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Reyes Ballista JM, Hoover AJ, Noble JT, Acciani MD, Miazgowicz KL, Harrison SA, Tabscott GAL, Duncan A, Barnes DN, Jimenez AR, Brindley MA. Chikungunya virus release is reduced by TIM-1 receptors through binding of envelope phosphatidylserine. J Virol 2024; 98:e0077524. [PMID: 39007616 PMCID: PMC11334481 DOI: 10.1128/jvi.00775-24] [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: 04/30/2024] [Accepted: 06/11/2024] [Indexed: 07/16/2024] Open
Abstract
T-cell immunoglobin and mucin domain protein-1 (TIM-1) mediates entry of chikungunya virus (CHIKV) into some mammalian cells through the interaction with envelope phospholipids. While this interaction enhances entry, TIM-1 has been shown to tether newly formed HIV and Ebola virus particles, limiting their efficient release. In this study, we investigate the ability of surface receptors such as TIM-1 to sequester newly budded virions on the surface of infected cells. We established a luminescence reporter system to produce chikungunya viral particles that integrate nano-luciferase and easily quantify viral particles. We found that TIM-1 on the surface of host cells significantly reduced CHIKV release efficiency in comparison to other entry factors. Removal of cell surface TIM-1 through direct cellular knock-out or altering the cellular lipid distribution enhanced CHIKV release. Over the course of infection, CHIKV was able to counteract the tethering effect by gradually decreasing the surface levels of TIM-1 in a process mediated by the nonstructural protein 2. This study highlights the importance of phosphatidylserine receptors in mediating not only the entry of CHIKV but also its release and could aid in developing cell lines capable of enhanced vaccine production. IMPORTANCE Chikungunya virus (CHIKV) is an enveloped alphavirus transmitted by the bites of infectious mosquitoes. Infection with CHIKV results in the development of fever, joint pain, and arthralgia that can become chronic and last for months after infection. Prevention of this disease is still highly focused on vector control strategies. In December 2023, a new live attenuated vaccine against CHIKV was approved by the FDA. We aimed to study the cellular factors involved in CHIKV release, to better understand CHIKV's ability to efficiently infect and spread among a wide variety of cell lines. We found that TIM-1 receptors can significantly abrogate CHIKV's ability to efficiently exit infected cells. This information can be beneficial for maximizing viral particle production in laboratory settings and during vaccine manufacturing.
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Affiliation(s)
- Judith M. Reyes Ballista
- Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, Georgia, USA
| | - Ashley J. Hoover
- Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, Georgia, USA
| | - Joseph T. Noble
- Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, Georgia, USA
| | - Marissa D. Acciani
- Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, Georgia, USA
| | - Kerri L. Miazgowicz
- Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, Georgia, USA
| | - Sarah A. Harrison
- Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, Georgia, USA
| | - Grace Andrea L. Tabscott
- Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, Georgia, USA
| | - Avery Duncan
- Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, Georgia, USA
| | - Don N. Barnes
- Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, Georgia, USA
| | - Ariana R. Jimenez
- Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, Georgia, USA
| | - Melinda A. Brindley
- Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, Georgia, USA
- Department of Population Health, College of Veterinary Medicine, University of Georgia, Athens, Georgia, USA
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Ballista JMR, Hoover AJ, Noble JT, Acciani MD, Miazgowicz KL, Harrison SA, Tabscott GAL, Duncan A, Barnes DN, Jimenez AR, Brindley MA. Chikungunya Virus Release is Reduced by TIM-1 Receptors Through Binding of Envelope Phosphatidylserine. BIORXIV : THE PREPRINT SERVER FOR BIOLOGY 2024:2024.01.25.577233. [PMID: 38328121 PMCID: PMC10849729 DOI: 10.1101/2024.01.25.577233] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 02/09/2024]
Abstract
T-cell immunoglobin and mucin domain protein-1 (TIM-1) mediates entry of Chikungunya virus (CHIKV) into some mammalian cells through the interaction with envelope phospholipids. While this interaction enhances entry, TIM has been shown to tether newly formed HIV and Ebola virus particles, limiting their efficient release. In this study, we investigate the ability of surface receptors such as TIM-1 to sequester newly budded virions on the surface of infected cells. We established a luminescence reporter system to produce Chikungunya viral particles that integrate nano-luciferase and easily quantify viral particles. We found that TIM-1 on the surface of host cells significantly reduced CHIKV release efficiency in comparison to other entry factors. Removal of cell surface TIM-1 through direct cellular knock-out or altering the cellular lipid distribution enhanced CHIKV release. Over the course of infection, CHIKV was able to counteract the tethering effect by gradually decreasing the surface levels of TIM-1 in a process that appears to be mediated by the nonstructural protein 2. This study highlights the importance of phosphatidylserine receptors in mediating not only the entry of CHIKV but also its release and could aid in developing cell lines capable of enhanced vaccine production.
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Affiliation(s)
- Judith M. Reyes Ballista
- Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, GA, USA
| | - Ashley J. Hoover
- Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, GA, USA
| | - Joseph T. Noble
- Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, GA, USA
| | - Marissa D. Acciani
- Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, GA, USA
| | - Kerri L. Miazgowicz
- Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, GA, USA
| | - Sarah A. Harrison
- Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, GA, USA
| | - Grace Andrea L. Tabscott
- Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, GA, USA
| | - Avery Duncan
- Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, GA, USA
| | - Don N. Barnes
- Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, GA, USA
| | - Ariana R. Jimenez
- Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, GA, USA
| | - Melinda A. Brindley
- Department of Infectious Diseases, College of Veterinary Medicine, University of Georgia, Athens, GA, USA
- Department of Population Health, College of Veterinary Medicine, University of Georgia, Athens, GA, USA
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Urban S, Neumann-Haefelin C, Lampertico P. Hepatitis D virus in 2021: virology, immunology and new treatment approaches for a difficult-to-treat disease. Gut 2021; 70:1782-1794. [PMID: 34103404 PMCID: PMC8355886 DOI: 10.1136/gutjnl-2020-323888] [Citation(s) in RCA: 126] [Impact Index Per Article: 31.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 04/12/2021] [Accepted: 05/26/2021] [Indexed: 02/06/2023]
Abstract
Approximately 5% of individuals infected with hepatitis B virus (HBV) are coinfected with hepatitis D virus (HDV). Chronic HBV/HDV coinfection is associated with an unfavourable outcome, with many patients developing liver cirrhosis, liver failure and eventually hepatocellular carcinoma within 5-10 years. The identification of the HBV/HDV receptor and the development of novel in vitro and animal infection models allowed a more detailed study of the HDV life cycle in recent years, facilitating the development of specific antiviral drugs. The characterisation of HDV-specific CD4+ and CD8+T cell epitopes in untreated and treated patients also permitted a more precise understanding of HDV immunobiology and possibly paves the way for immunotherapeutic strategies to support upcoming specific therapies targeting viral or host factors. Pegylated interferon-α has been used for treating HDV patients for the last 30 years with only limited sustained responses. Here we describe novel treatment options with regard to their mode of action and their clinical effectiveness. Of those, the entry-inhibitor bulevirtide (formerly known as myrcludex B) received conditional marketing authorisation in the European Union (EU) in 2020 (Hepcludex). One additional drug, the prenylation inhibitor lonafarnib, is currently under investigation in phase III clinical trials. Other treatment strategies aim at targeting hepatitis B surface antigen, including the nucleic acid polymer REP2139Ca. These recent advances in HDV virology, immunology and treatment are important steps to make HDV a less difficult-to-treat virus and will be discussed.
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Affiliation(s)
- Stephan Urban
- Department of Infectious Diseases, Molecular Virology, University Hospital Heidelberg, Heidelberg, Germany,German Center for Infection Research (DZIF) - Heidelberg Partner Site, Heidelberg, Germany
| | - Christoph Neumann-Haefelin
- Department of Medicine II, Freiburg University Medical Center, Faculty of Medicine, University of Freiburg, Freiburg, Germany
| | - Pietro Lampertico
- Division of Gastroenterology and Hepatology, Fondazione IRCCS Ca' Granda Ospedale Maggiore Policlinico, Milan, Italy,CRC “A. M. and A. Migliavacca” Center for Liver Disease, Department of Pathophysiology and Transplantation, University of Milan, Milan, Italy
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Bastolla U. Mathematical Model of SARS-Cov-2 Propagation Versus ACE2 Fits COVID-19 Lethality Across Age and Sex and Predicts That of SARS. Front Mol Biosci 2021; 8:706122. [PMID: 34322518 PMCID: PMC8311794 DOI: 10.3389/fmolb.2021.706122] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/06/2021] [Accepted: 06/30/2021] [Indexed: 12/11/2022] Open
Abstract
The fatality rate of Covid-19 escalates with age and is larger in men than women. I show that these variations correlate strongly with the level of the viral receptor protein ACE2 in rat lungs, which is consistent with the still limited data on human ACE2. Surprisingly, lower receptor levels correlate with higher fatality. I propose two possible explanations of this negative correlation: First, a previous mathematical model predicts that the velocity of viral progression in the organism as a function of the receptor level has a maximum and declines for abundant receptor. Secondly, degradation of ACE2 by the virus may cause the runaway inflammatory response that characterizes severe CoViD-19. I present here a mathematical model that predicts the lethality as a function of ACE2 protein level based on the two above hypothesis. The model fits Covid-19 fatality rate across age and sex in three countries with high accuracy (r 2 > 0.9 ) under the hypothesis that the speed of viral progression in the infected organism is a decreasing function of the ACE2 level. Moreover, rescaling the fitted parameters by the ratio of the binding rates of the spike proteins of SARS-CoV and SARS-CoV-2 allows predicting the fatality rate of SARS-CoV across age and sex, thus linking the molecular and epidemiological levels.
