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Alidjinou EK, Coulon P, Tetart M, Guigon A, Diarra A, Aissi E, Bazus H, Derdour V, Meybeck A, Viget N, Hober D, Bocket L, Robineau O. Defective HIV-1 DNA pol sequences are not associated with HIV-1 DNA levels and drive most APOBEC-context drug resistance mutations. J Antimicrob Chemother 2025; 80:941-946. [PMID: 39849847 DOI: 10.1093/jac/dkaf016] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/14/2024] [Accepted: 01/08/2025] [Indexed: 01/25/2025] Open
Abstract
INTRODUCTION The specificity of HIV-1 DNA genotypic resistance tests (GRTs) is hampered by the detection of the APOBEC-context drug resistance mutations (AC DRMs), usually harboured by replication-incompetent proviruses. We sought factors associated with defective sequences in the HIV-1 pol region. In addition, AC DRMs and their link with defective sequences were investigated. METHODS We included ART-treated patients with viral suppression or plasma viral load (VL) lower than 200 copies/mL, who underwent HIV-1 DNA genotyping, with successful sequencing of protease (PR), reverse transcriptase (RT) and integrase (IN) regions. Sequencing was performed using either the Sanger method or the Sentosa® NGS approach with a 20% cut-off. All hypermutated sequences and/or those containing at least one stop codon were considered defective. RESULTS A total of 613 HIV-1 DNA GRTs were analysed. Defective sequences were identified for 186 samples (30.3%) including 65 PR sequences, 92 RT sequences and 65 IN sequences. No association, including HIV-1 DNA levels, was found with the detection of defective pol sequences. A total of 226 AC DRMs were recorded in all sequences. Most of these mutations (78%) were harboured by defective sequences. AC DRMs did not emerge in the plasma viral population, and likely do not impact the virological response to ART. CONCLUSIONS Defectives pol sequences were not associated with HIV-1 DNA levels and harboured most of the AC DRMs. Such mutations likely have no clinical impact, and should not be reported in routine practice. Consensus guidelines for reporting HIV-1 DNA GRTs are needed, especially for the assessment and management of AC DRMs.
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Affiliation(s)
| | - Pauline Coulon
- Laboratoire de Virologie ULR3610, Univ. Lille, CHU de Lille, 59000 Lille, France
| | - Macha Tetart
- CH Tourcoing, Service Universitaire des Maladies Infectieuses, 59200 Tourcoing, France
| | - Aurélie Guigon
- Laboratoire de Virologie ULR3610, Univ. Lille, CHU de Lille, 59000 Lille, France
| | - Ava Diarra
- CH Tourcoing, Service Universitaire des Maladies Infectieuses, 59200 Tourcoing, France
| | - Emmanuelle Aissi
- CH Lens, Service Universitaire des Maladies Infectieuses, 62300 Lens, France
| | - Hélène Bazus
- CH Lens, Service Universitaire des Maladies Infectieuses, 62300 Lens, France
| | - Vincent Derdour
- CH Tourcoing, Service Universitaire des Maladies Infectieuses, 59200 Tourcoing, France
| | - Agnès Meybeck
- CH Tourcoing, Service Universitaire des Maladies Infectieuses, 59200 Tourcoing, France
| | - Nathalie Viget
- CH Tourcoing, Service Universitaire des Maladies Infectieuses, 59200 Tourcoing, France
| | - Didier Hober
- Laboratoire de Virologie ULR3610, Univ. Lille, CHU de Lille, 59000 Lille, France
| | - Laurence Bocket
- Laboratoire de Virologie ULR3610, Univ. Lille, CHU de Lille, 59000 Lille, France
| | - Olivier Robineau
- CH Tourcoing, Service Universitaire des Maladies Infectieuses, 59200 Tourcoing, France
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Zhao W, Modak A, Ross SR. DHX15 inhibits mouse APOBEC3 deamination. PLoS Pathog 2025; 21:e1013045. [PMID: 40168451 PMCID: PMC11990775 DOI: 10.1371/journal.ppat.1013045] [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: 08/23/2024] [Revised: 04/11/2025] [Accepted: 03/14/2025] [Indexed: 04/03/2025] Open
Abstract
APOBEC3 family proteins are critical host factors that counteract and prevent the replication of retroviruses and other viruses through cytidine deamination. Human APOBEC3 proteins inactivate HIV-1 through the introduction of lethal mutations to viral genomes. In contrast, mouse APOBEC3 does not induce DNA hypermutation of murine retroviruses, although it retains functional cytidine deaminase activity. Why mouse APOBEC3 does not effectively deaminate murine retroviruses is still unknown. In this study, we found that the dead box helicase DHX15 interacts with mouse APOBEC3 and inhibits its deamination activity. DHX15 was packaged into murine leukemia virus (MLV) virions independent of its binding with APOBEC3. Moreover, DHX15 knockdown inhibited MLV replication and resulted in more G-to-A mutations in proviral DNA. Finally, DHX15 knockdown induced DNA damage in murine cells, suggesting that it plays a role in preserving genome integrity in cells expressing mouse APOBEC3 protein.
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Affiliation(s)
- Wenming Zhao
- Department of Microbiology and Immunology, University of Illinois Chicago College of Medicine, Chicago, Illinois, United States of America
| | - Ayan Modak
- Department of Microbiology and Immunology, University of Illinois Chicago College of Medicine, Chicago, Illinois, United States of America
| | - Susan R. Ross
- Department of Microbiology and Immunology, University of Illinois Chicago College of Medicine, Chicago, Illinois, United States of America
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Leong S, Nasser H, Ikeda T. APOBEC3-Related Editing and Non-Editing Determinants of HIV-1 and HTLV-1 Restriction. Int J Mol Sci 2025; 26:1561. [PMID: 40004025 PMCID: PMC11855278 DOI: 10.3390/ijms26041561] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/30/2024] [Revised: 02/09/2025] [Accepted: 02/11/2025] [Indexed: 02/27/2025] Open
Abstract
The apolipoprotein B mRNA editing enzyme catalytic polypeptide-like 3 (APOBEC3/A3) family of cytosine deaminases serves as a key innate immune barrier against invading retroviruses and endogenous retroelements. The A3 family's restriction activity against these parasites primarily arises from their ability to catalyze cytosine-to-uracil conversions, resulting in genome editing and the accumulation of lethal mutations in viral genomes. Additionally, non-editing mechanisms, including deaminase-independent pathways, such as blocking viral reverse transcription, have been proposed as antiviral strategies employed by A3 family proteins. Although viral factors can influence infection progression, the determinants that govern A3-mediated restriction are critical in shaping retroviral infection outcomes. This review examines the interactions between retroviruses, specifically human immunodeficiency virus type 1 and human T-cell leukemia virus type 1, and A3 proteins to better understand how editing and non-editing activities contribute to the trajectory of these retroviral infections.
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Affiliation(s)
- Sharee Leong
- Division of Molecular Virology and Genetics, Joint Research Center for Human Retrovirus Infection, Kumamoto University, Kumamoto 860-0811, Japan
- Graduate School of Medical Sciences, Kumamoto University, Kumamoto 860-8556, Japan
| | - Hesham Nasser
- Division of Molecular Virology and Genetics, Joint Research Center for Human Retrovirus Infection, Kumamoto University, Kumamoto 860-0811, Japan
| | - Terumasa Ikeda
- Division of Molecular Virology and Genetics, Joint Research Center for Human Retrovirus Infection, Kumamoto University, Kumamoto 860-0811, Japan
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4
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Lehle J, Soleimanpour M, Mokhtari S, Ebrahimi D. Viral infection, APOBEC3 dysregulation, and cancer. Front Genet 2024; 15:1489324. [PMID: 39764440 PMCID: PMC11701051 DOI: 10.3389/fgene.2024.1489324] [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: 08/31/2024] [Accepted: 11/26/2024] [Indexed: 03/06/2025] Open
Abstract
Viral infection plays a significant role in the development and progression of many cancers. Certain viruses, such as Human Papillomavirus (HPV), Epstein-Barr Virus (EBV), and Hepatitis B and C viruses (HBV, HCV), are well-known for their oncogenic potential. These viruses can dysregulate specific molecular and cellular processes through complex interactions with host cellular mechanisms. One such interaction involves a family of DNA mutators known as APOBEC3 (Apolipoprotein B mRNA Editing Catalytic Polypeptide-like 3). The primary function of these cytidine deaminases is to provide protection against viral infections by inducing viral mutagenesis. However, induction and dysregulation of A3 enzymes, driven by viral infection, can inadvertently lead to cellular DNA tumorigenesis. This review focuses on the current knowledge regarding the interplay between viral infection, A3 dysregulation, and cancer, highlighting the molecular mechanisms underlying this relationship.
