics and Vaccine Development in Infectious Diseases, School of Public Health and School of Life Sciences, Xiamen University, 361102, Xiamen, China.
| | - Y Adam Yuan
- National University of Singapore (Suzhou) Research Institute, 377 Lin Quan Street, Suzhou Industrial Park, 215123, Jiangsu, China.,Department of Biological Sciences and Centre for Bioimaging Sciences, National University of Singapore, 14 Science Drive 4, Singapore, 117543, Singapore
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25
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Raghunath A, Sundarraj K, Arfuso F, Sethi G, Perumal E. Dysregulation of Nrf2 in Hepatocellular Carcinoma: Role in Cancer Progression and Chemoresistance. Cancers (Basel) 2018; 10:cancers10120481. [PMID: 30513925 PMCID: PMC6315366 DOI: 10.3390/cancers10120481] [Citation(s) in RCA: 135] [Impact Index Per Article: 19.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/31/2018] [Revised: 11/29/2018] [Accepted: 11/29/2018] [Indexed: 12/23/2022] Open
Abstract
The liver executes versatile functions and is the chief organ for metabolism of toxicants/xenobiotics. Hepatocellular carcinoma (HCC) is the most common primary liver malignancy and the third foremost cause of cancer death worldwide. Oxidative stress is a key factor related with the development and progression of HCC. Nuclear factor erythroid 2 [NF-E2]-related factor 2 (Nrf2) is a cytosolic transcription factor, which regulates redox homeostasis by activating the expression of an array of antioxidant response element-dependent genes. Nrf2 displays conflicting roles in normal, healthy liver and HCC; in the former, Nrf2 offers beneficial effects, whereas in the latter it causes detrimental effects favouring the proliferation and survival of HCC. Sustained Nrf2 activation has been observed in HCC and facilitates its progression and aggressiveness. This review summarizes the role and mechanism(s) of action of Nrf2 dysregulation in HCC and therapeutic options that can be employed to modulate this transcription factor.
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Affiliation(s)
- Azhwar Raghunath
- Molecular Toxicology Laboratory, Department of Biotechnology, Bharathiar University, Coimbatore 641 046, Tamilnadu, India.
| | - Kiruthika Sundarraj
- Molecular Toxicology Laboratory, Department of Biotechnology, Bharathiar University, Coimbatore 641 046, Tamilnadu, India.
| | - Frank Arfuso
- Stem Cell and Cancer Biology Laboratory, School of Pharmacy and Biomedical Sciences, Curtin Health Innovation Research Institute, Curtin University, Perth, WA 6009, Australia.
| | - Gautam Sethi
- Department of Pharmacology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117600, Singapore.
| | - Ekambaram Perumal
- Molecular Toxicology Laboratory, Department of Biotechnology, Bharathiar University, Coimbatore 641 046, Tamilnadu, India.
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26
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Cai X, Zheng W, Shi X, Chen L, Liu Z, Li Z. HBx-Derived Constrained Peptides Inhibit the Secretion of Hepatitis B Virus Antigens. Mol Pharm 2018; 15:5646-5652. [PMID: 30375875 DOI: 10.1021/acs.molpharmaceut.8b00807] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/16/2022]
Abstract
Hepatitis B virus (HBV) infection is the primary cause of cirrhosis and liver cancer. Protein-protein interactions (PPIs) between HBV x protein (HBx) and its host targets, including Bcl-2, are important for cell death and viral replication. No modulators targeting these PPIs have been reported yet. Here, we developed HBx-derived constrained peptides generated by a facile macrocyclization method by joining two methionine side chains of unprotected peptides with chemoselective alkylating linkers. The resulting constrained peptides with improved cell permeability and binding affinity were effective anti-HBV modulators by blocking the secretion of viral antigens. This study clearly demonstrated HBx as a potentially important PPI target and the potential application of this efficient peptide macrocyclization strategy for targeting key PPIs.
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Affiliation(s)
- Xiaodan Cai
- State Key Laboratory of Chemical Oncogenomics, School of Chemical Biology and Biotechnology , Peking University Shenzhen Graduate School , Shenzhen 518055 , China
| | - Weihao Zheng
- State Key Laboratory of Chemical Oncogenomics, School of Chemical Biology and Biotechnology , Peking University Shenzhen Graduate School , Shenzhen 518055 , China.,Division of Experimental Medicine, Department of Medicine , University of California, San Francisco , San Francisco , California 94110 , United States
| | - Xiaodong Shi
- State Key Laboratory of Chemical Oncogenomics, School of Chemical Biology and Biotechnology , Peking University Shenzhen Graduate School , Shenzhen 518055 , China
| | - Longjian Chen
- State Key Laboratory of Chemical Oncogenomics, School of Chemical Biology and Biotechnology , Peking University Shenzhen Graduate School , Shenzhen 518055 , China
| | - Zhihong Liu
- State Key Laboratory of Chemical Oncogenomics, School of Chemical Biology and Biotechnology , Peking University Shenzhen Graduate School , Shenzhen 518055 , China
| | - Zigang Li
- State Key Laboratory of Chemical Oncogenomics, School of Chemical Biology and Biotechnology , Peking University Shenzhen Graduate School , Shenzhen 518055 , China
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27
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Yin H, Zhao L, Wang Y, Li S, Huo H, Chen H. Duck enteritis virus activates CaMKKβ-AMPK to trigger autophagy in duck embryo fibroblast cells via increased cytosolic calcium. Virol J 2018; 15:120. [PMID: 30081955 PMCID: PMC6090797 DOI: 10.1186/s12985-018-1029-0] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/23/2018] [Accepted: 07/18/2018] [Indexed: 01/03/2023] Open
Abstract
Background The results of our previous study showed that impaired cellular energy metabolism contributes to duck enteritis virus-induced autophagy via the 5`-adenosine monophosphate-activated protein kinase (AMPK)/tuberous sclerosis complex 2/mammalian target of rapamycin pathway in duck embryo fibroblast (DEF) cells. However, it remains unknown whether any other underlying mechanisms of AMPK activation are involved in autophagy induction. Methods The activity of CaMKKβ and AMPK in DEF cells infected with DEV were evaluated.The Effect of inhibitory activity of CaMKKβ on DEV-induced autophagy was investigated. In addtion to, the cytosolic calcium level in DEF cells infected with DEV were evaluated.The Effect of inhibitory cytosolic calcium level on DEV-induced autophagy was investigated. Results In this study, duck enteritis virus (DEV) infection activated CaMKKβ and its substrate molecule AMPK at 36, 48, and 60 h post-infection (hpi). STO-609, a CaMKKβ inhibitor, or CaMKKβ siRNA significantly inhibited the activation of DEV to AMPK, LC3I to LC3II transformation, and GFP-LC3 puncta distribution. In addition, inhibition of CaMKKβ activity also significantly reduced progeny DEV titer and gB protein expression. Besides, cytosolic calcium (Ca2+) was higher in DEV-infected cells than mock controls at 36, 48, and 60 hpi, respectively. Treatment of DEV-infected cells with 1,2-Bis (2-aminophenoxy) ethane-N, N, N′, N-tetraacetic acid (BAPTA-AM) significantly reduced intracellular Ca2+ ion concentrations, as well as CaMKKβ and AMPK activities, and subsequent autophagy, in addition to viral protein synthesis and viral titer. Conclusions These results showed that elevated [Ca2+]cyto-mediated activation of CaMKKβ managed the activation of AMPK, which then positively regulated autophagy, thereby providing further insight into DEV–host interactions.