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Affiliation(s)
- Ugo Bastolla
- Centro de Biologia Molecular “Severo Ochoa”, CSIC-UAM Cantoblanco, Madrid, Spain
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Lempp FA, Schlund F, Rieble L, Nussbaum L, Link C, Zhang Z, Ni Y, Urban S. Recapitulation of HDV infection in a fully permissive hepatoma cell line allows efficient drug evaluation. Nat Commun 2019; 10:2265. [PMID: 31118422 PMCID: PMC6531471 DOI: 10.1038/s41467-019-10211-2] [Citation(s) in RCA: 38] [Impact Index Per Article: 6.3] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/27/2018] [Accepted: 04/22/2019] [Indexed: 12/18/2022] Open
Abstract
Hepatitis delta virus (HDV) depends on the helper function of hepatitis B virus (HBV), which provides the envelope proteins for progeny virus secretion. Current infection-competent cell culture models do not support assembly and secretion of HDV. By stably transducing HepG2 cells with genes encoding the NTCP-receptor and the HBV envelope proteins we produce a cell line (HepNB2.7) that allows continuous secretion of infectious progeny HDV following primary infection. Evaluation of antiviral drugs shows that the entry inhibitor Myrcludex B (IC50: 1.4 nM) and interferon-α (IC50: 28 IU/ml, but max. 60–80% inhibition) interfere with primary infection. Lonafarnib inhibits virus secretion (IC50: 36 nM) but leads to a substantial intracellular accumulation of large hepatitis delta antigen and replicative intermediates, accompanied by the induction of innate immune responses. This work provides a cell line that supports the complete HDV replication cycle and presents a convenient tool for antiviral drug evaluation. Hepatitis delta virus (HDV) depends on the envelope proteins of hepatitis B virus (HBV) for virion production. Here, Lempp et al. produce a cell line expressing HBV envelope proteins and their receptor, which allows continuous secretion of infectious progeny HDV and testing of antiviral drugs.
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Affiliation(s)
- Florian A Lempp
- Department of Infectious Diseases, Molecular Virology, University Hospital Heidelberg, Heidelberg, 69120, Germany.,German Centre for Infection Research (DZIF), partner site Heidelberg, Heidelberg, 69120, Germany
| | - Franziska Schlund
- Department of Infectious Diseases, Molecular Virology, University Hospital Heidelberg, Heidelberg, 69120, Germany
| | - Lisa Rieble
- Department of Infectious Diseases, Molecular Virology, University Hospital Heidelberg, Heidelberg, 69120, Germany
| | - Lea Nussbaum
- Department of Infectious Diseases, Molecular Virology, University Hospital Heidelberg, Heidelberg, 69120, Germany
| | - Corinna Link
- Department of Infectious Diseases, Molecular Virology, University Hospital Heidelberg, Heidelberg, 69120, Germany
| | - Zhenfeng Zhang
- Department of Infectious Diseases, Molecular Virology, University Hospital Heidelberg, Heidelberg, 69120, Germany
| | - Yi Ni
- Department of Infectious Diseases, Molecular Virology, University Hospital Heidelberg, Heidelberg, 69120, Germany.,German Centre for Infection Research (DZIF), partner site Heidelberg, Heidelberg, 69120, Germany
| | - Stephan Urban
- Department of Infectious Diseases, Molecular Virology, University Hospital Heidelberg, Heidelberg, 69120, Germany. .,German Centre for Infection Research (DZIF), partner site Heidelberg, Heidelberg, 69120, Germany.
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Aliyu IA, Ling KH, Md Hashim NF, Lam JY, Chee HY. Annexin II as a Dengue Virus Serotype 2 Interacting Protein Mediating Virus Interaction on Vero Cells. Viruses 2019; 11:v11040335. [PMID: 30970587 PMCID: PMC6520844 DOI: 10.3390/v11040335] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/31/2019] [Accepted: 02/21/2019] [Indexed: 01/01/2023] Open
Abstract
Recent evidence has demonstrated that dengue virus requires active filopodia formation for a successful infection. However, the cellular factor involved in the interaction has not been fully elucidated. We used a combination of virus overlay protein binding assay and LC-MS/MS, and identified annexin II as a dengue virus serotype 2 (DENV2) interacting protein on Vero cells, upon filopodia induction. Flow cytometry analysis showed annexin II on the Vero cells surface increased when DENV2 was added. The amount of annexin II in the plasma membrane fraction was reduced as the infection progressed. Antibody-mediated inhibition of infection and siRNA-mediated knockdown of annexin II expression significantly reduced DENV2 infection and production levels. Collectively, we demonstrated that annexin II is one of the host factor involved in DENV2 binding on Vero cells.
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Affiliation(s)
- Isah Abubakar Aliyu
- Department of Medical Microbiology and Parasitology, Faculty of Medicine and Health Science, Universiti Putra Malaysia, 43400 UPM Serdang, Selangor, Malaysia.
- Department of Medical Laboratory Science, Faculty of Allied Health Science, College of Health Science, Bayero University Kano, PMB 3011 Kano State, Nigeria.
| | - King-Hwa Ling
- NeuroBiology & Genetics Group, Department of Biomedical Sciences, Faculty of Medicine and Health Sciences, 43400 UPM Serdang, Selangor, Malaysia.
- Genetics and Regenerative Medicine Research Centre, Faculty of Medicine and Health Sciences, 43400 UPM Serdang, Selangor, Malaysia.
| | - Nur Fariesha Md Hashim
- Department of Biomedical Sciences, Faculty of Medicine and Health Sciences, 43400 UPM Serdang, Selangor, Malaysia.
| | - Jia-Yong Lam
- Department of Medical Microbiology and Parasitology, Faculty of Medicine and Health Science, Universiti Putra Malaysia, 43400 UPM Serdang, Selangor, Malaysia.
| | - Hui-Yee Chee
- Department of Medical Microbiology and Parasitology, Faculty of Medicine and Health Science, Universiti Putra Malaysia, 43400 UPM Serdang, Selangor, Malaysia.
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Abstract
Coinfections involving viruses are being recognized to influence the disease pattern that occurs relative to that with single infection. Classically, we usually think of a clinical syndrome as the consequence of infection by a single virus that is isolated from clinical specimens. However, this biased laboratory approach omits detection of additional agents that could be contributing to the clinical outcome, including novel agents not usually considered pathogens. The presence of an additional agent may also interfere with the targeted isolation of a known virus. Viral interference, a phenomenon where one virus competitively suppresses replication of other coinfecting viruses, is the most common outcome of viral coinfections. In addition, coinfections can modulate virus virulence and cell death, thereby altering disease severity and epidemiology. Immunity to primary virus infection can also modulate immune responses to subsequent secondary infections. In this review, various virological mechanisms that determine viral persistence/exclusion during coinfections are discussed, and insights into the isolation/detection of multiple viruses are provided. We also discuss features of heterologous infections that impact the pattern of immune responsiveness that develops.
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Chiu ES, Hoover EA, VandeWoude S. A Retrospective Examination of Feline Leukemia Subgroup Characterization: Viral Interference Assays to Deep Sequencing. Viruses 2018; 10:E29. [PMID: 29320424 PMCID: PMC5795442 DOI: 10.3390/v10010029] [Citation(s) in RCA: 30] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/01/2017] [Revised: 01/03/2018] [Accepted: 01/08/2018] [Indexed: 01/10/2023] Open
Abstract
Feline leukemia virus (FeLV) was the first feline retrovirus discovered, and is associated with multiple fatal disease syndromes in cats, including lymphoma. The original research conducted on FeLV employed classical virological techniques. As methods have evolved to allow FeLV genetic characterization, investigators have continued to unravel the molecular pathology associated with this fascinating agent. In this review, we discuss how FeLV classification, transmission, and disease-inducing potential have been defined sequentially by viral interference assays, Sanger sequencing, PCR, and next-generation sequencing. In particular, we highlight the influences of endogenous FeLV and host genetics that represent FeLV research opportunities on the near horizon.