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Affiliation(s)
- Jake Lehle
- Host-Pathogen Interaction Program, Texas Biomedical Research Institute, San Antonio, TX, United States
| | - Mohadeseh Soleimanpour
- Host-Pathogen Interaction Program, Texas Biomedical Research Institute, San Antonio, TX, United States
| | - Samira Mokhtari
- Host-Pathogen Interaction Program, Texas Biomedical Research Institute, San Antonio, TX, United States
| | - Diako Ebrahimi
- Host-Pathogen Interaction Program, Texas Biomedical Research Institute, San Antonio, TX, United States
- Department of Microbiology, Immunology, and Molecular Genetics, University of Texas Health San Antonio, San Antonio, TX, United States
- Department Molecular Microbiology and Immunology, The University of Texas at San Antonio, San Antonio, TX, United States
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5
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Hu Y, Delviks-Frankenberry KA, Wu C, Arizaga F, Pathak VK, Xiong Y. Structural insights into PPP2R5A degradation by HIV-1 Vif. Nat Struct Mol Biol 2024; 31:1492-1501. [PMID: 38789685 DOI: 10.1038/s41594-024-01314-6] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/12/2023] [Accepted: 04/11/2024] [Indexed: 05/26/2024]
Abstract
HIV-1 Vif recruits host cullin-RING-E3 ubiquitin ligase and CBFβ to degrade the cellular APOBEC3 antiviral proteins through diverse interactions. Recent evidence has shown that Vif also degrades the regulatory subunits PPP2R5(A-E) of cellular protein phosphatase 2A to induce G2/M cell cycle arrest. As PPP2R5 proteins bear no functional or structural resemblance to A3s, it is unclear how Vif can recognize different sets of proteins. Here we report the cryogenic-electron microscopy structure of PPP2R5A in complex with HIV-1 Vif-CBFβ-elongin B-elongin C at 3.58 Å resolution. The structure shows PPP2R5A binds across the Vif molecule, with biochemical and cellular studies confirming a distinct Vif-PPP2R5A interface that partially overlaps with those for A3s. Vif also blocks a canonical PPP2R5A substrate-binding site, indicating that it suppresses the phosphatase activities through both degradation-dependent and degradation-independent mechanisms. Our work identifies critical Vif motifs regulating the recognition of diverse A3 and PPP2R5A substrates, whereby disruption of these host-virus protein interactions could serve as potential targets for HIV-1 therapeutics.
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Affiliation(s)
- Yingxia Hu
- Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT, USA
| | - Krista A Delviks-Frankenberry
- Viral Mutation Section, HIV Dynamics and Replication Program, Center for Cancer Research, National Cancer Institute at Frederick, Frederick, MD, USA
| | - Chunxiang Wu
- Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT, USA
| | - Fidel Arizaga
- Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT, USA
| | - Vinay K Pathak
- Viral Mutation Section, HIV Dynamics and Replication Program, Center for Cancer Research, National Cancer Institute at Frederick, Frederick, MD, USA.
| | - Yong Xiong
- Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT, USA.
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Burdick RC, Duchon A, Hu WS, Pathak VK. Imaging HIV-1 Nuclear Import, Uncoating, and Proviral Transcription. Methods Mol Biol 2024; 2807:15-30. [PMID: 38743218 DOI: 10.1007/978-1-0716-3862-0_2] [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] [Indexed: 05/16/2024]
Abstract
Live-cell imaging has become a powerful tool for dissecting the behavior of viral complexes during HIV-1 infection with high temporal and spatial resolution. Very few HIV-1 particles in a viral population are infectious and successfully complete replication (~1/50). Single-particle live-cell imaging enables the study of these rare infectious viral particles, which cannot be accomplished in biochemical assays that measure the average property of the entire viral population, most of which are not infectious. The timing and location of many events in the early stage of the HIV-1 life cycle, including nuclear import, uncoating, and integration, have only recently been elucidated. Live-cell imaging also provides a valuable approach to study interactions of viral and host factors in distinct cellular compartments and at specific stages of viral replication. Successful live-cell imaging experiments require careful consideration of the fluorescent labeling method used and avoid or minimize its potential impact on normal viral replication and produce misleading results. Ideally, it is beneficial to utilize multiple virus labeling strategies and compare the results to ensure that the virion labeling did not adversely influence the viral replication step that is under investigation. Another potential benefit of using different labeling strategies is that they can provide information about the state of the viral complexes. Here, we describe our methods that utilize multiple fluorescent protein labeling approaches to visualize and quantify important events in the HIV-1 life cycle, including docking HIV-1 particles with the nuclear envelope (NE) and their nuclear import, uncoating, and proviral transcription.
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Affiliation(s)
- Ryan C Burdick
- Viral Mutation Section, HIV Dynamics and Replication Program, Center for Cancer Research, National Cancer Institute at Frederick, Frederick, MD, USA
| | - Alice Duchon
- Viral Recombination Section, HIV Dynamics and Replication Program, Center for Cancer Research, National Cancer Institute at Frederick, Frederick, MD, USA
| | - Wei-Shau Hu
- Viral Recombination Section, HIV Dynamics and Replication Program, Center for Cancer Research, National Cancer Institute at Frederick, Frederick, MD, USA
| | - Vinay K Pathak
- Viral Mutation Section, HIV Dynamics and Replication Program, Center for Cancer Research, National Cancer Institute at Frederick, Frederick, MD, USA.
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Delviks-Frankenberry KA, Ojha CR, Hermann KJ, Hu WS, Torbett BE, Pathak VK. Potent dual block to HIV-1 infection using lentiviral vectors expressing fusion inhibitor peptide mC46- and Vif-resistant APOBEC3G. MOLECULAR THERAPY. NUCLEIC ACIDS 2023; 33:794-809. [PMID: 37662965 PMCID: PMC10470399 DOI: 10.1016/j.omtn.2023.08.007] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 05/05/2023] [Accepted: 08/08/2023] [Indexed: 09/05/2023]
Abstract
Gene therapy strategies that effectively inhibit HIV-1 replication are needed to reduce the requirement for lifelong antiviral therapy and potentially achieve a functional cure. We previously designed self-activating lentiviral vectors that efficiently delivered and expressed a Vif-resistant mutant of APOBEC3G (A3G-D128K) to T cells, which potently inhibited HIV-1 replication and spread with no detectable virus. Here, we developed vectors that express A3G-D128K, membrane-associated fusion inhibitor peptide mC46, and O6-methylguanine-DNA-methyltransferase (MGMT) selectable marker for in vivo selection of transduced CD34+ hematopoietic stem and progenitor cells. MGMT-selected T cell lines MT4, CEM, and PM1 expressing A3G-D128K (with or without mC46) potently inhibited NL4-3 infection up to 45 days post infection with no detectable viral replication. Expression of mC46 was sufficient to block infection >80% in a single-cycle assay. Importantly, expression of mC46 provided a selective advantage to the A3G-D128K-modified T cells in the presence of replication competent virus. This combinational approach to first block HIV-1 entry with mC46, and then block any breakthrough infection with A3G-D128K, could provide an effective gene therapy treatment and a potential functional cure for HIV-1 infection.