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Affiliation(s)
- Haichang Yin
- State Key Laboratory of Veterinary Biotechnology, Heilongjiang Provincial Key Laboratory of Laboratory Animal and Comparative Medicine, Harbin Veterinary Research Institute, the Chinese Academy of Agriculture Sciences, 678 Haping Road, Harbin, 150069, People's Republic of China.,College of Life Science and Agriculture Forestry, Qiqihar University, Qiqihar, 161006, China.,Heilongjiang Provincial Key Laboratory of Resistance Gene Engineering and Protection of Biodiversity in Cold Areas, Qiqihar, Heilongjiang, 161006, China
| | - Lili Zhao
- State Key Laboratory of Veterinary Biotechnology, Heilongjiang Provincial Key Laboratory of Laboratory Animal and Comparative Medicine, Harbin Veterinary Research Institute, the Chinese Academy of Agriculture Sciences, 678 Haping Road, Harbin, 150069, People's Republic of China
| | - Yiping Wang
- State Key Laboratory of Veterinary Biotechnology, Heilongjiang Provincial Key Laboratory of Laboratory Animal and Comparative Medicine, Harbin Veterinary Research Institute, the Chinese Academy of Agriculture Sciences, 678 Haping Road, Harbin, 150069, People's Republic of China
| | - Siqi Li
- State Key Laboratory of Veterinary Biotechnology, Heilongjiang Provincial Key Laboratory of Laboratory Animal and Comparative Medicine, Harbin Veterinary Research Institute, the Chinese Academy of Agriculture Sciences, 678 Haping Road, Harbin, 150069, People's Republic of China
| | - Hong Huo
- State Key Laboratory of Veterinary Biotechnology, Heilongjiang Provincial Key Laboratory of Laboratory Animal and Comparative Medicine, Harbin Veterinary Research Institute, the Chinese Academy of Agriculture Sciences, 678 Haping Road, Harbin, 150069, People's Republic of China
| | - Hongyan Chen
- State Key Laboratory of Veterinary Biotechnology, Heilongjiang Provincial Key Laboratory of Laboratory Animal and Comparative Medicine, Harbin Veterinary Research Institute, the Chinese Academy of Agriculture Sciences, 678 Haping Road, Harbin, 150069, People's Republic of China.
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Wang Y, Wang H, Pan S, Hu T, Shen J, Zheng H, Xie S, Xie Y, Lu R, Guo L. Capable Infection of Hepatitis B Virus in Diffuse Large B-cell Lymphoma. J Cancer 2018; 9:1575-1581. [PMID: 29760795 PMCID: PMC5950586 DOI: 10.7150/jca.24384] [Citation(s) in RCA: 23] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/15/2017] [Accepted: 02/08/2018] [Indexed: 01/05/2023] Open
Abstract
Background: Diffuse large B-cell lymphoma (DLBCL) is the most common pathological type of non-Hodgkin lymphoma (NHL). It is strongly correlated to the host immunity and infection status. Aim: This study tested the hypothesis that hepatitis B virus (HBV) infection is also associated with DLBCL. Methods: Clinical analysis of the correlation between DLBCL and HBV infection, detection of HBV in situ of DLBCL tissue, and biological experiments that determined whether HBV infects B lymphocytes were conducted. Results: Our long-term clinical data showed that the positive rate of serum HBV was significantly increased in DLBCL patients (23.6%) compared to that in the general Chinese population (7.2%, P<0.001), especially in advanced stage lymphoma patients (P=0.003). In addition, HBV could infect B lymphocytes in vitro and the HBV antigen and nucleic acid could be detected intracellularly. Hepatitis B x protein (HBx) was also strongly expressed in tissues from DLBCL patients that were serum HBV surface antigen (HBsAg) positive. These patients responded less well to therapy with an odds ratio (OR) of 3.04. Conclusions: HBV can infect B lymphocytes. It might be related to the development of DLBCL and may also impact the efficacy of treatment.