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Affiliation(s)
- Elliott S Chiu
- Department of Microbiology, Immunology, and Pathology, Colorado State University, Fort Collins, CO 80524, USA.
| | - Edward A Hoover
- Department of Microbiology, Immunology, and Pathology, Colorado State University, Fort Collins, CO 80524, USA.
| | - Sue VandeWoude
- Department of Microbiology, Immunology, and Pathology, Colorado State University, Fort Collins, CO 80524, USA.
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Expression of Oncogenic Alleles Induces Multiple Blocks to Human Cytomegalovirus Infection. J Virol 2016; 90:4346-4356. [PMID: 26889030 DOI: 10.1128/jvi.00179-16] [Citation(s) in RCA: 23] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/29/2016] [Accepted: 02/08/2016] [Indexed: 12/26/2022] Open
Abstract
UNLABELLED In contrast to many viruses, human cytomegalovirus (HCMV) is unable to productively infect most cancer-derived cell lines. The mechanisms of this restriction are unclear. To explore this issue, we tested whether defined oncogenic alleles, including the simian virus 40 (SV40) T antigen (TAg) and oncogenic H-Ras, inhibit HCMV infection. We found that expression of SV40 TAg blocks HCMV infection in human fibroblasts, whereas the replication of a related herpesvirus, herpes simplex virus 1 (HSV-1), was not impacted. The earliest restriction of HCMV infection involves a block of viral entry, as TAg expression prevented the nuclear delivery of viral DNA and pp65. Subsequently, we found that TAg expression reduces the abundance of platelet-derived growth factor receptor α (PDGFRα), a host protein important for HCMV entry. Viral entry into TAg-immortalized fibroblasts could largely be rescued by PDGFRα overexpression. Similarly, PDGFRα overexpression in HeLa cells markedly increased the levels of HCMV gene expression and DNA replication. However, the robust production of viral progeny was not restored by PDGFRα overexpression in either HeLa cells or TAg-immortalized fibroblasts, suggesting additional restrictions associated with transformation and TAg expression. In TAg-expressing fibroblasts, expression of the immediate early 2 (IE2) protein was not rescued to the same extent as that of the immediate early 1 (IE1) protein, suggesting that TAg expression impacts the accumulation of major immediate early (MIE) transcripts. Transduction of IE2 largely rescued HCMV gene expression in TAg-expressing fibroblasts but did not rescue the production of infectious virions. Collectively, our data indicate that oncogenic alleles induce multiple restrictions to HCMV replication. IMPORTANCE HCMV cannot replicate in most cancerous cells, yet the causes of this restriction are not clear. The mechanisms that restrict viral replication in cancerous cells represent viral vulnerabilities that can potentially be exploited therapeutically in other contexts. Here we found that SV40 T antigen-mediated transformation inhibits HCMV infection at multiple points in the viral life cycle, including through inhibition of proper viral entry, normal expression of immediate early genes, and viral DNA replication. Our results suggest that the SV40 T antigen could be a valuable tool to dissect cellular activities that are important for successful infection, thereby potentially informing novel antiviral development strategies. This is an important consideration, given that HCMV is a leading cause of birth defects and causes severe infection in immunocompromised individuals.
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Li J, Tong S. From DCPD to NTCP: the long journey towards identifying a functional hepatitis B virus receptor. Clin Mol Hepatol 2015; 21:193-9. [PMID: 26523264 PMCID: PMC4612279 DOI: 10.3350/cmh.2015.21.3.193] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 05/05/2015] [Accepted: 08/15/2015] [Indexed: 12/13/2022] Open
Abstract
Hepatitis B virus (HBV) is the prototype of hepatotropic DNA viruses (hepadnaviruses) infecting a wide range of human and non-human hosts. Previous studies with duck hepatitis B virus (DHBV) identified duck carboxypeptidase D (dCPD) as a host specific binding partner for full-length large envelope protein, and p120 as a binding partner for several truncated versions of the large envelope protein. p120 is the P protein of duck glycine decarboxylase (dGLDC) with restricted expression in DHBV infectible tissues. Several lines of evidence suggest the importance of dCPD, and especially p120, in productive DHBV infection, although neither dCPD nor p120 cDNA could confer susceptibility to DHBV infection in any cell line. Recently, sodium taurocholate cotransporting polypeptide (NTCP) has been identified as a binding partner for the N-terminus of HBV large envelope protein. Importantly, knock down and reconstitution experiments unequivocally demonstrated that NTCP is both necessary and sufficient for in vitro infection by HBV and hepatitis delta virus (HDV), an RNA virus using HBV envelope proteins for its transmission. What remains unclear is whether NTCP is the major HBV receptor in vivo. The fact that some HBV patients are homozygous with an NTCP mutation known to abolish its receptor function suggests the existence of NTCP-independent pathways of HBV entry. Also, NTCP very likely mediates just one step of the HBV entry process, with additional co-factors for productive HBV infection still to be discovered. NTCP offers a novel therapeutic target for the control of chronic HBV infection.
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Affiliation(s)
- Jisu Li
- Liver Research Center, Rhode Island Hospital, The Warren Alpert Medical School of Brown University, Providence, USA
| | - Shuping Tong
- Liver Research Center, Rhode Island Hospital, The Warren Alpert Medical School of Brown University, Providence, USA. ; Key lab of Medical Molecular Virology, School of Basic Medical Sciences, Fudan University, Shanghai, China
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11
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Differential Ability of Primary HIV-1 Nef Isolates To Downregulate HIV-1 Entry Receptors. J Virol 2015; 89:9639-52. [PMID: 26178998 DOI: 10.1128/jvi.01548-15] [Citation(s) in RCA: 23] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/17/2015] [Accepted: 07/06/2015] [Indexed: 01/02/2023] Open
Abstract
UNLABELLED HIV-1 Nef downregulates the viral entry receptor CD4 as well as the coreceptors CCR5 and CXCR4 from the surface of HIV-infected cells, and this leads to promotion of viral replication through superinfection resistance and other mechanisms. Nef sequence motifs that modulate these functions have been identified via in vitro mutagenesis with laboratory HIV-1 strains. However, it remains unclear whether the same motifs contribute to Nef activity in patient-derived sequences and whether these motifs may differ in Nef sequences isolated at different infection stages and/or from patients with different disease phenotypes. Here, nef clones from 45 elite controllers (EC), 46 chronic progressors (CP), and 43 acute progressors (AP) were examined for their CD4, CCR5, and CXCR4 downregulation functions. Nef clones from EC exhibited statistically significantly impaired CD4 and CCR5 downregulation ability and modestly impaired CXCR4 downregulation activity compared to those from CP and AP. Nef's ability to downregulate CD4 and CCR5 correlated positively in all cohorts, suggesting that they are functionally linked in vivo. Moreover, impairments in Nef's receptor downregulation functions increased the susceptibility of Nef-expressing cells to HIV-1 infection. Mutagenesis studies on three functionally impaired EC Nef clones revealed that multiple residues, including those at novel sites, were involved in the alteration of Nef functions and steady-state protein levels. Specifically, polymorphisms at highly conserved tryptophan residues (e.g., Trp-57 and Trp-183) and immune escape-associated sites were responsible for reduced Nef functions in these clones. Our results suggest that the functional modulation of primary Nef sequences is mediated by complex polymorphism networks. IMPORTANCE HIV-1 Nef, a key factor for viral pathogenesis, downregulates functionally important molecules from the surface of infected cells, including the viral entry receptor CD4 and coreceptors CCR5 and CXCR4. This activity enhances viral replication by protecting infected cells from cytotoxicity associated with superinfection and may also serve as an immune evasion strategy. However, how these activities are maintained under selective pressure in vivo remains elusive. We addressed this question by analyzing functions of primary Nef clones isolated from patients at various infection stages and with different disease phenotypes, including elite controllers, who spontaneously control HIV-1 viremia to undetectable levels. The results indicated that downregulation of HIV-1 entry receptors, particularly CCR5, is impaired in Nef clones from elite controllers. These functional impairments were driven by rare Nef polymorphisms and adaptations associated with cellular immune responses, underscoring the complex molecular pathways responsible for maintaining and attenuating viral protein function in vivo.