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Affiliation(s)
- Krista A. Delviks-Frankenberry
- Viral Mutation Section, HIV Dynamics and Replication Program, National Cancer Institute at Frederick, Frederick, MD 21702, USA
| | - Chet R. Ojha
- Center for Immunity and Immunotherapies, Seattle Children’s Research Institute, Seattle, WA 98101, USA
| | - Kip J. Hermann
- Department of Immunology and Microbiology, The Scripps Research Institute, La Jolla, CA 92037, USA
| | - Wei-Shau Hu
- Viral Recombination Section, HIV Dynamics and Replication Program, National Cancer Institute, Frederick, MD 21702, USA
| | - Bruce E. Torbett
- Center for Immunity and Immunotherapies, Seattle Children’s Research Institute, Seattle, WA 98101, USA
- Department of Immunology and Microbiology, The Scripps Research Institute, La Jolla, CA 92037, USA
- Department of Pediatrics, University of Washington School of Medicine, Seattle, WA 98101, USA
| | - Vinay K. Pathak
- Viral Mutation Section, HIV Dynamics and Replication Program, National Cancer Institute at Frederick, Frederick, MD 21702, USA
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Ajoge HO, Renner TM, Bélanger K, Greig M, Dankar S, Kohio HP, Coleman MD, Ndashimye E, Arts EJ, Langlois MA, Barr SD. Antiretroviral APOBEC3 cytidine deaminases alter HIV-1 provirus integration site profiles. Nat Commun 2023; 14:16. [PMID: 36627271 PMCID: PMC9832166 DOI: 10.1038/s41467-022-35379-y] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/11/2019] [Accepted: 11/30/2022] [Indexed: 01/12/2023] Open
Abstract
APOBEC3 (A3) proteins are host-encoded deoxycytidine deaminases that provide an innate immune barrier to retroviral infection, notably against HIV-1. Low levels of deamination are believed to contribute to the genetic evolution of HIV-1, while intense catalytic activity of these proteins can induce catastrophic hypermutation in proviral DNA leading to near-total HIV-1 restriction. So far, little is known about how A3 cytosine deaminases might impact HIV-1 proviral DNA integration sites in human chromosomal DNA. Using a deep sequencing approach, we analyze the influence of catalytic active and inactive APOBEC3F and APOBEC3G on HIV-1 integration site selections. Here we show that DNA editing is detected at the extremities of the long terminal repeat regions of the virus. Both catalytic active and non-catalytic A3 mutants decrease insertions into gene coding sequences and increase integration sites into SINE elements, oncogenes and transcription-silencing non-B DNA features. Our data implicates A3 as a host factor influencing HIV-1 integration site selection and also promotes what appears to be a more latent expression profile.
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Affiliation(s)
- Hannah O Ajoge
- Western University, Schulich School of Medicine and Dentistry, Department of Microbiology and Immunology, London, ON, Canada
| | - Tyler M Renner
- Department of Biochemistry, Microbiology and Immunology, Faculty of Medicine, University of Ottawa, Ottawa, ON, Canada
| | - Kasandra Bélanger
- Department of Biochemistry, Microbiology and Immunology, Faculty of Medicine, University of Ottawa, Ottawa, ON, Canada
| | - Matthew Greig
- Department of Biochemistry, Microbiology and Immunology, Faculty of Medicine, University of Ottawa, Ottawa, ON, Canada
| | - Samar Dankar
- Department of Biochemistry, Microbiology and Immunology, Faculty of Medicine, University of Ottawa, Ottawa, ON, Canada
| | - Hinissan P Kohio
- Western University, Schulich School of Medicine and Dentistry, Department of Microbiology and Immunology, London, ON, Canada
| | - Macon D Coleman
- Western University, Schulich School of Medicine and Dentistry, Department of Microbiology and Immunology, London, ON, Canada
| | - Emmanuel Ndashimye
- Western University, Schulich School of Medicine and Dentistry, Department of Microbiology and Immunology, London, ON, Canada
| | - Eric J Arts
- Western University, Schulich School of Medicine and Dentistry, Department of Microbiology and Immunology, London, ON, Canada
| | - Marc-André Langlois
- Department of Biochemistry, Microbiology and Immunology, Faculty of Medicine, University of Ottawa, Ottawa, ON, Canada. .,Ottawa Center for Infection, Immunity and Inflammation (CI3), Ottawa, ON, Canada.
| | - Stephen D Barr
- Western University, Schulich School of Medicine and Dentistry, Department of Microbiology and Immunology, London, ON, Canada.
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Dominant Negative Mutants of Human Immunodeficiency Virus Type 1 Viral Infectivity Factor (Vif) Disrupt Core-Binding Factor Beta-Vif Interaction. J Virol 2022; 96:e0055522. [PMID: 35950859 PMCID: PMC9472641 DOI: 10.1128/jvi.00555-22] [Citation(s) in RCA: 3] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
Apolipoprotein B mRNA-editing catalytic polypeptide-like 3 family members (APOBEC3s) are host restriction factors that inhibit viral replication. Viral infectivity factor (Vif), a human immunodeficiency virus type 1 (HIV-1) accessory protein, mediates the degradation of APOBEC3s by forming the Vif-E3 complex, in which core-binding factor beta (CBFβ) is an essential molecular chaperone. Here, we screened nonfunctional Vif mutants with high affinity for CBFβ to inhibit HIV-1 in a dominant negative manner. We applied the yeast surface display technology to express Vif random mutant libraries, and mutants showing high CBFβ affinity were screened using flow cytometry. Most of the screened Vif mutants containing random mutations of different frequencies were able to rescue APOBEC3G (A3G). In the subsequent screening, three of the mutants restricted HIV-1, recovered G-to-A hypermutation, and rescued APOBEC3s. Among them, Vif-6M showed a cross-protection effect toward APOBEC3C, APOBEC3F, and African green monkey A3G. Stable expression of Vif-6M in T lymphocytes inhibited the viral replication in newly HIV-1-infected cells and the chronically infected cell line H9/HXB2. Furthermore, the expression of Vif-6M provided a survival advantage to T lymphocytes infected with HIV-1. These results suggest that dominant negative Vif mutants acting on the Vif-CBFβ target potently restrict HIV-1. IMPORTANCE Antiviral therapy cannot eliminate HIV and exhibits disadvantages such as drug resistance and toxicity. Therefore, novel strategies for inhibiting viral replication in patients with HIV are urgently needed. APOBEC3s in host cells are able to inhibit viral replication but are antagonized by HIV-1 Vif-mediated degradation. Therefore, we screened nonfunctional Vif mutants with high affinity for CBFβ to compete with the wild-type Vif (wtVif) as a potential strategy to assist with HIV-1 treatment. Most screened mutants rescued the expression of A3G in the presence of wtVif, especially Vif-6M, which could protect various APOBEC3s and improve the incorporation of A3G into HIV-1 particles. Transduction of Vif-6M into T lymphocytes inhibited the replication of the newly infected virus and the chronically infected virus. These data suggest that Vif mutants targeting the Vif-CBFβ interaction may be promising in the development of a new AIDS therapeutic strategy.
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Tikhonov AS, Mintaev RR, Glazkova DV, Bogoslovskaya EV, Shipulin GA. HIV Restriction Factor APOBEC3G and Prospects for Its Use in Gene Therapy for HIV. Mol Biol 2022. [DOI: 10.1134/s0026893322040112] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/23/2022]
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11
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A Conserved uORF Regulates APOBEC3G Translation and Is Targeted by HIV-1 Vif Protein to Repress the Antiviral Factor. Biomedicines 2021; 10:biomedicines10010013. [PMID: 35052693 PMCID: PMC8773096 DOI: 10.3390/biomedicines10010013] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/03/2021] [Revised: 12/17/2021] [Accepted: 12/18/2021] [Indexed: 11/17/2022] Open
Abstract
The HIV-1 Vif protein is essential for viral fitness and pathogenicity. Vif decreases expression of cellular restriction factors APOBEC3G (A3G), A3F, A3D and A3H, which inhibit HIV-1 replication by inducing hypermutation during reverse transcription. Vif counteracts A3G at several levels (transcription, translation, and protein degradation) that altogether reduce the levels of A3G in cells and prevent its incorporation into viral particles. How Vif affects A3G translation remains unclear. Here, we uncovered the importance of a short conserved uORF (upstream ORF) located within two critical stem-loop structures of the 5′ untranslated region (5′-UTR) of A3G mRNA for this process. A3G translation occurs through a combination of leaky scanning and translation re-initiation and the presence of an intact uORF decreases the extent of global A3G translation under normal conditions. Interestingly, the uORF is also absolutely required for Vif-mediated translation inhibition and redirection of A3G mRNA into stress granules. Overall, we discovered that A3G translation is regulated by a small uORF conserved in the human population and that Vif uses this specific feature to repress its translation.