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Affiliation(s)
- Yanchun Wang
- Department of Clinical Laboratory, Fudan University Shanghai Cancer Center, Shanghai, China.,Department of Oncology, Shanghai Medical College, Fudan University, Shanghai, China
| | - Huijie Wang
- Department of Oncology, Shanghai Medical College, Fudan University, Shanghai, China.,Department of Oncology, Fudan University Shanghai Cancer Center, Shanghai, China
| | - Shaokun Pan
- Department of Pathogen Biology, Shanghai Medical College, Fudan University, Shanghai, China
| | - Tao Hu
- Department of Clinical Laboratory, Fudan University Shanghai Cancer Center, Shanghai, China
| | - Jiabin Shen
- Department of Clinical Laboratory, Fudan University Shanghai Cancer Center, Shanghai, China
| | - Hui Zheng
- Department of Clinical Laboratory, Fudan University Shanghai Cancer Center, Shanghai, China.,Department of Oncology, Shanghai Medical College, Fudan University, Shanghai, China
| | - Suhong Xie
- Department of Clinical Laboratory, Fudan University Shanghai Cancer Center, Shanghai, China.,Department of Oncology, Shanghai Medical College, Fudan University, Shanghai, China
| | - Youhua Xie
- Department of Pathogen Biology, Shanghai Medical College, Fudan University, Shanghai, China
| | - Renquan Lu
- Department of Clinical Laboratory, Fudan University Shanghai Cancer Center, Shanghai, China.,Department of Oncology, Shanghai Medical College, Fudan University, Shanghai, China
| | - Lin Guo
- Department of Clinical Laboratory, Fudan University Shanghai Cancer Center, Shanghai, China.,Department of Oncology, Shanghai Medical College, Fudan University, Shanghai, China
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29
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Lei L, Wu S, Lu S, Liu M, Song Y, Fu Z, Shi H, Raley-Susman KM, He D. Microplastic particles cause intestinal damage and other adverse effects in zebrafish Danio rerio and nematode Caenorhabditis elegans. THE SCIENCE OF THE TOTAL ENVIRONMENT 2018; 619-620:1-8. [PMID: 29136530 DOI: 10.1016/j.scitotenv.2017.11.103] [Citation(s) in RCA: 732] [Impact Index Per Article: 104.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 09/20/2017] [Revised: 11/08/2017] [Accepted: 11/08/2017] [Indexed: 05/18/2023]
Abstract
Microplastics have been frequently detected in aquatic environments, and there are increasing concerns about potential effects on biota. In this study, zebrafish Danio rerio and nematode Caenorhabditis elegans were used as model organisms for microplastic exposure in freshwater pelagic (i.e. water column) and benthic (i.e. sediment) environments. We investigated the toxic effects of five common types of microplastics: polyamides (PA), polyethylene (PE), polypropylene (PP), polyvinyl chloride (PVC) and polystyrene (PS) particles. Results showed no or low lethality in D. rerio after exposure for 10d at 0.001-10.0mgL-1 microplastics. The PA, PE, PP and/or PVC microplastics with ~70μm size caused intestinal damage including cracking of villi and splitting of enterocytes. Exposure to 5.0mgm-2 microplastics for 2d significantly inhibited survival rates, body length and reproduction of C. elegans. Moreover, exposure to microplastics reduced calcium levels but increased expression of the glutathione S-transferase 4 enzyme in the intestine, which indicates intestinal damage and oxidative stress are major effects of microplastic exposure. Among 0.1, 1.0 and 5.0μm sizes of fluorescently labeled PS, 1.0μm particles caused the highest lethality, the maximum accumulation, the lowest Ca2+ level in the intestine and the highest expression of glutathione S-transferase 4 in nematodes. Taken together, these findings suggest that intestinal damage is a key effect of microplastics; and that the toxicity of microplastics is closely dependent on their size, rather than their composition.
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Affiliation(s)
- Lili Lei
- Lab of Toxicology, School of Ecological and Environmental Sciences, East China Normal University, 500# DongChuan RD, Shanghai 200241, China
| | - Siyu Wu
- Lab of Toxicology, School of Ecological and Environmental Sciences, East China Normal University, 500# DongChuan RD, Shanghai 200241, China
| | - Shibo Lu
- Lab of Toxicology, School of Ecological and Environmental Sciences, East China Normal University, 500# DongChuan RD, Shanghai 200241, China
| | - Mengting Liu
- Lab of Toxicology, School of Ecological and Environmental Sciences, East China Normal University, 500# DongChuan RD, Shanghai 200241, China
| | - Yang Song
- Lab of Toxicology, School of Ecological and Environmental Sciences, East China Normal University, 500# DongChuan RD, Shanghai 200241, China
| | - Zhenhuan Fu
- Lab of Toxicology, School of Ecological and Environmental Sciences, East China Normal University, 500# DongChuan RD, Shanghai 200241, China
| | - Huahong Shi
- State Key Laboratory of Estuarine and Coastal Research, East China Normal University, Shanghai 200062, China
| | | | - Defu He
- Lab of Toxicology, School of Ecological and Environmental Sciences, East China Normal University, 500# DongChuan RD, Shanghai 200241, China; Shanghai Key Laboratory for Urban Ecological Processes and Eco-Restoration, East China Normal University, Shanghai 200241, China.
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30
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Yao JH, Liu ZJ, Yi JH, Wang J, Liu YN. Hepatitis B Virus X Protein Upregulates Intracellular Calcium Signaling by Binding C-terminal of Orail Protein. Curr Med Sci 2018; 38:26-34. [PMID: 30074148 DOI: 10.1007/s11596-018-1843-z] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/20/2017] [Revised: 01/15/2018] [Indexed: 12/15/2022]
Abstract
Hepatitis B virus X (HBx) protein plays a pivotal role in the development of hepatitis B virus (HBV)-associated hepatocellular carcinoma. Although regulation of cytosolic calcium is essential for HBV replication and is mediated by HBx protein, the mechanism of HBx protein regulating intracellular calcium level remains poorly understood. The present study examined whether HBx protein elevated the intracellular calcium through interacting with storeoperated calcium entry (SOCE) components, Orail and stromal interaction molecule 1, and then identified the targets of HBx protein, with an attempt to understand the mechanism of HBx protein upsetting intracellular calcium homeostasis. By employing co-immunoprecipitation and GST-pull-down assay, we found that Orail protein interacted with HBx protein, and the C-terminus of Orail was implicated in the interaction. Confocal microscopy also revealed that HBx protein could co-localize with full-length Orail protein in HEK293 cells. Moreover, live cell calcium imaging exhibited that HBx protein elevated intracellular calcium, possibly by binding to SOCE components. Our results suggest that HBx protein binds to STIM1-Orail complexes to positively regulate the activity of plasma membrane store-operated calcium channels.
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Affiliation(s)
- Jing-Hong Yao
- Department of Infectious Diseases, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430022, China
| | - Zi-Jian Liu
- Department of Anatomy, School of Basic Medical Sciences, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430030, China.
| | - Jian-Hua Yi
- Department of Infectious Diseases, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430022, China
| | - Jun Wang
- Department of Gastroenterology, the Central Hospital of Wuhan, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430014, China
| | - Ya-Nan Liu
- Department of Infectious Diseases, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, 430022, China
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31
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Casciano JC, Bouchard MJ. Hepatitis B virus X protein modulates cytosolic Ca 2+ signaling in primary human hepatocytes. Virus Res 2018; 246:23-27. [PMID: 29307794 DOI: 10.1016/j.virusres.2018.01.001] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/19/2017] [Revised: 01/02/2018] [Accepted: 01/03/2018] [Indexed: 12/22/2022]
Abstract
Worldwide, approximately 240 million people are chronically infected with the hepatitis B virus (HBV); chronic HBV infection is associated with the development of life-threatening liver diseases. The HBV HBx protein alters hepatocyte physiology to promote HBV replication. We previously reported that HBx modulates calcium signaling to stimulate HBV replication in human hepatoblastoma HepG2 cells and primary rat hepatocytes. Whether HBx modulates calcium signaling in a primary human hepatocyte, the natural site of an HBV infection, has not been determined. Here, we report the effect of HBx on calcium signaling in primary human hepatocytes and show that HBx modulates calcium signaling via enhanced calcium entry through store-operated calcium channels and elevated mitochondrial calcium, similar to HBx effects in HepG2 cells and primary rat hepatocytes. In addition to demonstrating that HBV and HBx affect calcium signaling in human hepatocytes, these studies also show that HBV and HBx regulation of calcium signaling is identical in primary human and rat hepatocytes, further validating the use of cultured primary rat hepatocytes for HBV studies.