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12
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A novel human endogenous retroviral protein inhibits cell-cell fusion. Sci Rep 2013; 3:1462. [PMID: 23492904 PMCID: PMC3598002 DOI: 10.1038/srep01462] [Citation(s) in RCA: 71] [Impact Index Per Article: 5.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/31/2012] [Accepted: 02/28/2013] [Indexed: 11/30/2022] Open
Abstract
While common in viral infections and neoplasia, spontaneous cell-cell fusion, or syncytialization, is quite restricted in healthy tissues. Such fusion is essential to human placental development, where interactions between trophoblast-specific human endogenous retroviral (HERV) envelope proteins, called syncytins, and their widely-distributed cell surface receptors are centrally involved. We have identified the first host cell-encoded protein that inhibits cell fusion in mammals. Like the syncytins, this protein, called suppressyn, is HERV-derived, placenta-specific and well-conserved over simian evolution. In vitro, suppressyn binds to the syn1 receptor and inhibits syn1-, but not syn2-mediated trophoblast syncytialization. Suppressyn knock-down promotes cell-cell fusion in trophoblast cells and cell-associated and secreted suppressyn binds to the syn1 receptor, ASCT2. Identification of the first host cell-encoded inhibitor of mammalian cell fusion may encourage improved understanding of cell fusion mechanisms, of placental morphogenesis and of diseases resulting from abnormal cell fusion.
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13
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Dijkman R, Jebbink MF, Deijs M, Milewska A, Pyrc K, Buelow E, van der Bijl A, van der Hoek L. Replication-dependent downregulation of cellular angiotensin-converting enzyme 2 protein expression by human coronavirus NL63. J Gen Virol 2012; 93:1924-1929. [DOI: 10.1099/vir.0.043919-0] [Citation(s) in RCA: 115] [Impact Index Per Article: 8.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/22/2022] Open
Abstract
Like severe acute respiratory syndrome coronavirus (SARS-CoV), human coronavirus (HCoV)-NL63 employs angiotensin-converting enzyme 2 (ACE2) as a receptor for cellular entry. SARS-CoV infection causes robust downregulation of cellular ACE2 expression levels and it has been suggested that the SARS-CoV effect on ACE2 is involved in the severity of disease. We investigated whether cellular ACE2 downregulation occurs at optimal replication conditions of HCoV-NL63 infection. The expression of the homologue of ACE2, the ACE protein not used as a receptor by HCoV-NL63, was measured as a control. A specific decrease for ACE2 protein level was observed when HCoV-NL63 was cultured at 34 °C. Culturing the virus at the suboptimal temperature of 37 °C resulted in low replication of the virus and the effect on ACE2 expression was lost. We conclude that the decline of ACE2 expression is dependent on the efficiency of HCoV-NL63 replication, and that HCoV-NL63 and SARS-CoV both affect cellular ACE2 expression during infection.
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Affiliation(s)
- Ronald Dijkman
- Laboratory of Experimental Virology, Department of Medical Microbiology, Center for Infection and Immunity Amsterdam (CINIMA), Academic Medical Center, University of Amsterdam, The Netherlands
| | - Maarten F. Jebbink
- Laboratory of Experimental Virology, Department of Medical Microbiology, Center for Infection and Immunity Amsterdam (CINIMA), Academic Medical Center, University of Amsterdam, The Netherlands
| | - Martin Deijs
- Laboratory of Experimental Virology, Department of Medical Microbiology, Center for Infection and Immunity Amsterdam (CINIMA), Academic Medical Center, University of Amsterdam, The Netherlands
| | - Aleksandra Milewska
- Laboratory of Experimental Virology, Department of Medical Microbiology, Center for Infection and Immunity Amsterdam (CINIMA), Academic Medical Center, University of Amsterdam, The Netherlands
| | - Krzysztof Pyrc
- Microbiology Department, Faculty of Biochemistry, Biophysics and Biotechnology, Jagiellonian University, Krakow, Poland
| | - Elena Buelow
- Laboratory of Experimental Virology, Department of Medical Microbiology, Center for Infection and Immunity Amsterdam (CINIMA), Academic Medical Center, University of Amsterdam, The Netherlands
| | - Anna van der Bijl
- Laboratory of Experimental Virology, Department of Medical Microbiology, Center for Infection and Immunity Amsterdam (CINIMA), Academic Medical Center, University of Amsterdam, The Netherlands
| | - Lia van der Hoek
- Laboratory of Experimental Virology, Department of Medical Microbiology, Center for Infection and Immunity Amsterdam (CINIMA), Academic Medical Center, University of Amsterdam, The Netherlands
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14
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Biard-Piechaczyk M, Borel S, Espert L, de Bettignies G, Coux O. HIV-1, ubiquitin and ubiquitin-like proteins: the dialectic interactions of a virus with a sophisticated network of post-translational modifications. Biol Cell 2012; 104:165-87. [PMID: 22188301 DOI: 10.1111/boc.201100112] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/05/2011] [Accepted: 12/14/2011] [Indexed: 11/26/2022]
Abstract
The modification of intracellular proteins by ubiquitin (Ub) and ubiquitin-like (UbL) proteins is a central mechanism for regulating and fine-tuning all cellular processes. Indeed, these modifications are widely used to control the stability, activity and localisation of many key proteins and, therefore, they are instrumental in regulating cellular functions as diverse as protein degradation, cell signalling, vesicle trafficking and immune response. It is thus no surprise that pathogens in general, and viruses in particular, have developed multiple strategies to either counteract or exploit the complex mechanisms mediated by the Ub and UbL protein conjugation pathways. The aim of this review is to provide an overview on the intricate and conflicting relationships that intimately link HIV-1 and these sophisticated systems of post-translational modifications.
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Affiliation(s)
- Martine Biard-Piechaczyk
- Centre d'étude d'agents Pathogènes et Biotechnologies pour la Santé (CPBS-CNRS), Montpellier Cedex 5, France.
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15
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Mwimanzi P, Hasan Z, Tokunaga M, Gatanaga H, Oka S, Ueno T. Naturally arising HIV-1 Nef variants conferring escape from cytotoxic T lymphocytes influence viral entry co-receptor expression and susceptibility to superinfection. Biochem Biophys Res Commun 2010; 403:422-7. [PMID: 21093412 DOI: 10.1016/j.bbrc.2010.11.047] [Citation(s) in RCA: 9] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/26/2010] [Accepted: 11/12/2010] [Indexed: 11/17/2022]
Abstract
HIV-1 Nef is a key factor for pathogenesis and is known to down-regulate functionally important molecules, including viral entry co-receptor CCR5 and CXCR4, from the surface of HIV-infected cells. Some of these Nef activities are mediated by the well-conserved proline-rich region of Nef, and this region is highly targeted by cytotoxic T lymphocytes (CTLs). In the present study, we asked whether Nef variants selected under CTL-mediated selective pressure in vivo may constrain these important Nef activities. The analysis of autologous nef sequences isolated from a cohort of total 235 subjects in Japan revealed that the subjects showing amino acid variations, such as Arg75Thr and Tyr85Phe, located within the proline-rich region were significantly over-represented by those having HLA-B*3501. CTL assays corroborated that these mutations conferred escape from HLA-B(∗)3501-restricted CTLs. The Arg75Thr variant Nef selectively impaired CCR5, but not CXCR4, down-regulation activity from the cell surface; whereas the Tyr85Phe variant Nef affected neither CCR5 nor CXCR4 down-regulation activity. Moreover, the cells expressing the Arg75Thr variant Nef significantly impaired protection from superinfection by CCR5-tropic, but not CXCR4-tropic, viruses. These results highlighted the importance of certain Nef-specific CTLs in modulation of viral co-receptor down-regulation activity and protection from HIV-1 superinfection, providing us with additional insight into vaccine design.
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Affiliation(s)
- Philip Mwimanzi
- Center for AIDS Research, Kumamoto University, Kumamoto, Japan
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16
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Laguette N, Brégnard C, Benichou S, Basmaciogullari S. Human immunodeficiency virus (HIV) type-1, HIV-2 and simian immunodeficiency virus Nef proteins. Mol Aspects Med 2010; 31:418-33. [PMID: 20594957 DOI: 10.1016/j.mam.2010.05.003] [Citation(s) in RCA: 42] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/15/2010] [Accepted: 05/26/2010] [Indexed: 11/19/2022]
Abstract
The genomes of all retroviruses encode the Gag Pol and Env structural proteins. Human and simian lentiviruses have acquired non-structural proteins among which Nef plays a major role in the evolution of viral infection towards an immunodeficiency syndrome. Indeed, in the absence of a functional nef gene, primate lentiviruses are far less pathogenic than their wild type counterparts. The multiple protein-protein interactions in which Nef is involved all contribute to explain the role played by Nef in HIV- and SIV-associated disease progression. This review summarizes common and distinct features among Nef proteins and how they contribute to increasing HIV and SIV fitness towards their respective hosts.