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Abdi B, Lambert-Niclot S, Wirden M, Jary A, Teyssou E, Sayon S, Palich R, Tubiana R, Simon A, Valantin MA, Katlama C, Morand-Joubert L, Calvez V, Marcelin AG, Soulie C. Presence of HIV-1 G-to-A mutations linked to APOBEC editing is more prevalent in non-B HIV-1 subtypes and is associated with lower HIV-1 reservoir. J Antimicrob Chemother 2021; 76:2148-2152. [PMID: 33930161 DOI: 10.1093/jac/dkab123] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/09/2020] [Accepted: 03/14/2021] [Indexed: 12/16/2022] Open
Abstract
OBJECTIVES APOBEC3 editing activity contributes to sequences variation and viral diversification. We aimed to characterize virological and clinical factors associated with G-to-A mutations and stop codons in the HIV-1 reservoir, markers of APOBEC3 footprints, in order to better understand HIV-1 diversity among virologically suppressed HIV-1-infected patients. METHODS Immuno-virological and clinical factors were compared between 92 patients harbouring G-to-A mutations and stop codons (APOBEC+) in the reverse transcriptase gene and 92 patients without G-to-A mutations (APOBEC-) and stop codons in their DNA genotypes. RESULTS Patients were predominantly men (74.5%) and were mostly infected by B-subtype (69.0%), with 44.1% and 55.9% in APOBEC+ and APOBEC- groups, respectively. At time of HIV DNA genotypes, the total cell-associated HIV-1 DNA load was 2.34 log10 copies/106 cells (IQR 1.85-2.67) and 33.2% of them had a detectable ultrasensitive plasma viral load. Hypermutated sequences were identified in 28.2% of the APOBEC+ group. The median total cell-associated HIV-1 DNA level was significantly lower in APOBEC+ than APOBEC- group: 2.13 log10 copies/106 cells (IQR 1.60-2.60) versus 2.52 log10 copies/106 cells (IQR 2.19-2.71) (P < 0.001), respectively. Presence of G-to-A mutations and stop codon was independently associated with HIV-1 subtype non-B (P = 0.017). CONCLUSIONS These results show an independent association between the presence of G-to-A mutations and stop codons with HIV-1 subtype non-B and low proviral DNA that could be explained by the APOBEC3 footprints and restriction of DNA synthesis and integration. However, further investigations are needed to study the contribution of Vif amino acid variability among HIV-1 subtypes.
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Affiliation(s)
- Basma Abdi
- Sorbonne Université, INSERM, Institut Pierre Louis d'Epidémiologie et de Santé Publique (IPLESP UMRS 1136), AP-HP, Hôpital Pitié Salpêtrière, Laboratoire de virologie, Paris, France
| | - Sidonie Lambert-Niclot
- Sorbonne Université, INSERM, Institut Pierre Louis d'Epidémiologie et de Santé Publique (IPLESP UMRS 1136), AP-HP, Hôpital Saint Antoine, Laboratoire de virologie, Paris, France
| | - Marc Wirden
- Sorbonne Université, INSERM, Institut Pierre Louis d'Epidémiologie et de Santé Publique (IPLESP UMRS 1136), AP-HP, Hôpital Pitié Salpêtrière, Laboratoire de virologie, Paris, France
| | - Aude Jary
- Sorbonne Université, INSERM, Institut Pierre Louis d'Epidémiologie et de Santé Publique (IPLESP UMRS 1136), AP-HP, Hôpital Pitié Salpêtrière, Laboratoire de virologie, Paris, France
| | - Elisa Teyssou
- Sorbonne Université, INSERM, Institut Pierre Louis d'Epidémiologie et de Santé Publique (IPLESP UMRS 1136), AP-HP, Hôpital Pitié Salpêtrière, Laboratoire de virologie, Paris, France
| | - Sophie Sayon
- Sorbonne Université, INSERM, Institut Pierre Louis d'Epidémiologie et de Santé Publique (IPLESP UMRS 1136), AP-HP, Hôpital Pitié Salpêtrière, Laboratoire de virologie, Paris, France
| | - Romain Palich
- Sorbonne Université, INSERM, Institut Pierre Louis d'Epidémiologie et de Santé Publique (IPLESP UMRS 1136), AP-HP, Hôpital Pitié Salpêtrière, Service des Maladies Infectieuses, Paris, France
| | - Roland Tubiana
- Sorbonne Université, INSERM, Institut Pierre Louis d'Epidémiologie et de Santé Publique (IPLESP UMRS 1136), AP-HP, Hôpital Pitié Salpêtrière, Service des Maladies Infectieuses, Paris, France
| | - Anne Simon
- Sorbonne Université, INSERM, Institut Pierre Louis d'Epidémiologie et de Santé Publique (IPLESP UMRS 1136), AP-HP, Hôpital Pitié Salpêtrière, Service de Médecine Interne, Paris, France
| | - Marc-Antoine Valantin
- Sorbonne Université, INSERM, Institut Pierre Louis d'Epidémiologie et de Santé Publique (IPLESP UMRS 1136), AP-HP, Hôpital Pitié Salpêtrière, Service des Maladies Infectieuses, Paris, France
| | - Christine Katlama
- Sorbonne Université, INSERM, Institut Pierre Louis d'Epidémiologie et de Santé Publique (IPLESP UMRS 1136), AP-HP, Hôpital Pitié Salpêtrière, Service des Maladies Infectieuses, Paris, France
| | - Laurence Morand-Joubert
- Sorbonne Université, INSERM, Institut Pierre Louis d'Epidémiologie et de Santé Publique (IPLESP UMRS 1136), AP-HP, Hôpital Saint Antoine, Laboratoire de virologie, Paris, France
| | - Vincent Calvez
- Sorbonne Université, INSERM, Institut Pierre Louis d'Epidémiologie et de Santé Publique (IPLESP UMRS 1136), AP-HP, Hôpital Pitié Salpêtrière, Laboratoire de virologie, Paris, France
| | - Anne-Geneviève Marcelin
- Sorbonne Université, INSERM, Institut Pierre Louis d'Epidémiologie et de Santé Publique (IPLESP UMRS 1136), AP-HP, Hôpital Pitié Salpêtrière, Laboratoire de virologie, Paris, France
| | - Cathia Soulie
- Sorbonne Université, INSERM, Institut Pierre Louis d'Epidémiologie et de Santé Publique (IPLESP UMRS 1136), AP-HP, Hôpital Pitié Salpêtrière, Laboratoire de virologie, Paris, France
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13
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Sadeghpour S, Khodaee S, Rahnama M, Rahimi H, Ebrahimi D. Human APOBEC3 Variations and Viral Infection. Viruses 2021; 13:1366. [PMID: 34372572 PMCID: PMC8310219 DOI: 10.3390/v13071366] [Citation(s) in RCA: 36] [Impact Index Per Article: 9.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/11/2021] [Revised: 07/07/2021] [Accepted: 07/08/2021] [Indexed: 12/13/2022] Open
Abstract
Human APOBEC3 (apolipoprotein B mRNA-editing catalytic polypeptide-like 3) enzymes are capable of inhibiting a wide range of endogenous and exogenous viruses using deaminase and deaminase-independent mechanisms. These enzymes are essential components of our innate immune system, as evidenced by (a) their strong positive selection and expansion in primates, (b) the evolution of viral counter-defense mechanisms, such as proteasomal degradation mediated by HIV Vif, and (c) hypermutation and inactivation of a large number of integrated HIV-1 proviruses. Numerous APOBEC3 single nucleotide polymorphisms, haplotypes, and splice variants have been identified in humans. Several of these variants have been reported to be associated with differential antiviral immunity. This review focuses on the current knowledge in the field about these natural variations and their roles in infectious diseases.