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Affiliation(s)
- Jessica C Casciano
- Program in Molecular and Cellular Biology and Genetics, Graduate School of Biomedical Sciences and Professional Studies, Drexel University College of Medicine, Philadelphia, PA, USA
| | - Michael J Bouchard
- Department of Biochemistry and Molecular Biology, Drexel University College of Medicine, Philadelphia, PA, USA.
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32
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van Sluijs L, Pijlman GP, Kammenga JE. Why do Individuals Differ in Viral Susceptibility? A Story Told by Model Organisms. Viruses 2017; 9:E284. [PMID: 28973976 PMCID: PMC5691635 DOI: 10.3390/v9100284] [Citation(s) in RCA: 18] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/22/2017] [Revised: 09/22/2017] [Accepted: 09/26/2017] [Indexed: 01/30/2023] Open
Abstract
Viral susceptibility and disease progression is determined by host genetic variation that underlies individual differences. Genetic polymorphisms that affect the phenotype upon infection have been well-studied for only a few viruses, such as HIV-1 and Hepatitis C virus. However, even for well-studied viruses the genetic basis of individual susceptibility differences remains elusive. Investigating the effect of causal polymorphisms in humans is complicated, because genetic methods to detect rare or small-effect polymorphisms are limited and genetic manipulation is not possible in human populations. Model organisms have proven a powerful experimental platform to identify and characterize polymorphisms that underlie natural variations in viral susceptibility using quantitative genetic tools. We summarize and compare the genetic tools available in three main model organisms, Mus musculus, Drosophila melanogaster, and Caenorhabditis elegans, and illustrate how these tools can be applied to detect polymorphisms that determine the viral susceptibility. Finally, we analyse how candidate polymorphisms from model organisms can be used to shed light on the underlying mechanism of individual variation. Insights in causal polymorphisms and mechanisms underlying individual differences in viral susceptibility in model organisms likely provide a better understanding in humans.
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Affiliation(s)
- Lisa van Sluijs
- Laboratory of Nematology, Wageningen University, 6708 PB Wageningen, The Netherlands.
- Laboratory of Virology, Wageningen University, 6708 PB Wageningen, The Netherlands.
| | - Gorben P Pijlman
- Laboratory of Virology, Wageningen University, 6708 PB Wageningen, The Netherlands.
| | - Jan E Kammenga
- Laboratory of Nematology, Wageningen University, 6708 PB Wageningen, The Netherlands.
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33
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Interference of Apoptosis by Hepatitis B Virus. Viruses 2017; 9:v9080230. [PMID: 28820498 PMCID: PMC5580487 DOI: 10.3390/v9080230] [Citation(s) in RCA: 27] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/30/2017] [Revised: 08/07/2017] [Accepted: 08/10/2017] [Indexed: 12/16/2022] Open
Abstract
Hepatitis B virus (HBV) causes liver diseases that have been a consistent problem for human health, leading to more than one million deaths every year worldwide. A large proportion of hepatocellular carcinoma (HCC) cases across the world are closely associated with chronic HBV infection. Apoptosis is a programmed cell death and is frequently altered in cancer development. HBV infection interferes with the apoptosis signaling to promote HCC progression and viral proliferation. The HBV-mediated alteration of apoptosis is achieved via interference with cellular signaling pathways and regulation of epigenetics. HBV X protein (HBX) plays a major role in the interference of apoptosis. There are conflicting reports on the HBV interference of apoptosis with the majority showing inhibition of and the rest reporting induction of apoptosis. In this review, we described recent studies on the mechanisms of the HBV interference with the apoptosis signaling during the virus infection and provided perspective.
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34
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Kusunoki H, Tanaka T, Kohno T, Kimura H, Hosoda K, Wakamatsu K, Hamaguchi I. Expression, purification and characterization of hepatitis B virus X protein BH3-like motif-linker-Bcl-x L fusion protein for structural studies. Biochem Biophys Rep 2016; 9:159-165. [PMID: 29114584 PMCID: PMC5632712 DOI: 10.1016/j.bbrep.2016.12.006] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/01/2016] [Revised: 12/12/2016] [Accepted: 12/19/2016] [Indexed: 11/24/2022] Open
Abstract
Hepatitis B virus X protein (HBx) is a multifunctional protein that interacts directly with many host proteins. For example, HBx interacts with anti-apoptotic proteins, Bcl-2 and Bcl-xL, through its BH3-like motif, which leads to elevated cytosolic calcium levels, efficient viral DNA replication and the induction of apoptosis. To facilitate sample preparation and perform detailed structural characterization of the complex between HBx and Bcl-xL, we designed and purified a recombinant HBx BH3-like motif-linker-Bcl-xL fusion protein produced in E. coli. The fusion protein was characterized by size exclusion chromatography, circular dichroism and nuclear magnetic resonance experiments. Our results show that the fusion protein is a monomer in aqueous solution, forms a stable intramolecular complex, and likely retains the native conformation of the complex between Bcl-xL and the HBx BH3-like motif. Furthermore, the HBx BH3-like motif of the intramolecular complex forms an α-helix. These observations indicate that the fusion protein should facilitate structural studies aimed at understanding the interaction between HBx and Bcl-xL at the atomic level.
Soluble HBx BH3-like motif-linker-Bcl-xL fusion protein was produced in E. coli. The fusion protein behaves as a monomer and forms a stable intramolecular complex. The HBx BH3-like motif of the fusion protein forms an α-helix. The fusion protein likely retains the native conformation of the complex.