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Affiliation(s)
- Nadine Laguette
- Institut Cochin, CNRS UMR8104, Université Paris Descartes, Paris, France
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17
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Joseph SB, Hanley KA, Chao L, Burch CL. Coinfection rates in Φ6 bacteriophage are enhanced by virus-induced changes in host cells. Evol Appl 2009; 2:24-31. [PMID: 25567844 PMCID: PMC3352419 DOI: 10.1111/j.1752-4571.2008.00055.x] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/02/2008] [Accepted: 11/26/2008] [Indexed: 11/28/2022] Open
Abstract
Two or more viruses infecting the same host cell can interact in ways that profoundly affect disease dynamics and control, yet the factors determining coinfection rates are incompletely understood. Previous studies have focused on the mechanisms that viruses use to suppress coinfection, but recently the phenomenon of enhanced coinfection has also been documented. In the experiments described here, we explore the hypothesis that enhanced coinfection rates in the bacteriophage Φ6 are achieved by virus-induced upregulation of the Φ6 receptor, which is the bacterial pilus. First, we confirmed that coinfection enhancement in Φ6 is virus-mediated by showing that Φ6 attaches significantly faster to infected cells than to uninfected cells. Second, we explored the hypothesis that coinfection enhancement in Φ6 depends upon changes in the expression of an inducible receptor. Consistent with this hypothesis, the closely related phage, Φ12, that uses constitutively expressed lipopolysaccharide as its receptor, attaches to infected and uninfected cells at the same rate. Our results, along with the previous finding that coinfection in Φ6 is limited to two virions, suggest that viruses may closely regulate rates of coinfection through mechanisms for both coinfection enhancement and exclusion.
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Affiliation(s)
- Sarah B Joseph
- Department of Biology, University of North Carolina Chapel Hill, NC, USA
| | - Kathryn A Hanley
- Department of Biology, New Mexico State University Las Cruces, NM, USA
| | - Lin Chao
- Division of Biological Sciences, University of California San Diego, CA, USA
| | - Christina L Burch
- Department of Biology, University of North Carolina Chapel Hill, NC, USA
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18
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Thiel L, Diederich S, Erbar S, Pfaff D, Augustin HG, Maisner A. Ephrin-B2 expression critically influences Nipah virus infection independent of its cytoplasmic tail. Virol J 2008; 5:163. [PMID: 19108727 PMCID: PMC2628893 DOI: 10.1186/1743-422x-5-163] [Citation(s) in RCA: 13] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/11/2008] [Accepted: 12/24/2008] [Indexed: 11/11/2022] Open
Abstract
Background Cell entry and cell-to-cell spread of the highly pathogenic Nipah virus (NiV) requires binding of the NiV G protein to cellular ephrin receptors and subsequent NiV F-mediated fusion. Since expression levels of the main NiV entry receptor ephrin-B2 (EB2) are highly regulated in vivo to fulfill the physiological functions in axon guidance and angiogenesis, the goal of this study was to determine if changes in the EB2 expression influence NiV infection. Results Surprisingly, transfection of increasing EB2 plasmid concentrations reduced cell-to-cell fusion both in cells expressing the NiV glycoproteins and in cells infected with NiV. This effect was attributed to the downregulation of the NiV glycoproteins from the cell surface. In addition to the influence on cell-to-cell fusion, increased EB2 expression significantly reduced the total amount of NiV-infected cells, thus interfered with virus entry. To determine if the negative effect of elevated EB2 expression on virus entry is a result of an increased EB2 signaling, receptor function of a tail-truncated and therefore signaling-defective ΔcEB2 was tested. Interestingly, ΔcEB2 fully functioned as NiV entry and fusion receptor, and overexpression also interfered with virus replication. Conclusion Our findings clearly show that EB2 signaling does not account for the striking negative impact of elevated receptor expression on NiV infection, but rather that the ratio between the NiV envelope glycoproteins and surface receptors critically influence cell-to-cell fusion and virus entry.
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Affiliation(s)
- Lena Thiel
- Institute of Virology, Philipps University of Marburg, Marburg, Germany.
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19
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Claus C, Tzeng WP, Liebert UG, Frey TK. Rubella virus-induced superinfection exclusion studied in cells with persisting replicons. J Gen Virol 2007; 88:2769-2773. [PMID: 17872530 DOI: 10.1099/vir.0.83092-0] [Citation(s) in RCA: 17] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/18/2022] Open
Abstract
For the first time, homologous superinfection exclusion was documented for rubella virus (RUB) by using Vero cells harbouring persisting RUB replicons. Infection with wild-type RUB was reduced by tenfold, whereas Sindbis virus infection was unaffected. Replication following infection with packaged replicons and transfection with replicon transcripts was also restricted in these cells, indicating that restriction occurred after penetration and entry. Translation of such 'supertransfecting' replicon transcripts was not impaired, but no accumulation of supertransfecting replicon RNA could be detected. We tested the hypothesis favoured in the related alphaviruses that superinfection exclusion is mediated by cleavage of the incoming non-structural precursor by the pre-existing non-structural (NS) protease, resulting in an inhibition of minus-strand RNA synthesis. However, cleavage of a precursor translated from a supertransfecting replicon transcript with an NS protease catalytic-site mutation was not detected and the event in the replication cycle at which superinfection exclusion is executed remains to be elucidated.
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Affiliation(s)
- Claudia Claus
- Institute of Virology, University of Leipzig, Leipzig, Germany
| | - Wen-Pin Tzeng
- Department of Biology, Georgia State University, Atlanta, GA 30303, USA
| | - Uwe G Liebert
- Institute of Virology, University of Leipzig, Leipzig, Germany
| | - Teryl K Frey
- Department of Biology, Georgia State University, Atlanta, GA 30303, USA
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20
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Sawatsky B, Grolla A, Kuzenko N, Weingartl H, Czub M. Inhibition of henipavirus infection by Nipah virus attachment glycoprotein occurs without cell-surface downregulation of ephrin-B2 or ephrin-B3. J Gen Virol 2007; 88:582-591. [PMID: 17251577 DOI: 10.1099/vir.0.82427-0] [Citation(s) in RCA: 12] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/02/2023] Open
Abstract
Nipah virus (NiV) and Hendra virus (HeV) are newly identified members of the family Paramyxoviridae and have been classified in the new genus Henipavirus based on unique genetic characteristics distinct from other paramyxoviruses. Transgenic cell lines were generated that expressed either the attachment protein (G) or the fusion protein (F) of NiV. Functional expression of NiV F and G was verified by complementation with the corresponding glycoprotein, which resulted in the development of syncytia. When exposed to NiV and HeV, expression of NiV G in Crandall feline kidney cells resulted in a qualitative inhibition of both cytopathic effect (CPE) and cell death by both viruses. RT-PCR analysis of surviving exposed cells showed a complete absence of viral positive-sense mRNA and genomic negative-sense viral RNA. Cells expressing NiV G were also unable to fuse with cells co-expressing NiV F and G in a fluorescent fusion inhibition assay. Cell-surface staining for the cellular receptors for NiV and HeV (ephrin-B2 and ephrin-B3) indicated that they were located on the surface of cells, regardless of NiV G expression or infection by NiV. These results indicated that viral interference can be established for henipaviruses and requires only the expression of the attachment protein, G. Furthermore, it was found that this interference probably occurs at the level of virus entry, as fusion was not observed in cells expressing NiV G. Finally, expression of NiV G by either transient transfection or NiV infection did not alter the cell-surface levels of the two known viral receptors.