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Affiliation(s)
- Shiva Sadeghpour
- Department of Biological Science, University of California Irvine, Irvine, CA 92697, USA;
| | - Saeideh Khodaee
- Department of Bioinformatics, Institute of Biochemistry and Biophysics, University of Tehran, Tehran 1417614335, Iran;
| | - Mostafa Rahnama
- Department of Plant Pathology, University of Kentucky, Lexington, KY 40546, USA;
| | - Hamzeh Rahimi
- Department of Molecular Medicine, Biotechnology Research Center, Pasteur Institute of Iran, Tehran 1316943551, Iran;
| | - Diako Ebrahimi
- Texas Biomedical Research Institute, San Antonio, TX 78227, USA
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14
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Hakata Y, Miyazawa M. Deaminase-Independent Mode of Antiretroviral Action in Human and Mouse APOBEC3 Proteins. Microorganisms 2020; 8:microorganisms8121976. [PMID: 33322756 PMCID: PMC7764128 DOI: 10.3390/microorganisms8121976] [Citation(s) in RCA: 16] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/08/2020] [Accepted: 12/09/2020] [Indexed: 02/06/2023] Open
Abstract
Apolipoprotein B mRNA editing enzyme, catalytic polypeptide-like 3 (APOBEC3) proteins (APOBEC3s) are deaminases that convert cytosines to uracils predominantly on a single-stranded DNA, and function as intrinsic restriction factors in the innate immune system to suppress replication of viruses (including retroviruses) and movement of retrotransposons. Enzymatic activity is supposed to be essential for the APOBEC3 antiviral function. However, it is not the only way that APOBEC3s exert their biological function. Since the discovery of human APOBEC3G as a restriction factor for HIV-1, the deaminase-independent mode of action has been observed. At present, it is apparent that both the deaminase-dependent and -independent pathways are tightly involved not only in combating viruses but also in human tumorigenesis. Although the deaminase-dependent pathway has been extensively characterized so far, understanding of the deaminase-independent pathway remains immature. Here, we review existing knowledge regarding the deaminase-independent antiretroviral functions of APOBEC3s and their molecular mechanisms. We also discuss the possible unidentified molecular mechanism for the deaminase-independent antiretroviral function mediated by mouse APOBEC3.
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Affiliation(s)
- Yoshiyuki Hakata
- Department of Immunology, Kindai University Faculty of Medicine, 377-2 Ohno-Higashi, Osaka-Sayama, Osaka 589-8511, Japan;
- Correspondence: ; Tel.: +81-72-367-7660
| | - Masaaki Miyazawa
- Department of Immunology, Kindai University Faculty of Medicine, 377-2 Ohno-Higashi, Osaka-Sayama, Osaka 589-8511, Japan;
- Kindai University Anti-Aging Center, 3-4-1 Kowakae, Higashiosaka, Osaka 577-8502, Japan
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15
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Mika T, Ladigan-Badura S, Maghnouj A, Mustafa B, Klein-Scory S, Baraniskin A, Döhring S, Fuchs I, Ehl S, Hahn SA, Schroers R. Altered T-Lymphocyte Biology Following High-Dose Melphalan and Autologous Stem Cell Transplantation With Implications for Adoptive T-Cell Therapy. Front Oncol 2020; 10:568056. [PMID: 33363008 PMCID: PMC7759611 DOI: 10.3389/fonc.2020.568056] [Citation(s) in RCA: 16] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/31/2020] [Accepted: 10/20/2020] [Indexed: 01/18/2023] Open
Abstract
In relapsed and refractory multiple myeloma (MM), adoptive cell therapies (ACT) including CAR-T-cells are under clinical investigation. However, relapse due to T-cell exhaustion or limited persistence is an obstacle. Before ACT are considered in MM, high-dose (HD) melphalan followed by autologous stem-cell transplantation (autoSCT) has been administered in most clinical situations. Yet, the impact of HD chemotherapy on T-cells in MM with respect to ACT is unclear. In this study, T-lymphocytes’ phenotypes, expansion properties, lentiviral transduction efficacy, and gene expression were examined with special respect to patients following HD melphalan. Significant impairment of T-cells’ expansion and transduction rates could be demonstrated. Expansion was diminished due to inherent disadvantages of the predominant T-cell phenotype but restored over time. The quantitative fraction of CD27−/CD28− T-cells before expansion was predictive of T-cell yield. Following autoSCT, the transduction efficacy was reduced by disturbed lentiviral genome integration. Moreover, an unfavorable T-cell phenotype after expansion was demonstrated. In initial analyses of CD107a degranulation impaired T-cell cytotoxicity was detected in one patient following melphalan and autoSCT. The findings of our study have potential implications regarding the time point of leukapheresis for CAR-T-cell manufacturing. Our results point to a preferred interval of more than 3 months until patients should undergo cell separation for CAR-T therapy in the specific situation post-HD melphalan/autoSCT. Monitoring of CD27−/CD28− T-cells, has the potential to influence clinical decision making before apheresis in MM.
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Affiliation(s)
- Thomas Mika
- Department of Medicine, Hematology and Oncology, Ruhr-University Bochum, Bochum, Germany.,Department of Molecular Gastrointestinal Oncology, Ruhr-University Bochum, Bochum, Germany
| | - Swetlana Ladigan-Badura
- Department of Medicine, Hematology and Oncology, Ruhr-University Bochum, Bochum, Germany.,Department of Molecular Gastrointestinal Oncology, Ruhr-University Bochum, Bochum, Germany
| | - Abdelouahid Maghnouj
- Department of Molecular Gastrointestinal Oncology, Ruhr-University Bochum, Bochum, Germany
| | - Bakr Mustafa
- Department of Molecular Gastrointestinal Oncology, Ruhr-University Bochum, Bochum, Germany
| | - Susanne Klein-Scory
- Department of Medicine, Hematology and Oncology, Ruhr-University Bochum, Bochum, Germany
| | - Alexander Baraniskin
- Department of Medicine, Hematology and Oncology, Ruhr-University Bochum, Bochum, Germany
| | - Sascha Döhring
- Department of Molecular Gastrointestinal Oncology, Ruhr-University Bochum, Bochum, Germany
| | - Ilka Fuchs
- Center for Chronic Immunodeficiency, Faculty of Medicine, University of Freiburg, Freiburg im Breisgau, Germany
| | - Stephan Ehl
- Center for Chronic Immunodeficiency, Faculty of Medicine, University of Freiburg, Freiburg im Breisgau, Germany
| | - Stephan A Hahn
- Department of Molecular Gastrointestinal Oncology, Ruhr-University Bochum, Bochum, Germany
| | - Roland Schroers
- Department of Medicine, Hematology and Oncology, Ruhr-University Bochum, Bochum, Germany
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16
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Jaguva Vasudevan AA, Balakrishnan K, Gertzen CGW, Borvető F, Zhang Z, Sangwiman A, Held U, Küstermann C, Banerjee S, Schumann GG, Häussinger D, Bravo IG, Gohlke H, Münk C. Loop 1 of APOBEC3C Regulates its Antiviral Activity against HIV-1. J Mol Biol 2020; 432:6200-6227. [PMID: 33068636 DOI: 10.1016/j.jmb.2020.10.014] [Citation(s) in RCA: 11] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/31/2020] [Revised: 10/09/2020] [Accepted: 10/09/2020] [Indexed: 01/10/2023]
Abstract
APOBEC3 deaminases (A3s) provide mammals with an anti-retroviral barrier by catalyzing dC-to-dU deamination on viral ssDNA. Within primates, A3s have undergone a complex evolution via gene duplications, fusions, arms race, and selection. Human APOBEC3C (hA3C) efficiently restricts the replication of viral infectivity factor (vif)-deficient Simian immunodeficiency virus (SIVΔvif), but for unknown reasons, it inhibits HIV-1Δvif only weakly. In catarrhines (Old World monkeys and apes), the A3C loop 1 displays the conserved amino acid pair WE, while the corresponding consensus sequence in A3F and A3D is the largely divergent pair RK, which is also the inferred ancestral sequence for the last common ancestor of A3C and of the C-terminal domains of A3D and A3F in primates. Here, we report that modifying the WE residues in hA3C loop 1 to RK leads to stronger interactions with substrate ssDNA, facilitating catalytic function, which results in a drastic increase in both deamination activity and in the ability to restrict HIV-1 and LINE-1 replication. Conversely, the modification hA3F_WE resulted only in a marginal decrease in HIV-1Δvif inhibition. We propose that the two series of ancestral gene duplications that generated A3C, A3D-CTD and A3F-CTD allowed neo/subfunctionalization: A3F-CTD maintained the ancestral RK residues in loop 1, while diversifying selection resulted in the RK → WE modification in Old World anthropoids' A3C, possibly allowing for novel substrate specificity and function.