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Affiliation(s)
- Hideki Kusunoki
- Department of Research on Blood and Biological Products, National Institute of Infectious Diseases, 4-7-1 Gakuen, Musashimurayama, Tokyo 208-0011, Japan
| | - Toshiyuki Tanaka
- Graduate School of Life and Environmental Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8572, Japan
| | - Toshiyuki Kohno
- Department of Biochemistry, Kitasato University School of Medicine, 1-15-1 Kitasato, Minami-ku, Sagamihara, Kanagawa 252-0374, Japan
| | - Hirokazu Kimura
- Infectious Disease Surveillance Center, National Institute of Infectious Diseases, 4-7-1 Gakuen, Musashimurayama, Tokyo 208-0011, Japan
| | - Kazuo Hosoda
- Department of Molecular Science, Graduate School of Science and Technology, Gunma University, 1-5-1 Tenjin-cho, Kiryu, Gunma 376-8515, Japan
| | - Kaori Wakamatsu
- Department of Molecular Science, Graduate School of Science and Technology, Gunma University, 1-5-1 Tenjin-cho, Kiryu, Gunma 376-8515, Japan
| | - Isao Hamaguchi
- Department of Research on Blood and Biological Products, National Institute of Infectious Diseases, 4-7-1 Gakuen, Musashimurayama, Tokyo 208-0011, Japan
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35
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Kim H, Lee SA, Kim BJ. X region mutations of hepatitis B virus related to clinical severity. World J Gastroenterol 2016; 22:5467-5478. [PMID: 27350725 PMCID: PMC4917607 DOI: 10.3748/wjg.v22.i24.5467] [Citation(s) in RCA: 28] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 03/24/2016] [Revised: 05/17/2016] [Accepted: 06/02/2016] [Indexed: 02/06/2023] Open
Abstract
Chronic hepatitis B virus (HBV) infection remains a major health problem, with more than 240 million people chronically infected worldwide and potentially 650000 deaths per year due to advanced liver diseases including liver cirrhosis and hepatocellular carcinoma (HCC). HBV-X protein (HBx) contributes to the biology and pathogenesis of HBV via stimulating virus replication or altering host gene expression related to HCC. The HBV X region contains only 465 bp encoding the 16.5 kDa HBx protein, which also contains several critical cis-elements such as enhancer II, the core promoter and the microRNA-binding region. Thus, mutations in this region may affect not only the HBx open reading frame but also the overlapped cis-elements. Recently, several types of HBx mutations significantly associated with clinical severity have been described, although the functional mechanism in most of these cases remains unsolved. This review article will mainly focus on the HBx mutations proven to be significantly related to clinical severity via epidemiological studies.
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Cai JG, Zhu TT, Liu FR, Huang ZG. Correlation between serum levels of Golgi protein 73 and antiviral efficacy of entecavir in patients with chronic hepatitis B. Shijie Huaren Xiaohua Zazhi 2016; 24:2379-2383. [DOI: 10.11569/wcjd.v24.i15.2379] [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 investigate the correlation between serum levels of Golgi protein 73 (GP73) and antiviral efficacy of entecavir (ETV) in patients with chronic hepatitis B (CHB) to evaluate the value of GP73 in monitoring the progression and outcome of CHB.
METHODS: A total of 483 CHB patients were included, and 63 healthy persons were used as controls. Serum levels of GP73 were detected by ELISA, and liver tissues were collected from 51 CHB patients for detecting GP73 expression by immunohistochemical staining. The correlation between serum GP73 concentrations and GP73 expression in liver tissues was examined. One hundred and eighty-three CHB patients received ETV antiviral treatment for more than 1 year, and serum GP73 concentrations were determined before treatment and 1, 3, 6, 9, and 12 after treatment.
RESULTS: In patients with chronic HBV infection, serum GP73 level was significantly higher than that of the healthy control group (F = 191.60, P = 0.000). With the progression of the disease, serum GP73 concentration continued to significantly rise (P = 0.000). Serum GP73 levels were positively correlated with severity of chronic HBV infection (r = 0.576, P = 0.000). With the increase of GP73 expression in liver tissue, serum GP73 concentrations also increased simultaneously (F = 7.285, P = 0.01). Serum GP73 concentration was positively correlated with the expression of GP73 in liver tissues (r = 0.592, P = 0.00). In 186 CHB patients treated with ETV for more than 1 years, with the recovery of the disease, serum GP73 concentrations significantly decreased, and there was a significant difference between before and after treatment (P = 0.005).
CONCLUSION: Serum GP73 levels are closely related to the disease progression of chronic HBV infection. The decrease of serum GP73 concentration in the course of ETV antiviral therapy suggests that the liver inflammatory injury is relieved. Serum GP73 can be used to monitor the prognosis of patients with chronic liver disease.
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Porcine Circovirus Type 2 Activates CaMMKβ to Initiate Autophagy in PK-15 Cells by Increasing Cytosolic Calcium. Viruses 2016; 8:v8050135. [PMID: 27213427 PMCID: PMC4885090 DOI: 10.3390/v8050135] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/12/2016] [Revised: 05/11/2016] [Accepted: 05/12/2016] [Indexed: 12/25/2022] Open
Abstract
Porcine circovirus type 2 (PCV2) induces autophagy via the 5′ adenosine monophosphate-activated protein kinase (AMPK)/extracellular signal-regulated kinase (ERK)/tuberous sclerosis complex 2 (TSC2)/mammalian target of rapamycin (mTOR) pathway in pig kidney PK-15 cells. However, the underlying mechanisms of AMPK activation in autophagy induction remain unknown. With specific inhibitors and RNA interference (RNAi), we show that PCV2 infection upregulated calcium/calmodulin-dependent protein kinase kinase-beta (CaMKKβ) by increasing cytosolic Ca2+ via inositol 1,4,5-trisphosphate receptor (IP3R). Elevation of cytosolic calcium ion (Ca2+) did not seem to involve inositol 1,4,5-trisphosphate (IP3) release from phosphatidylinositol 4,5-bisphosphate (PIP2) by phosphoinositide phospholipase C-gamma (PLC-γ). CaMKKβ then activated both AMPK and calcium/calmodulin-dependent protein kinase I (CaMKI). PCV2 employed CaMKI and Trp-Asp (WD) repeat domain phosphoinositide-interacting protein 1 (WIPI1) as another pathway additional to AMPK signaling in autophagy initiation. Our findings could help better understanding of the signaling pathways of autophagy induction as part of PCV2 pathogenesis. Further research is warranted to study if PCV2 interacts directly with IP3R or indirectly with the molecules that antagonize IP3R activity responsible for increased cytosolic Ca2+ both in PK-15 cells and PCV2-targeted primary cells from pigs.