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Affiliation(s)
- Bevan Sawatsky
- Department of Medical Microbiology, University of Manitoba, 730 William Avenue, Winnipeg, MB R3E 0W3, Canada
- National Microbiology Laboratory, Canadian Science Centre for Human and Animal Health, 1015 Arlington Street, Winnipeg, MB R3E 3R2, Canada
| | - Allen Grolla
- National Microbiology Laboratory, Canadian Science Centre for Human and Animal Health, 1015 Arlington Street, Winnipeg, MB R3E 3R2, Canada
| | - Nina Kuzenko
- Department of Medical Microbiology, University of Manitoba, 730 William Avenue, Winnipeg, MB R3E 0W3, Canada
- National Microbiology Laboratory, Canadian Science Centre for Human and Animal Health, 1015 Arlington Street, Winnipeg, MB R3E 3R2, Canada
| | - Hana Weingartl
- National Centre for Foreign Animal Disease, Canadian Science Centre for Human and Animal Health, 1015 Arlington Street, Winnipeg, MB R3E 3R2, Canada
- Department of Medical Microbiology, University of Manitoba, 730 William Avenue, Winnipeg, MB R3E 0W3, Canada
| | - Markus Czub
- Department of Medical Microbiology, University of Manitoba, 730 William Avenue, Winnipeg, MB R3E 0W3, Canada
- National Microbiology Laboratory, Canadian Science Centre for Human and Animal Health, 1015 Arlington Street, Winnipeg, MB R3E 3R2, Canada
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21
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Abstract
Hepadnaviridae is a family of hepatotropic DNA viruses that is divided into the genera orthohepadnavirus of mammals and avihepadnavirus of birds. All members of this family can cause acute and chronic hepatic infection, which in the case of human hepatitis B virus (HBV) constitutes a major global health problem. Although our knowledge about the molecular biology of these highly liver-specific viruses has profoundly increased in the last two decades, the mechanisms of attachment and productive entrance into the differentiated host hepatocytes are still enigmatic. The difficulties in studying hepadnaviral entry were primarily caused by the lack of easily accessible in vitro infection systems. Thus, for more than twenty years, differentiated primary hepatocytes from the respective species were the only in vitro models for both orthohepadnaviruses (e.g. HBV) and avihepadnaviruses (e.g. duck hepatitis B virus [DHBV]). Two important discoveries have been made recently regarding HBV: (1) primary hepatocytes from tree-shrews; i.e., Tupaia belangeri, can be substituted for primary human hepatocytes, and (2) a human hepatoma cell line (HepaRG) was established that gains susceptibility for HBV infection upon induction of differentiation in vitro. A number of potential HBV receptor candidates have been described in the past, but none of them have been confirmed to function as a receptor. For DHBV and probably all other avian hepadnaviruses, carboxypeptidase D (CPD) has been shown to be indispensable for infection, although the exact role of this molecule is still under debate. While still restricted to the use of primary duck hepatocytes (PDH), investigations performed with DHBV provided important general concepts on the first steps of hepadnaviral infection. However, with emerging data obtained from the new HBV infection systems, the hope that DHBV utilizes the same mechanism as HBV only partially held true. Nevertheless, both HBV and DHBV in vitro infection systems will help to: (1) functionally dissect the hepadnaviral entry pathways, (2) perform reverse genetics (e.g. test the fitness of escape mutants), (3) titrate and map neutralizing antibodies, (4) improve current vaccines to combat acute and chronic infections of hepatitis B, and (5) develop entry inhibitors for future clinical applications.
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Affiliation(s)
- Dieter Glebe
- Institute of Medical Virology, Justus-Liebig University of Giessen, Frankfurter Strasse 107, D-35392 Giessen, Germany.
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22
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Venzke S, Michel N, Allespach I, Fackler OT, Keppler OT. Expression of Nef downregulates CXCR4, the major coreceptor of human immunodeficiency virus, from the surfaces of target cells and thereby enhances resistance to superinfection. J Virol 2006; 80:11141-52. [PMID: 16928758 PMCID: PMC1642143 DOI: 10.1128/jvi.01556-06] [Citation(s) in RCA: 63] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
Lentiviral Nef proteins are key factors for pathogenesis and are known to downregulate functionally important molecules, including CD4 and major histocompatibility complex class I (MHC-I), from the surfaces of infected cells. Recently, we demonstrated that Nef reduces cell surface levels of the human immunodeficiency virus type 1 (HIV-1) entry coreceptor CCR5 (N. Michel, I. Allespach, S. Venzke, O. T. Fackler, and O. T. Keppler, Curr. Biol. 15:714-723, 2005). Here, we report that Nef downregulates the second major HIV-1 coreceptor, CXCR4, from the surfaces of HIV-infected primary CD4 T lymphocytes with efficiencies comparable to those of the natural CXCR4 ligand, stromal cell-derived factor-1 alpha. Analysis of a panel of mutants of HIV-1(SF2) Nef revealed that the viral protein utilized the same signature motifs for downmodulation of CXCR4 and MHC-I, including the proline-rich motif P(73)P(76)P(79)P(82) and the acidic cluster motif E(66)E(67)E(68)E(69.) Expression of wild-type Nef, but not of specific Nef mutants, resulted in a perinuclear accumulation of the coreceptor. Remarkably, the carboxy terminus of CXCR4, which harbors the classical motifs critical for basal and ligand-induced receptor endocytosis, was dispensable for the Nef-mediated reduction of surface exposure. Functionally, the ability of Nef to simultaneously downmodulate CXCR4 and CD4 correlated with maximum-level protection of Nef-expressing target cells from fusion with cells exposing X4 HIV-1 envelopes. Furthermore, the Nef-mediated downregulation of CXCR4 alone on target T lymphocytes was sufficient to diminish cells' susceptibility to X4 HIV-1 virions at the entry step. The downregulation of chemokine coreceptors is a conserved activity of Nef to modulate infected cells, an important functional consequence of which is an enhanced resistance to HIV superinfection.
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Affiliation(s)
- Stephanie Venzke
- Department of Virology, University of Heidelberg, Im Neuenheimer Feld 324, 69120 Heidelberg, Germany
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23
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Marzi A, Wegele A, Pöhlmann S. Modulation of virion incorporation of Ebolavirus glycoprotein: effects on attachment, cellular entry and neutralization. Virology 2006; 352:345-56. [PMID: 16777170 DOI: 10.1016/j.virol.2006.04.038] [Citation(s) in RCA: 17] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/24/2006] [Revised: 02/23/2006] [Accepted: 04/27/2006] [Indexed: 01/09/2023]
Abstract
The filoviruses Ebolavirus (EBOV) and Marburgvirus (MARV) cause severe hemorrhagic fever in humans and are potential agents of biological warfare. The envelope glycoprotein (GP) of filoviruses mediates viral entry into cells and is an attractive target for therapeutic intervention and vaccine design. Here, we asked if the efficiency of virion incorporation of EBOV-GP impacts attachment and entry into target cells and modulates susceptibility to neutralizing antibodies. In order to control the level of EBOV-GP expression, we generated cell lines expressing the GPs of the four known EBOV subspecies in an inducible fashion. Regulated expression of GP on the cell surface allowed production of reporter viruses harboring different amounts of GP. A pronounced reduction of virion incorporation of EBOV-GP had relatively little effect on virion infectivity, suggesting that only a few copies of GP might be sufficient for efficient engagement of cellular receptors. In contrast, optimal interactions with cellular attachment factors like the DC-SIGN protein required incorporation of high amounts of GP. Antibody-mediated neutralization of virions bearing high amounts of GP was slightly more efficient than neutralization of virions harboring low amounts of GP, suggesting that the efficiency of GP incorporation into virions might modulate susceptibility to neutralizing antibodies. Finally, regulated expression of GP in permissive 293 cells did not reduce EBOV-GP-driven infection but diminished vesicular stomatitis virus GP (VSV-G) and amphotropic murine leukemia virus (A-MLV) GP mediated entry in a dose-dependent manner. Therefore, intracellular GP does not seem to downmodulate expression of its receptor(s) but might alter expression and/or function of molecules involved in VSV-G and A-MLV-GP-dependent entry. Our results suggest that the efficiency of virion incorporation of GP could impact EBOV attachment to target cells and might modulate control of viral spread by the humoral immune response.
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Affiliation(s)
- Andrea Marzi
- Institute for Clinical and Molecular Virology, University Erlangen-Nürnberg, 91054 Erlangen, Germany
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24
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Dandri M, Volz TK, Lütgehetmann M, Petersen J. Animal models for the study of HBV replication and its variants. J Clin Virol 2005; 34 Suppl 1:S54-62. [PMID: 16461225 DOI: 10.1016/s1386-6532(05)80011-3] [Citation(s) in RCA: 33] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
Abstract
Enormous progresses in hepatitis B virus research have been made through the identification of avian and mammalian HBV related viruses, which offer ample opportunities for studies in naturally occurring hosts. However, none of these natural hosts belongs to the commonly used laboratory animals, and the development of various mouse strains carrying HBV transgenes offered unique opportunities to investigate some mechanisms of viral pathogenesis. Furthermore, the need to perform infection studies in a system harbouring HBV-permissive hepatocytes has lately led researchers to create new challenging human mouse chimera models of HBV infection. In this review, we will overview the type of animal models currently available in hepadnavirus research.