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Affiliation(s)
- Ananda Ayyappan Jaguva Vasudevan
- Clinic for Gastroenterology, Hepatology, and Infectiology, Medical Faculty, Heinrich Heine University Düsseldorf, Düsseldorf, Germany.
| | - Kannan Balakrishnan
- Clinic for Gastroenterology, Hepatology, and Infectiology, Medical Faculty, Heinrich Heine University Düsseldorf, Düsseldorf, Germany; Department of Biochemistry, School of Life Sciences, University of Hyderabad, Gachibowli, Hyderabad, India
| | - Christoph G W Gertzen
- Institute for Pharmaceutical and Medicinal Chemistry, Heinrich Heine University Düsseldorf, Düsseldorf, Germany; John von Neumann Institute for Computing (NIC), Jülich Supercomputing Centre & Institute of Biological Information Processing (IBI-7: Structural Biochemistry), Forschungszentrum Jülich GmbH, Jülich, Germany; Center for Structural Studies (CSS), Heinrich Heine University Düsseldorf, Düsseldorf, Germany
| | - Fanni Borvető
- Centre National de la Recherche Scientifique, Laboratory MIVEGEC (CNRS, IRD, Uni Montpellier), Montpellier, France
| | - Zeli Zhang
- Clinic for Gastroenterology, Hepatology, and Infectiology, Medical Faculty, Heinrich Heine University Düsseldorf, Düsseldorf, Germany
| | - Anucha Sangwiman
- Clinic for Gastroenterology, Hepatology, and Infectiology, Medical Faculty, Heinrich Heine University Düsseldorf, Düsseldorf, Germany
| | - Ulrike Held
- Division of Medical Biotechnology, Paul-Ehrlich-Institute, Langen, Germany
| | | | - Sharmistha Banerjee
- Department of Biochemistry, School of Life Sciences, University of Hyderabad, Gachibowli, Hyderabad, India
| | - Gerald G Schumann
- Division of Medical Biotechnology, Paul-Ehrlich-Institute, Langen, Germany
| | - Dieter Häussinger
- Clinic for Gastroenterology, Hepatology, and Infectiology, Medical Faculty, Heinrich Heine University Düsseldorf, Düsseldorf, Germany
| | - Ignacio G Bravo
- Centre National de la Recherche Scientifique, Laboratory MIVEGEC (CNRS, IRD, Uni Montpellier), Montpellier, France
| | - Holger Gohlke
- Institute for Pharmaceutical and Medicinal Chemistry, Heinrich Heine University Düsseldorf, Düsseldorf, Germany; John von Neumann Institute for Computing (NIC), Jülich Supercomputing Centre & Institute of Biological Information Processing (IBI-7: Structural Biochemistry), Forschungszentrum Jülich GmbH, Jülich, Germany
| | - Carsten Münk
- Clinic for Gastroenterology, Hepatology, and Infectiology, Medical Faculty, Heinrich Heine University Düsseldorf, Düsseldorf, Germany.
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17
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Murine Leukemia Virus P50 Protein Counteracts APOBEC3 by Blocking Its Packaging. J Virol 2020; 94:JVI.00032-20. [PMID: 32641479 DOI: 10.1128/jvi.00032-20] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/09/2020] [Accepted: 07/01/2020] [Indexed: 11/20/2022] Open
Abstract
Apolipoprotein B editing enzyme, catalytic polypeptide 3 (APOBEC3) family members are cytidine deaminases that play important roles in intrinsic responses to retrovirus infection. Complex retroviruses like human immunodeficiency virus type 1 (HIV-1) encode the viral infectivity factor (Vif) protein to counteract APOBEC3 proteins. Vif induces degradation of APOBEC3G and other APOBEC3 proteins and thereby prevents their packaging into virions. It is not known if murine leukemia virus (MLV) encodes a Vif-like protein. Here, we show that the MLV P50 protein, produced from an alternatively spliced gag RNA, interacts with the C terminus of mouse APOBEC3 and prevents its packaging without causing its degradation. By infecting APOBEC3 knockout (KO) and wild-type (WT) mice with Friend or Moloney MLV P50-deficient viruses, we found that APOBEC3 restricts the mutant viruses more than WT viruses in vivo Replication of P50-mutant viruses in an APOBEC3-expressing stable cell line was also much slower than that of WT viruses, and overexpressing P50 in this cell line enhanced mutant virus replication. Thus, MLV encodes a protein, P50, that overcomes APOBEC3 restriction by preventing its packaging into virions.IMPORTANCE MLV has existed in mice for at least a million years, in spite of the existence of host restriction factors that block infection. Although MLV is considered a simple retrovirus compared to lentiviruses, it does encode proteins generated from alternatively spliced RNAs. Here, we show that P50, generated from an alternatively spliced RNA encoded in gag, counteracts APOBEC3 by blocking its packaging. MLV also encodes a protein, glycoGag, that increases capsid stability and limits APOBEC3 access to the reverse transcription complex (RTC). Thus, MLV has evolved multiple means of preventing APOBEC3 from blocking infection, explaining its survival as an infectious pathogen in mice.
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18
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Moreira DA, Lamarca AP, Soares RF, Coelho AMA, Furtado C, Scherer NM, Moreira MAM, Seuánez HN, Boroni M. Transcriptome of the Southern Muriqui Brachyteles arachnoides (Primates:Platyrrhini), a Critically Endangered New World Monkey: Evidence of Adaptive Evolution. Front Genet 2020; 11:831. [PMID: 32849820 PMCID: PMC7412869 DOI: 10.3389/fgene.2020.00831] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/16/2020] [Accepted: 07/09/2020] [Indexed: 12/03/2022] Open
Abstract
The southern muriqui (Brachyteles arachnoides) is the largest neotropical primate. This species is endemic to Brazil and is currently critically endangered due to its habitat destruction. The genetic basis underlying adaptive traits of New World monkeys has been a subject of interest to several investigators, with significant concern about genes related to the immune system. In the absence of a reference genome, RNA-seq and de novo transcriptome assembly have proved to be valuable genetic procedures for accessing gene sequences and testing evolutionary hypotheses. We present here a first report on the sequencing, assembly, annotation and adaptive selection analysis for thousands of transcripts of B. arachnoides from two different samples, corresponding to 13 different blood cells and fibroblasts. We assembled 284,283 transcripts with N50 of 2,940 bp, with a high rate of complete transcripts, with a median high scoring pair coverage of 88.2%, including low expressed transcripts, accounting for 72.3% of complete BUSCOs. We could predict and extract 81,400 coding sequences with 79.8% of significant BLAST hit against the Euarchontoglires SwissProt dataset. Of these 64,929 sequences, 34,084 were considered homologous to Supraprimate proteins, and of the remaining sequences (30,845), 94% were associated with a protein domain or a KEGG Orthology group, indicating potentially novel or specific protein-coding genes of B. arachnoides. We use the predicted protein sequences to perform a comparative analysis with 10 other primates. This analysis revealed, for the first time in an Atelid species, an expansion of APOBEC3G, extending this knowledge to all NWM families. Using a branch-site model, we searched for evidence of positive selection in 4,533 orthologous sets. This evolutionary analysis revealed 132 amino acid sites in 30 genes potentially evolving under positive selection, shedding light on primate genome evolution. These genes belonged to a wide variety of categories, including those encoding the innate immune system proteins (APOBEC3G, OAS2, and CEACAM1) among others related to the immune response. This work generated a set of thousands of complete sequences that can be used in other studies on molecular evolution and may help to unveil the evolution of primate genes. Still, further functional studies are required to provide an understanding of the underlying evolutionary forces modeling the primate genome.