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Structural and biochemical analysis of Bcl-2 interaction with the hepatitis B virus protein HBx. Proc Natl Acad Sci U S A 2016; 113:2074-9. [PMID: 26858413 DOI: 10.1073/pnas.1525616113] [Citation(s) in RCA: 43] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/25/2022] Open
Abstract
HBx is a hepatitis B virus protein that is required for viral infectivity and replication. Anti-apoptotic Bcl-2 family members are thought to be among the important host targets of HBx. However, the structure and function of HBx are poorly understood and the molecular mechanism of HBx-induced carcinogenesis remains unknown. In this study, we report biochemical and structural characterization of HBx. The recombinant HBx protein contains metal ions, in particular iron and zinc. A BH3-like motif in HBx (residues 110-135) binds Bcl-2 with a dissociation constant of ∼193 μM, which is drastically lower than that for a canonical BH3 motif from Bim or Bad. Structural analysis reveals that, similar to other BH3 motifs, the BH3-like motif of HBx adopts an amphipathic α-helix and binds the conserved BH3-binding groove on Bcl-2. Unlike the helical Bim or Bad BH3 motif, the C-terminal portion of the bound HBx BH3-like motif has an extended conformation and makes considerably fewer interactions with Bcl-2. These observations suggest that HBx may modulate Bcl-2 function in a way that is different from that of the classical BH3-only proteins.
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39
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Bacillus thuringiensis Crystal Protein Cry6Aa Triggers Caenorhabditis elegans Necrosis Pathway Mediated by Aspartic Protease (ASP-1). PLoS Pathog 2016; 12:e1005389. [PMID: 26795495 PMCID: PMC4721865 DOI: 10.1371/journal.ppat.1005389] [Citation(s) in RCA: 47] [Impact Index Per Article: 5.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/22/2015] [Accepted: 12/18/2015] [Indexed: 02/07/2023] Open
Abstract
Cell death plays an important role in host-pathogen interactions. Crystal proteins (toxins) are essential components of Bacillus thuringiensis (Bt) biological pesticides because of their specific toxicity against insects and nematodes. However, the mode of action by which crystal toxins to induce cell death is not completely understood. Here we show that crystal toxin triggers cell death by necrosis signaling pathway using crystal toxin Cry6Aa-Caenorhabditis elegans toxin-host interaction system, which involves an increase in concentrations of cytoplasmic calcium, lysosomal lyses, uptake of propidium iodide, and burst of death fluorescence. We find that a deficiency in the necrosis pathway confers tolerance to Cry6Aa toxin. Intriguingly, the necrosis pathway is specifically triggered by Cry6Aa, not by Cry5Ba, whose amino acid sequence is different from that of Cry6Aa. Furthermore, Cry6Aa-induced necrosis pathway requires aspartic protease (ASP-1). In addition, ASP-1 protects Cry6Aa from over-degradation in C. elegans. This is the first demonstration that deficiency in necrosis pathway confers tolerance to Bt crystal protein, and that Cry6A triggers necrosis represents a newly added necrosis paradigm in the C. elegans. Understanding this model could lead to new strategies for nematode control. Necrosis contributes to many devastating pathological conditions, such as neurodegenerative diseases and microbial pathogenesis. Bacillus thuringiensis crystal proteins are effective biopesticides. Our study reveals that B. thuringiensis Cry6Aa protein triggers the necrosis pathway using Caenorhabditis elegans as a model. We show that aspartic protease ASP-1 is required for Cry6Aa protein-induced necrosis, whereas intrinsic insults induce necrosis mediated by ASP-3 and ASP-4. Our findings contribute to the understanding of the mechanism of Bt crystal protein action and host-pathogen interactions. Because necrosis mechanisms are conserved from nematodes to humans, the fact that necrosis can be induced by Cry6Aa provides a model system for studying necrosis mechanisms in human diseases.
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Hepatocyte Factor JMJD5 Regulates Hepatitis B Virus Replication through Interaction with HBx. J Virol 2016; 90:3530-42. [PMID: 26792738 DOI: 10.1128/jvi.02776-15] [Citation(s) in RCA: 26] [Impact Index Per Article: 2.9] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/29/2015] [Accepted: 12/30/2015] [Indexed: 12/12/2022] Open
Abstract
UNLABELLED Hepatitis B virus (HBV) is a causative agent for chronic liver diseases such as hepatitis, cirrhosis, and hepatocellular carcinoma (HCC). HBx protein encoded by the HBV genome plays crucial roles not only in pathogenesis but also in replication of HBV. Although HBx has been shown to bind to a number of host proteins, the molecular mechanisms by which HBx regulates HBV replication are largely unknown. In this study, we identified jumonji C-domain-containing 5 (JMJD5) as a novel binding partner of HBx interacting in the cytoplasm. DNA microarray analysis revealed that JMJD5-knockout (JMJD5KO) Huh7 cells exhibited a significant reduction in the expression of transcriptional factors involved in hepatocyte differentiation, such as HNF4A, CEBPA, and FOXA3. We found that hydroxylase activity of JMJD5 participates in the regulation of these transcriptional factors. Moreover, JMJD5KO Huh7 cells exhibited a severe reduction in HBV replication, and complementation of HBx expression failed to rescue replication of a mutant HBV deficient in HBx, suggesting that JMJD5 participates in HBV replication through an interaction with HBx. We also found that replacing Gly(135) with Glu in JMJD5 abrogates binding with HBx and replication of HBV. Moreover, the hydroxylase activity of JMJD5 was crucial for HBV replication. Collectively, these results suggest that direct interaction of JMJD5 with HBx facilitates HBV replication through the hydroxylase activity of JMJD5. IMPORTANCE HBx protein encoded by hepatitis B virus (HBV) plays important roles in pathogenesis and replication of HBV. We identified jumonji C-domain-containing 5 (JMJD5) as a novel binding partner to HBx. JMJD5 was shown to regulate several transcriptional factors to maintain hepatocyte function. Although HBx had been shown to support HBV replication, deficiency of JMJD5 abolished contribution of HBx in HBV replication, suggesting that HBx-mediated HBV replication is largely dependent on JMJD5. We showed that hydroxylase activity of JMJD5 in the C terminus region is crucial for expression of HNF4A and replication of HBV. Furthermore, a mutant JMJD5 with Gly(135) replaced by Glu failed to interact with HBx and to rescue the replication of HBV in JMJD5-knockout cells. Taken together, our data suggest that interaction of JMJD5 with HBx facilitates HBV replication through the hydroxylase activity of JMJD5.