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Affiliation(s)
- M Dandri
- Department of Medicine, University Hospital Eppendorf University of Hamburg, Martinistr 52, D-20246 Hamburg, Germany
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25
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Michel N, Allespach I, Venzke S, Fackler OT, Keppler OT. The Nef protein of human immunodeficiency virus establishes superinfection immunity by a dual strategy to downregulate cell-surface CCR5 and CD4. Curr Biol 2005; 15:714-23. [PMID: 15854903 DOI: 10.1016/j.cub.2005.02.058] [Citation(s) in RCA: 128] [Impact Index Per Article: 6.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/23/2004] [Revised: 02/21/2005] [Accepted: 02/21/2005] [Indexed: 10/25/2022]
Abstract
BACKGROUND Viruses frequently render cells refractory to subsequent infection with the same virus. This state of superinfection immunity counteracts potentially detrimental consequences for the infected cell and facilitates high-level replication and viral spread in the host. RESULTS Here, we show that human immunodeficiency virus (HIV) employs its early gene product Nef to efficiently interfere with superinfection at the viral-entry step. In this context, we identify the downregulation of cell-surface CCR5, the major HIV coreceptor, as a novel and highly conserved activity of Nef. Nef targets the CCR5 coreceptor and the HIV binding receptor CD4 via distinct cellular machineries to enhance the endocytosis rate of both HIV receptor components and to accelerate their degradation. Functionally, these genetically separable actions by Nef synergized to efficiently protect cells from HIV superinfection at the level of fusion of the viral envelope with the plasma membrane. CONCLUSIONS HIV has evolved two independent activities for Nef to downregulate the receptor complex and to facilitate its efficient replication and spread. This evasion strategy likely represents a mechanism by which the pathogenicity factor Nef elevates viral replication in vivo and thus promotes AIDS pathogenesis.
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Affiliation(s)
- Nico Michel
- Department of Virology, University of Heidelberg, Germany
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26
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Ludlow M, McQuaid S, Cosby SL, Cattaneo R, Rima BK, Duprex WP. Measles virus superinfection immunity and receptor redistribution in persistently infected NT2 cells. J Gen Virol 2005; 86:2291-2303. [PMID: 16033977 DOI: 10.1099/vir.0.81052-0] [Citation(s) in RCA: 19] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/15/2022] Open
Abstract
A recombinant measles virus (MV) expressing red fluorescent protein (MVDsRed1) was used to produce a persistently infected cell line (piNT2-MVDsRed1) from human neural precursor (NT2) cells. A similar cell line (piNT2-MVeGFP) was generated using a virus that expresses enhanced green fluorescent protein. Intracytoplasmic inclusions containing the viral nucleocapsid protein were evident in all cells and viral glycoproteins were present at the cell surface. Nevertheless, the cells did not release infectious virus nor did they fuse to generate syncytia. Uninfected NT2 cells express the MV receptor CD46 uniformly over their surface, whereas CD46 was present in cell surface aggregates in the piNT2 cells. There was no decrease in the overall amount of CD46 in piNT2 compared to NT2 cells. Cell-to-cell fusion was observed when piNT2 cells were overlaid onto confluent monolayers of MV receptor-positive cells, indicating that the viral glycoproteins were correctly folded and processed. Infectious virus was released from the underlying cells, indicating that persistence was not due to gross mutations in the virus genome. Persistently infected cells were superinfected with MV or canine distemper virus and cytopathic effects were not observed. However, mumps virus could readily infect the cells, indicating that superinfection immunity is not caused by general soluble antiviral factors. As MVeGFP and MVDsRed1 are antigenically indistinguishable but phenotypically distinct it was possible to use them to measure the degree of superinfection immunity in the absence of any cytopathic effect. Only small numbers of non-fusing green fluorescent piNT2-MVDsRed1 cells (1 : 300 000) were identified in which superinfecting MVeGFP entered, replicated and expressed its genes.
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Affiliation(s)
- Martin Ludlow
- School of Biology and Biochemistry, The Queen's University of Belfast, Belfast BT9 7BL, Northern Ireland, UK
| | - Stephen McQuaid
- Molecular Pathology Laboratory, Royal Group of Hospitals Trust, Belfast BT12 6BL, Northern Ireland, UK
| | - S Louise Cosby
- School of Medicine, The Queen's University of Belfast, Belfast BT9 7BL, Northern Ireland, UK
| | - Roberto Cattaneo
- Molecular Medicine Program, Mayo Clinic, Guggenheim 18, Rochester, MN 55905, USA
| | - Bert K Rima
- School of Biology and Biochemistry, The Queen's University of Belfast, Belfast BT9 7BL, Northern Ireland, UK
| | - W Paul Duprex
- School of Biology and Biochemistry, The Queen's University of Belfast, Belfast BT9 7BL, Northern Ireland, UK
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Thepparit C, Smith DR. Serotype-specific entry of dengue virus into liver cells: identification of the 37-kilodalton/67-kilodalton high-affinity laminin receptor as a dengue virus serotype 1 receptor. J Virol 2004; 78:12647-56. [PMID: 15507651 PMCID: PMC525075 DOI: 10.1128/jvi.78.22.12647-12656.2004] [Citation(s) in RCA: 184] [Impact Index Per Article: 8.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/31/2022] Open
Abstract
Dengue virus, the causative agent of dengue fever, dengue shock syndrome, and dengue hemorrhagic fever, infects susceptible cells by initially binding to a receptor(s) located on the host cell surface. Evidence to date suggests that receptor usage may be cell and serotype specific, and this study sought to identify dengue virus serotype 1 binding proteins on the surface of liver cells, a known target organ. By using a virus overlay protein binding assay (VOPBA), in both nondenaturing and denaturing gel systems, a putative dengue virus serotype 1 binding protein of approximately 37 kDa expressed on the surface of liver (HepG2) cells was identified. Mass spectrometry analysis identified a candidate protein, the 37/67-kDa high-affinity laminin receptor. Entry of the dengue virus serotype 1 was significantly inhibited in a dose-dependent manner by both antibodies directed against the 37/67-kDa high-affinity laminin receptor and soluble laminin. No inhibition of virus entry was seen with dengue virus serotypes 2, 3, or 4, demonstrating that the 37/67-kDa high-affinity laminin receptor is a serotype-specific receptor for dengue virus entry into liver cells.
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Affiliation(s)
- Chutima Thepparit
- Molecular Pathology Laboratory, Institute of Molecular Biology and Genetics, Mahidol University, 25/25 Phuttamontol Sai 4, Salaya, Nakorn Pathom, Thailand 73170
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Walters KA, Joyce MA, Addison WR, Fischer KP, Tyrrell DLJ. Superinfection exclusion in duck hepatitis B virus infection is mediated by the large surface antigen. J Virol 2004; 78:7925-37. [PMID: 15254165 PMCID: PMC446106 DOI: 10.1128/jvi.78.15.7925-7937.2004] [Citation(s) in RCA: 43] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/28/2004] [Accepted: 03/19/2004] [Indexed: 12/17/2022] Open
Abstract
Superinfection exclusion is the phenomenon whereby a virus prevents the subsequent infection of an already infected host cell. The Pekin duck hepatitis B virus (DHBV) model was used to investigate superinfection exclusion in hepadnavirus infections. Superinfection exclusion was shown to occur both in vivo and in vitro with a genetically marked DHBV, DHBV-ClaI, which was unable to establish an infection in either DHBV-infected ducklings or DHBV-infected primary duck hepatocytes (PDHs). In addition, exclusion occurred in vivo even when the second virus had a replicative advantage. Superinfection exclusion appears to be restricted to DHBV, as adenovirus, herpes simplex virus type 1, and vesicular stomatitis virus were all capable of efficiently infecting DHBV-infected PDHs. Exclusion was dependent on gene expression by the original infecting virus, since UV-irradiated DHBV was unable to mediate the exclusion of DHBV-ClaI. Using recombinant adenoviruses expressing DHBV proteins, we determined that the large surface antigen mediated exclusion. The large surface antigen is known to cause down-regulation of a DHBV receptor, carboxypeptidase D (CPD). Receptor down-regulation is a mechanism of superinfection exclusion seen in other viral infections, and so it was investigated as a possible mechanism of DHBV-mediated exclusion. However, a mutant large surface antigen which did not down-regulate CPD was still capable of inhibiting DHBV infection of PDHs. In addition, exclusion of DHBV-ClaI did not correlate with a decrease in CPD levels. Finally, virus binding assays and confocal microscopy analysis of infected PDHs indicated that the block in infection occurs after internalization of the second virus. We suggest that superinfection exclusion may result from the role of the L surface antigen as a regulator of intracellular trafficking.