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Affiliation(s)
- Daniel A Moreira
- Laboratory of Bioinformatics and Computational Biology, Division of Experimental and Translational Research, Brazilian National Cancer Institute (INCA), Rio de Janeiro, Brazil
| | - Alessandra P Lamarca
- Laboratory of Bioinformatics and Molecular Evolution, Department of Genetics, Institute of Biology, Federal University of Rio de Janeiro (UFRJ), Rio de Janeiro, Brazil
| | - Rafael Ferreira Soares
- Laboratory for Functional Genomics and Bioinformatics, Oswaldo Cruz Institute, Fiocruz, Rio de Janeiro, Brazil
| | - Ana M A Coelho
- Department of Genetics, Institute of Biology, Federal University of Rio de Janeiro (UFRJ), Rio de Janeiro, Brazil
| | - Carolina Furtado
- Genetics Program, Division of Experimental and Translational Research, Brazilian National Cancer Institute (INCA), Rio de Janeiro, Brazil
| | - Nicole M Scherer
- Laboratory of Bioinformatics and Computational Biology, Division of Experimental and Translational Research, Brazilian National Cancer Institute (INCA), Rio de Janeiro, Brazil
| | - Miguel A M Moreira
- Genetics Program, Division of Experimental and Translational Research, Brazilian National Cancer Institute (INCA), Rio de Janeiro, Brazil
| | - Hector N Seuánez
- Genetics Program, Division of Experimental and Translational Research, Brazilian National Cancer Institute (INCA), Rio de Janeiro, Brazil
| | - Mariana Boroni
- Laboratory of Bioinformatics and Computational Biology, Division of Experimental and Translational Research, Brazilian National Cancer Institute (INCA), Rio de Janeiro, Brazil
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19
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Delviks-Frankenberry KA, Desimmie BA, Pathak VK. Structural Insights into APOBEC3-Mediated Lentiviral Restriction. Viruses 2020; 12:E587. [PMID: 32471198 PMCID: PMC7354603 DOI: 10.3390/v12060587] [Citation(s) in RCA: 18] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/23/2020] [Revised: 05/22/2020] [Accepted: 05/24/2020] [Indexed: 01/18/2023] Open
Abstract
Mammals have developed clever adaptive and innate immune defense mechanisms to protect against invading bacterial and viral pathogens. Human innate immunity is continuously evolving to expand the repertoire of restriction factors and one such family of intrinsic restriction factors is the APOBEC3 (A3) family of cytidine deaminases. The coordinated expression of seven members of the A3 family of cytidine deaminases provides intrinsic immunity against numerous foreign infectious agents and protects the host from exogenous retroviruses and endogenous retroelements. Four members of the A3 proteins-A3G, A3F, A3H, and A3D-restrict HIV-1 in the absence of virion infectivity factor (Vif); their incorporation into progeny virions is a prerequisite for cytidine deaminase-dependent and -independent activities that inhibit viral replication in the host target cell. HIV-1 encodes Vif, an accessory protein that antagonizes A3 proteins by targeting them for polyubiquitination and subsequent proteasomal degradation in the virus producing cells. In this review, we summarize our current understanding of the role of human A3 proteins as barriers against HIV-1 infection, how Vif overcomes their antiviral activity, and highlight recent structural and functional insights into A3-mediated restriction of lentiviruses.
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Affiliation(s)
| | | | - Vinay K. Pathak
- Viral Mutation Section, HIV Dynamics and Replication Program, National Cancer Institute at Frederick, Frederick, MD 21702, USA; (K.A.D.-F.); (B.A.D.)
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20
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Delviks-Frankenberry KA, Ackerman D, Timberlake ND, Hamscher M, Nikolaitchik OA, Hu WS, Torbett BE, Pathak VK. Development of Lentiviral Vectors for HIV-1 Gene Therapy with Vif-Resistant APOBEC3G. MOLECULAR THERAPY. NUCLEIC ACIDS 2019; 18:1023-1038. [PMID: 31778955 PMCID: PMC6889484 DOI: 10.1016/j.omtn.2019.10.024] [Citation(s) in RCA: 15] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 08/11/2019] [Revised: 10/17/2019] [Accepted: 10/23/2019] [Indexed: 12/29/2022]
Abstract
Strategies to control HIV-1 replication without antiviral therapy are needed to achieve a functional cure. To exploit the innate antiviral function of restriction factor cytidine deaminase APOBEC3G (A3G), we developed self-activating lentiviral vectors that efficiently deliver HIV-1 Vif-resistant mutant A3G-D128K to target cells. To circumvent APOBEC3 expression in virus-producing cells, which diminishes virus infectivity, a vector containing two overlapping fragments of A3G-D128K was designed that maintained the gene in an inactive form in the virus-producer cells. However, during transduction of target cells, retroviral recombination between the direct repeats reconstituted an active A3G-D128K in 89%-98% of transduced cells. Lentiviral vectors that expressed A3G-D128K transduced CD34+ hematopoietic stem and progenitor cells with a high efficiency (>30%). A3G-D128K expression in T cell lines CEM, CEMSS, and PM1 potently inhibited spreading infection of several HIV-1 subtypes by C-to-U deamination leading to lethal G-to-A hypermutation and inhibition of reverse transcription. SIVmac239 and HIV-2 were not inhibited, since their Vifs degraded A3G-D128K. A3G-D128K expression in CEM cells potently suppressed HIV-1 replication for >3.5 months without detectable resistant virus, suggesting a high genetic barrier for the emergence of A3G-D128K resistance. Because of this, A3G-D128K expression in HIV-1 target cells is a potential anti-HIV gene therapy approach that could be combined with other therapies for the treatment and functional cure of HIV-1 infection.
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Affiliation(s)
- Krista A Delviks-Frankenberry
- Viral Mutation Section, HIV Dynamics and Replication Program, National Cancer Institute at Frederick, Frederick, MD 21702, USA
| | - Daniel Ackerman
- Viral Mutation Section, HIV Dynamics and Replication Program, National Cancer Institute at Frederick, Frederick, MD 21702, USA
| | | | - Maria Hamscher
- Viral Mutation Section, HIV Dynamics and Replication Program, National Cancer Institute at Frederick, Frederick, MD 21702, USA
| | - Olga A Nikolaitchik
- Viral Recombination Section, HIV Dynamics and Replication Program, National Cancer Institute at Frederick, Frederick, MD 21702, USA
| | - Wei-Shau Hu
- Viral Recombination Section, HIV Dynamics and Replication Program, National Cancer Institute at Frederick, Frederick, MD 21702, USA
| | | | - Vinay K Pathak
- Viral Mutation Section, HIV Dynamics and Replication Program, National Cancer Institute at Frederick, Frederick, MD 21702, USA.
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21
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Wang X, Liu J, Zhou L, Ho WZ. Morphine Withdrawal Enhances HIV Infection of Macrophages. Front Immunol 2019; 10:2601. [PMID: 31803178 PMCID: PMC6872497 DOI: 10.3389/fimmu.2019.02601] [Citation(s) in RCA: 9] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/10/2019] [Accepted: 10/21/2019] [Indexed: 12/29/2022] Open
Abstract
Opioid withdrawal recurs at high rates in opioid use disorder and compromises the immune system. In general, there are two types of opioid withdrawal: abrupt withdrawal (AW) and precipitated withdrawal (PW). In this study, we examined the effect of morphine AW or morphine PW on HIV infection of human blood monocyte-derived macrophages. We observed that both morphine AW and PW enhanced the susceptibility of macrophages to HIV infection. In addition, both AW and PW activated HIV replication in the latently infected myeloid cells (U1 and OM10.1). Investigation of mechanisms responsible for these observations showed that both AW and PW could inhibit the expression of multiple intracellular HIV inhibitory factors, including APOBE3G/F, SAMHD1, MX2, and HIV restriction microRNAs (miR-28, miR-125b, and miR-150) in macrophages. These findings provide additional evidence to support the notion that opioid use compromises the intracellular anti-HIV immunity and facilitates HIV infection and persistence in macrophages.
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Affiliation(s)
| | | | | | - Wen-Zhe Ho
- Department of Pathology and Laboratory Medicine, Lewis Katz School of Medicine, Temple University, Philadelphia, PA, United States
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Profiling of LINE-1-Related Genes in Hepatocellular Carcinoma. Int J Mol Sci 2019; 20:ijms20030645. [PMID: 30717368 PMCID: PMC6387036 DOI: 10.3390/ijms20030645] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/31/2018] [Revised: 01/26/2019] [Accepted: 01/29/2019] [Indexed: 12/14/2022] Open
Abstract
Hepatocellular carcinoma (HCC) is a prime public health concern that accounts for most of the primary liver malignancies in humans. The most common etiological factor of HCC is hepatitis B virus (HBV). Despite recent advances in treatment strategies, there has been little success in improving the survival of HCC patients. To develop a novel therapeutic approach, evaluation of a working hypothesis based on different viewpoints might be important. Long interspersed element 1 (L1) retrotransposons have been suggested to play a role in HCC. However, the molecular machineries that can modulate L1 biology in HBV-related HCC have not been well-evaluated. Here, we summarize the profiles of expression and/or activation status of L1-related genes in HBV-related HCC, and HBV- and HCC-related genes that may impact L1-mediated tumorigenesis. L1 restriction factors appear to be suppressed by HBV infection. Since some of the L1 restriction factors also limit HBV, these factors may be exhausted in HBV-infected cells, which causes de-suppression of L1. Several HBV- and HCC-related genes that interact with L1 can affect oncogenic processes. Thus, L1 may be a novel prime therapeutic target for HBV-related HCC. Studies in this area will provide insights into HCC and other types of cancers.