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Teng YC, Shen ZQ, Kao CH, Tsai TF. Hepatocellular carcinoma mouse models: Hepatitis B virus-associated hepatocarcinogenesis and haploinsufficient tumor suppressor genes. World J Gastroenterol 2016; 22:300-325. [PMID: 26755878 PMCID: PMC4698494 DOI: 10.3748/wjg.v22.i1.300] [Citation(s) in RCA: 19] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/18/2015] [Revised: 10/14/2015] [Accepted: 11/24/2015] [Indexed: 02/06/2023] Open
Abstract
The multifactorial and multistage pathogenesis of hepatocellular carcinoma (HCC) has fascinated a wide spectrum of scientists for decades. While a number of major risk factors have been identified, their mechanistic roles in hepatocarcinogenesis still need to be elucidated. Many tumor suppressor genes (TSGs) have been identified as being involved in HCC. These TSGs can be classified into two groups depending on the situation with respect to allelic mutation/loss in the tumors: the recessive TSGs with two required mutated alleles and the haploinsufficient TSGs with one required mutated allele. Hepatitis B virus (HBV) is one of the most important risk factors associated with HCC. Although mice cannot be infected with HBV due to the narrow host range of HBV and the lack of a proper receptor, one advantage of mouse models for HBV/HCC research is the numerous and powerful genetic tools that help investigate the phenotypic effects of viral proteins and allow the dissection of the dose-dependent action of TSGs. Here, we mainly focus on the application of mouse models in relation to HBV-associated HCC and on TSGs that act either in a recessive or in a haploinsufficient manner. Discoveries obtained using mouse models will have a great impact on HCC translational medicine.
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Takaki A, Yamamoto K. Control of oxidative stress in hepatocellular carcinoma: Helpful or harmful? World J Hepatol 2015; 7:968-979. [PMID: 25954479 PMCID: PMC4419100 DOI: 10.4254/wjh.v7.i7.968] [Citation(s) in RCA: 73] [Impact Index Per Article: 7.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/28/2014] [Revised: 12/03/2014] [Accepted: 02/02/2015] [Indexed: 02/06/2023] Open
Abstract
Oxidative stress is becoming recognized as a key factor in the progression of chronic liver disease (CLD) and hepatocarcinogenesis. The metabolically important liver is a major reservoir of mitochondria that serve as sources of reactive oxygen species, which are apparently responsible for the initiation of necroinflammation. As a result, CLD could be a major inducer of oxidative stress. Chronic hepatitis C is a powerful generator of oxidative stress, causing a high rate of hepatocarcinogenesis among patients with cirrhosis. Non-alcoholic steatohepatitis is also associated with oxidative stress although its hepatocarcinogenic potential is lower than that of chronic hepatitis C. Analyses of serum markers and histological findings have shown that hepatocellular carcinoma correlates with oxidative stress and experimental data indicate that oxidative stress increases the likelihood of developing hepatocarcinogenesis. However, the results of antioxidant therapy have not been favorable. Physiological oxidative stress is a necessary biological response, and thus adequate control of oxidative stress and a balance between oxidative and anti-oxidative responses is important. Several agents including metformin and L-carnitine can reportedly control mechanistic oxidative stress. This study reviews the importance of oxidative stress in hepatocarcinogenesis and of control strategies for the optimal survival of patients with CLD and hepatocellular carcinoma.
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Li W, Goto K, Matsubara Y, Ito S, Muroyama R, Li Q, Kato N. The characteristic changes in hepatitis B virus x region for hepatocellular carcinoma: a comprehensive analysis based on global data. PLoS One 2015; 10:e0125555. [PMID: 25942596 PMCID: PMC4420286 DOI: 10.1371/journal.pone.0125555] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/31/2014] [Accepted: 03/25/2015] [Indexed: 12/12/2022] Open
Abstract
OBJECTIVES Mutations in hepatitis B virus (HBV) X region (HBx) play important roles in hepatocarcinogenesis while the results remain controversial. We sought to clarify potential hepatocellular carcinoma (HCC)-characteristic mutations in HBx from HBV genotype C-infected patients and the distribution of those mutations in different disease phases and genotypes. METHODS HBx sequences downloaded from an online global HBV database were screened and then classified into Non-HCC or HCC group by diagnosis information. Patients' data of patient age, gender, country or area, and viral genotype were also extracted. Logistic regression was performed to evaluate the effects of mutations on HCC risk. RESULTS 1) Full length HBx sequences (HCC: 161; Non-HCC: 954) originated from 1115 human sera across 29 countries/areas were extracted from the downloaded 5956 HBx sequences. Genotype C occupied 40.6% of Non-HCC (387/954) and 89.4% of HCC (144/161). 2) Sixteen nucleotide positions showed significantly different distributions between genotype C HCC and Non-HCC groups. 3) Logistic regression showed that mutations A1383C (OR: 2.32, 95% CI: 1.34-4.01), R1479C/T (OR: 1.96, 95% CI: 1.05-3.64; OR: 5.15, 95% CI: 2.53-10.48), C1485T (OR: 2.40, 95% CI: 1.41-4.08), C1631T (OR: 4.09, 95% CI: 1.41-11.85), C1653T (OR: 2.58, 95% CI: 1.59-4.19), G1719T (OR: 2.11, 95% CI: 1.19-3.73), and T1800C (OR: 23.59, 95% CI: 2.25-247.65) were independent risk factors for genotype C HBV-related HCC, presenting different trends among individual disease phases. 4) Several genotype C HCC risk mutations pre-existed, even as major types, in early disease phases with other genotypes. CONCLUSIONS Mutations associated with HCC risk were mainly located in HBx transactivation domain, viral promoter, protein/miRNA binding sites, and the area for immune epitopes. Furthermore, the signatures of these mutations were unique to disease phases leading to HCC, suggesting molecular counteractions between the virus and host during hepatocarcinogenesis.
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Affiliation(s)
- Wenwen Li
- Division of advanced genome medicine, Advanced clinical research center, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan
| | - Kaku Goto
- Division of advanced genome medicine, Advanced clinical research center, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan
| | - Yasuo Matsubara
- Division of advanced genome medicine, Advanced clinical research center, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan
| | - Sayaka Ito
- Division of advanced genome medicine, Advanced clinical research center, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan
| | - Ryosuke Muroyama
- Division of advanced genome medicine, Advanced clinical research center, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan
| | - Qiang Li
- Affiliated Infectious Disease Hospital of Shandong University, Jinan, Shandong, China
| | - Naoya Kato
- Division of advanced genome medicine, Advanced clinical research center, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan
- * E-mail:
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Xie Y, Liu S, Zhao Y, Guo Z, Xu J. X protein mutations in hepatitis B virus DNA predict postoperative survival in hepatocellular carcinoma. Tumour Biol 2014; 35:10325-31. [PMID: 25034530 DOI: 10.1007/s13277-014-2331-0] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/21/2014] [Accepted: 07/08/2014] [Indexed: 12/13/2022] Open
Abstract
Hepatitis B virus (HBV) DNA is prone to mutations because of the proofreading deficiencies of HBV polymerase. The postoperative prognostic value of HBV mutations in HBV X protein (HBx) gene was assessed in HBV associated hepatocellular carcinoma (HCC) patients. The HBx gene was amplified and sequenced, the HBV mutations was identified according to NCBI database ( http://www.ncbi.nlm.nih.gov/genome/5536 ). The relationship between the HBV mutations and HCC survival was compared. Survival curves were generated using the Kaplan-Meier method, and comparisons between the curves were made using the log-rank test. Multivariate survival analysis was performed using a Cox proportional hazards model. After adjusting for clinical characteristics, the following eight mutational sites were identified as statistically significant independent predictors of HCC survival: 1383, 1461, 1485, 1544, 1613, 1653, 1719, and 1753. In addition, the following four mutational sites were identified for their association with survival at a border-line significance level: 1527, 1637, 1674, and 1762/1764. A total of 12 mutations in HBx gene region were identified as independent predictors of postoperative survival in HCC patients. The analysis of HBV DNA mutations may help identify patient subgroups with poor prognosis and may help refine therapeutic decisions regarding HCC patients.