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Affiliation(s)
- Kathie-Anne Walters
- Department of Medical Microbiology and Immunology, University of Alberta, Edmonton, Alberta, Canada
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Cooper A, Paran N, Shaul Y. The earliest steps in hepatitis B virus infection. BIOCHIMICA ET BIOPHYSICA ACTA 2003; 1614:89-96. [PMID: 12873769 DOI: 10.1016/s0005-2736(03)00166-4] [Citation(s) in RCA: 23] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/23/2022]
Abstract
The early steps in hepatitis B virus (HBV) infection, a human hepadnavirus, initiates from cell attachment followed by entry and delivery of the genetic information to the nucleus. Despite the fact that these steps determine the virus-related pathogenesis, their molecular basis is poorly understood. Cumulative data suggest that this process can be divided to cell attachment, endocytosis, membrane fusion and post-fusion consecutive steps. These steps are likely to be regulated by the viral envelope proteins and by the cellular membrane, receptors and extracellular matrix. In the absence of animal model for HBV, the duck hepadnavirus DHBV turned out to be a fruitful animal model. Therefore data concerning the early, post-attachment steps in hepadnaviral entry are largely based on studies performed with DHBV in primary duck liver hepatocytes. These studies are now starting to illuminate the mechanisms of hepadnavirus route of cell entry and to provide some new insights on the molecular basis of the strict species specificity of hepadnavirus infection.
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Affiliation(s)
- Arik Cooper
- Department of Molecular Genetics, Weizmann Institute of Science, Rehovot 76100, Israel
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Yao E, Schaller H, Tavis JE. The duck hepatitis B virus polymerase and core proteins accumulate in different patterns from their common mRNA. Virology 2003; 311:81-8. [PMID: 12832205 DOI: 10.1016/s0042-6822(03)00142-9] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/15/2023]
Abstract
Hepadnaviral reverse transcription occurs in capsids in which the core (C) protein surrounds the reverse transcriptase (P) and pregenomic RNA (pgRNA). We analyzed the accumulation patterns of duck hepatitis B virus P, C, and pgRNA in transfected LMH cells, infected primary duck hepatocytes (PDH), and infected duck liver. In all three systems, P accumulated over time in a different pattern compared with C, despite translation of both proteins from the pgRNA. Although the accumulation patterns of the proteins varied between the systems, in each case P became detectable at the same time or earlier than C and the ratio of P relative to C dropped with time. These accumulation patterns were consistent with the translation rates and half-lives of P and C. Comparing the translation rates of P and C with the pgRNA level over time revealed that translation of P and C was negatively regulated in LMH cells. These data provide a framework for comparing replication studies performed in LMH cells, PDHs and ducks.
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Affiliation(s)
- Ermei Yao
- Department of Molecular Microbiology and Immunology, Saint Louis University School of Medicine, 1402 South Grand Boulevard, St. Louis, MO 63104, USA
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31
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Abstract
Virus infection is initiated by recognition and attachment of the virus to the cell surface. Despite the fact that this interaction determines the virus-related pathogenesis, its molecular basis remained obscure for HBV. This process is mediated primarily by the viral envelope and the cellular receptors. HBV infection is not exceptional in this regard but its putative receptors have not been identified yet. The recent development of protocols to establish HBV susceptible cell lines and unique tools to measure HBV-cell attachment at a single cell resolution set the stage for the study of HBV-host cell interaction. These studies revealed that the QLDPAF epitope of the HBV surface antigen large protein (LHBsAg) plays a major role in this process. Quantitative measurements suggested the presence of a second player in this process and both act synergistically to improve cell attachment. As the step of virus-cell attachment is potentially susceptible to specific inhibitors, understanding the molecular basis of virus-cell attachment can be expected to have therapeutic impacts.
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Affiliation(s)
- Nir Paran
- Department of Molecular Genetics, Weizmann Institute of Science, Rehovot 76100, Israel
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Geib T, Sauder C, Venturelli S, Hässler C, Staeheli P, Schwemmle M. Selective virus resistance conferred by expression of Borna disease virus nucleocapsid components. J Virol 2003; 77:4283-90. [PMID: 12634385 PMCID: PMC150622 DOI: 10.1128/jvi.77.7.4283-4290.2003] [Citation(s) in RCA: 61] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/21/2022] Open
Abstract
Persistent viral infections can render host cells resistant to superinfection with closely related viruses by largely uncharacterized mechanisms. We present evidence for superinfection exclusion in brains of Borna disease virus (BDV)-infected rats and in persistently infected Vero cells, and we suggest that acquired resistance to BDV is due to unbalanced intracellular levels of viral nucleocapsid components. We observed that expression of BDV protein P, N, or X rendered human cells resistant to subsequent challenge with BDV but not with other RNA viruses, indicating that incorrect stoichiometry of nucleocapsid components selectively blocked the polymerase activity of incoming viruses. Vero cells containing high levels of an untranslatable BDV-N transcript remained virus susceptible, demonstrating that viral protein rather than RNA mediated resistance. Transient overexpression of BDV-P in persistently infected Vero cells was also remarkably effective against BDV, indicating that the intracellular balance of viral nucleocapsid components could serve as a target for future therapeutic antiviral strategies.
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Affiliation(s)
- Till Geib
- Department of Virology, Institute for Medical Microbiology and Hygiene, University of Freiburg, Germany
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Spangenberg HC, Lee HB, Li J, Tan F, Skidgel R, Wands JR, Tong S. A short sequence within domain C of duck carboxypeptidase D is critical for duck hepatitis B virus binding and determines host specificity. J Virol 2001; 75:10630-42. [PMID: 11602705 PMCID: PMC114645 DOI: 10.1128/jvi.75.22.10630-10642.2001] [Citation(s) in RCA: 17] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022] Open
Abstract
Virus-cell surface receptor interactions are of major interest. Hepadnaviruses are a family of partially double-stranded DNA viruses with liver tropism and a narrow host range of susceptibility to infection. At least in the case of duck hepatitis B virus (DHBV), host specificity seems controlled partly at the receptor level. The middle portion in the pre-S region of the viral large envelope protein binds specifically to duck carboxypeptidase D (DCPD) but not to its human or chicken homologue. Although domain C of DCPD is implicated in ligand binding, the exact pre-S contact site remains to be determined. We prepared and tested a panel of chimeric constructs consisting of DCPD and human carboxypeptidase D (HCPD). Our results indicate that a short region at the N terminus of domain C (residues 920 to 949) is critical to DHBV binding and is a major determinant for the host specificity of DHBV infection. Replacing this region of the DCPD molecule with its human homologue abolished the DHBV interaction, whereas introducing this DCPD sequence into HCPD conferred efficient DHBV binding. Extensive analysis of site-directed mutants revealed that both conserved and nonconserved residues were important for the pre-S interaction. There were primary sequence variations and secondary structural differences that contributed to the inability of HCPD to bind the DHBV pre-S domain.
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Affiliation(s)
- H C Spangenberg
- Liver Research Center, Rhode Island Hospital and Brown University School of Medicine, Providence, Rhode Island 02903, USA
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Chiou HC, Lucas MA, Coffin CC, Banaszczyk MG, Ill CR, Lollo CP. Gene therapy strategies for the treatment of chronic viral hepatitis. Expert Opin Biol Ther 2001; 1:629-39. [PMID: 11727499 DOI: 10.1517/14712598.1.4.629] [Citation(s) in RCA: 10] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/26/2022]
Abstract
Chronic viral hepatitis is a major clinical problem, with over half a billion persons infected worldwide. Current therapies, principally treatment with recombinant IFN-alpha protein, have limited benefit. Recent studies suggest that gene-based expression of IFN-alpha is a possible therapeutic alternative that may improve the effectiveness of treatment. Gene delivery to the liver and consequent IFN-alpha expression therein, has the potential to concentrate the protein at the target organ and provide more continuous exposure to the therapeutic agent. Other potential gene and nucleic acid therapeutics for viral hepatitis are also being investigated. Key to the deployment of these future therapies is a suitable method of gene delivery. Although recombinant viral vector systems, such as adenovirus, are currently the most effective means of gene delivery to the liver, their use presents many concerns. These include immune and inflammatory reactions to the viral vector and possible adverse interactions between the recombinant virus and the pre-existing viral infection. Non-viral gene delivery systems would be a preferred treatment modality. The efficiency of current non-viral systems is not adequate for systemically administered liver gene therapy. However, recent use of membrane permeabilisation techniques has shown that high efficiency non-viral gene transfer agents are possible. The future coupling of these improved delivery systems with gene- or nucleic acid-based therapeutics currently in development holds out great promise for new generations of antihepatitis therapies.
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Affiliation(s)
- H C Chiou
- Immune Response Corporation, 5935 Darwin Court, Carlsbad, CA 92008, USA.
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