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Affiliation(s)
- Ying Xie
- Hebei Key Lab of Laboratory Animal Science, Hebei Medical University, Shijiazhuang, People's Republic of China
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Diogo J, Bratanich A. The nematode Caenorhabditis elegans as a model to study viruses. Arch Virol 2014; 159:2843-51. [PMID: 25000902 DOI: 10.1007/s00705-014-2168-2] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/28/2014] [Accepted: 06/28/2014] [Indexed: 12/15/2022]
Abstract
Caenorhabditis elegans is a worm that has been extensively studied, and it is today an accepted model in many different biological fields. C. elegans is cheap to maintain, it is transparent, allowing easy localization studies, and it develops from egg to adult in around 4 days. Many mutants, available to the scientific community, have been developed. This has facilitated the study of the role of particular genes in many cellular pathways, which are highly conserved when compared with higher eukaryotes. This review describes the advantages of C. elegans as a laboratory model and the known mechanisms utilized by this worm to fight pathogens. In particular, we describe the strong C. elegans RNAi machinery, which plays an important role in the antiviral response. This has been shown in vitro (C. elegans cell cultures) as well as in vivo (RNAi-deficient strains) utilizing recently described viruses that have the worm as a host. Infections with mammalian viruses have also been achieved using chemical treatment. The role of viral genes involved in pathogenesis has been addressed by evaluating the phenotypes of transgenic strains of C. elegans expressing those genes. Very simple approaches such as feeding the worm with bacteria transformed with viral genes have also been utilized. The advantages and limitations of different approaches are discussed.
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Affiliation(s)
- Jesica Diogo
- Department of Virology, School of Veterinary Sciences, University of Buenos Aires, Av. Chorroarin 280, 1427, Buenos Aires, Argentina
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Kusunoki H, Tanaka T, Kohno T, Wakamatsu K, Hamaguchi I. Structural characterization of the BH3-like motif of hepatitis B virus X protein. Biochem Biophys Res Commun 2014; 450:741-5. [PMID: 24950407 DOI: 10.1016/j.bbrc.2014.06.042] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/28/2014] [Accepted: 06/10/2014] [Indexed: 01/26/2023]
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Xu C, Zhou W, Wang Y, Qiao L. Hepatitis B virus-induced hepatocellular carcinoma. Cancer Lett 2014; 345:216-222. [PMID: 23981576 DOI: 10.1016/j.canlet.2013.08.035] [Citation(s) in RCA: 107] [Impact Index Per Article: 9.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/14/2013] [Revised: 08/10/2013] [Accepted: 08/18/2013] [Indexed: 12/12/2022]
Abstract
Many factors are considered to contribute to hepatitis B virus (HBV) associated hepatocellular carcinoma (HCC), including products of HBV, HBV integration and mutation, and host susceptibility. HBV X protein (HBx) can interfere with several signal pathways that associated with cell proliferation and apoptosis, and the impact of HBx C-terminal truncation in the development of HCC has been implicated. Recent studies by advanced sequencing technologies have revealed recurrent HBV DNA integration sites in hepatoma cells and susceptible genes/SNPs play an important role in the pathogenesis of liver cancer. Epigenetic changes, immune and inflammatory factors are also important contributing factors for liver cancer. This mini-review provides an overview on the recent development of HBV induced HCC.
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Affiliation(s)
- Cheng Xu
- Institute for Infectious Diseases, Southwest Hospital, Third Military Medical University, Chongqing 400038, China
| | - Wence Zhou
- The Department of General Surgery II, The First Hospital of Lanzhou University, Lanzhou 730000, Gansu Province, China
| | - Yuming Wang
- Institute for Infectious Diseases, Southwest Hospital, Third Military Medical University, Chongqing 400038, China.
| | - Liang Qiao
- Storr Liver Unit, University of Sydney, Westmead Millennium Institute, Westmead Hospital, Westmead, NSW 2145, Australia.
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Chung YL. Defective DNA damage response and repair in liver cells expressing hepatitis B virus surface antigen. FASEB J 2013; 27:2316-27. [PMID: 23444429 DOI: 10.1096/fj.12-226639] [Citation(s) in RCA: 16] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/26/2022]
Abstract
Hepatitis B virus (HBV) is implicated in liver cancer. The aim of this study was to find out whether HBV or its components [HBV surface antigen (HBsAg), HBV core protein (HBc), and HBV X protein (HBx)] could interfere with the host DNA damage response and repair pathway. The full HBV genome or individual HBV open-reading frame (ORF) was introduced into HepG2 cells to examine the effect on host genomic stability, DNA repair efficacy in response to double-strand DNA damage, and DNA damage-induced cell death. Responses to apoptosis induction in the HBV ORF-transfected HepG2 cells were also compared with those in HBV-positive and HBV-negative human hepatocellular carcinoma (HCC) cells. In the absence of HBV replication, accumulation of HBsAg in liver cells without other HBV proteins enhanced DNA repair protein and tumor suppressor promyelocytic leukemia (PML) degradation, which resulted in resistance to apoptosis induction and deficient double-strand DNA repair. However, HBsAg-positive cells exhibited increased cell death with exposure to the poly(ADP-ribose) polymerase inhibitor that blocks single-strand DNA repair. These results indicate that suppression of PML by HBsAg disrupts cellular mechanisms that respond to double-strand DNA damage for DNA repair or apoptosis induction, which may facilitate hepatocarcinogenesis and open up a synthetic lethality strategy for HBsAg-positive HCC treatment.
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Affiliation(s)
- Yih-Lin Chung
- Department of Radiation Oncology, Koo Foundation Sun Yat-Sen Cancer Center, Taipei, Taiwan.
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