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Abstract
Pandemic HIV-1, a human lentivirus, is the result of zoonotic transmission of SIV from chimpanzees (SIVcpz). How SIVcpz established spread in humans after spillover is an outstanding question. Lentiviral cross-species transmissions are exceptionally rare events. Nevertheless, the chimpanzee and the gorilla were part of the transmission chains that resulted in sustained infections that evolved into HIV-1. Although many restriction factors can repress the early stages of lentiviral replication, others target replication during the late phases. In some cases, viruses incorporate host proteins that interfere with subsequent rounds of replication. Though limited and small, HIVs and SIVs, including SIVcpz can use their genome products to modulate and escape some of these barriers and thus establish a chronic infection.
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Affiliation(s)
- Augustin Penda Twizerimana
- Clinic for Gastroenterology, Hepatology & Infectiology, Medical Faculty, Heinrich-Heine-University, Moorenstr. 5, 40225 Düsseldorf, Germany
| | - Rachel Scheck
- Clinic for Gastroenterology, Hepatology & Infectiology, Medical Faculty, Heinrich-Heine-University, Moorenstr. 5, 40225 Düsseldorf, Germany
| | - Dieter Häussinger
- Clinic for Gastroenterology, Hepatology & Infectiology, Medical Faculty, Heinrich-Heine-University, Moorenstr. 5, 40225 Düsseldorf, Germany
| | - Carsten Münk
- Clinic for Gastroenterology, Hepatology & Infectiology, Medical Faculty, Heinrich-Heine-University, Moorenstr. 5, 40225 Düsseldorf, Germany
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Castro-Gonzalez S, Colomer-Lluch M, Serra-Moreno R. Barriers for HIV Cure: The Latent Reservoir. AIDS Res Hum Retroviruses 2018; 34:739-759. [PMID: 30056745 PMCID: PMC6152859 DOI: 10.1089/aid.2018.0118] [Citation(s) in RCA: 80] [Impact Index Per Article: 11.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/29/2022] Open
Abstract
Thirty-five years after the identification of HIV-1 as the causative agent of AIDS, we are still in search of vaccines and treatments to eradicate this devastating infectious disease. Progress has been made in understanding the molecular pathogenesis of this infection, which has been crucial for the development of the current therapy regimens. However, despite their efficacy at limiting active viral replication, these drugs are unable to purge the latent reservoir: a pool of cells that harbor transcriptionally inactive, but replication-competent HIV-1 proviruses, and that represent the main barrier to eradicate HIV-1 from affected individuals. In this review, we discuss advances in the field that have allowed a better understanding of HIV-1 latency, including the diverse cell types that constitute the latent reservoir, factors influencing latency, tools to study HIV-1 latency, as well as current and prospective therapeutic approaches to target these latently infected cells, so a functional cure for HIV/AIDS can become a reality.
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Affiliation(s)
- Sergio Castro-Gonzalez
- Department of Biological Sciences, College of Arts and Sciences, Texas Tech University, Lubbock, Texas
| | - Marta Colomer-Lluch
- IrsiCaixa AIDS Research Institute, Hospital Germans Trias i Pujol, Badalona, Spain
| | - Ruth Serra-Moreno
- Department of Biological Sciences, College of Arts and Sciences, Texas Tech University, Lubbock, Texas
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Borzooee F, Asgharpour M, Quinlan E, Grant MD, Larijani M. Viral subversion of APOBEC3s: Lessons for anti-tumor immunity and tumor immunotherapy. Int Rev Immunol 2018; 37:151-164. [PMID: 29211501 DOI: 10.1080/08830185.2017.1403596] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/28/2022]
Abstract
APOBEC3s (A3) are endogenous DNA-editing enzymes that are expressed in immune cells including T lymphocytes. A3s target and mutate the genomes of retroviruses that infect immune tissues such as the human immunodeficiency virus (HIV). Therefore, A3s were classically defined as host anti-viral innate immune factors. In contrast, we and others showed that A3s can also benefit the virus by mediating escape from adaptive immune recognition and drugs. Crucially, whether A3-mediated mutations help or hinder HIV, is not up to chance. Rather, the virus has evolved multiple mechanisms to actively and maximally subvert A3 activity. More recently, extensive A3 mutational footprints in tumor genomes have been observed in many different cancers. This suggests a role for A3s in cancer initiation and progression. On the other hand, multiple anti-tumor activities of A3s have also come to light, including impact on immune checkpoint molecules and possible generation of tumor neo-antigens. Here, we review the studies that reshaped the view of A3s from anti-viral innate immune agents to host factors exploited by HIV to escape from immune recognition. Viruses and tumors share many attributes, including rapid evolution and adeptness at exploiting mutations. Given this parallel, we then discuss the pro- and anti-tumor roles of A3s, and suggest that lessons learned from studying A3s in the context of anti-viral immunity can be applied to tumor immunotherapy.
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Affiliation(s)
- Faezeh Borzooee
- a Program in Immunology and Infectious Diseases, Division of Biomedical Sciences, Faculty of Medicine , Memorial University of Newfoundland , St. John's, Newfoundland A1B 3V6 , Canada
| | - Mahdi Asgharpour
- a Program in Immunology and Infectious Diseases, Division of Biomedical Sciences, Faculty of Medicine , Memorial University of Newfoundland , St. John's, Newfoundland A1B 3V6 , Canada
| | - Emma Quinlan
- a Program in Immunology and Infectious Diseases, Division of Biomedical Sciences, Faculty of Medicine , Memorial University of Newfoundland , St. John's, Newfoundland A1B 3V6 , Canada
| | - Michael D Grant
- a Program in Immunology and Infectious Diseases, Division of Biomedical Sciences, Faculty of Medicine , Memorial University of Newfoundland , St. John's, Newfoundland A1B 3V6 , Canada
| | - Mani Larijani
- a Program in Immunology and Infectious Diseases, Division of Biomedical Sciences, Faculty of Medicine , Memorial University of Newfoundland , St. John's, Newfoundland A1B 3V6 , Canada
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Feline APOBEC3s, Barriers to Cross-Species Transmission of FIV? Viruses 2018; 10:v10040186. [PMID: 29642583 PMCID: PMC5923480 DOI: 10.3390/v10040186] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/27/2018] [Revised: 04/06/2018] [Accepted: 04/07/2018] [Indexed: 02/07/2023] Open
Abstract
The replication of lentiviruses highly depends on host cellular factors, which defines their species-specific tropism. Cellular restriction factors that can inhibit lentiviral replication were recently identified. Feline immunodeficiency virus (FIV) was found to be sensitive to several feline cellular restriction factors, such as apolipoprotein B mRNA-editing enzyme, catalytic polypeptide-like 3 (APOBEC3) and tetherin, but FIV evolved to counteract them. Here, we describe the molecular mechanisms by which feline APOBEC3 restriction factors inhibit FIV replication and discuss the molecular interaction of APOBEC3 proteins with the viral antagonizing protein Vif. We speculate that feline APOBEC3 proteins could explain some of the observed FIV cross-species transmissions described in wild Felids.
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Human-APOBEC3G-dependent restriction of porcine endogenous retrovirus replication is mediated by cytidine deamination and inhibition of DNA strand transfer during reverse transcription. Arch Virol 2018; 163:1907-1914. [PMID: 29610985 DOI: 10.1007/s00705-018-3822-x] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/05/2017] [Accepted: 03/18/2018] [Indexed: 10/17/2022]
Abstract
Although human apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like 3G (APOBEC3G, hA3G)-mediated deamination is the major mechanism used to restrict the infectivity of a broad range of retroviruses, it is unclear whether porcine endogenous retrovirus (PERV) is affected by hA3G or porcine A3F (poA3F). To determine whether DNA deamination is required for hA3G- and poA3F-dependent inhibition of PERV transmission, we developed VSV-pseudotype PERV-B expressing hA3G, mutant hA3G-E67Q (encapsidation and RNA binding activity-deficient), mutant hA3G-E259Q (deaminase-deficient), or poA3F. hA3G-E67Q decreased virus infectivity by ~ 93% compared to the ~ 99% decrease of viral infectivity by wild-type hA3G, while hA3G-E259Q decreased the infectivity of PERV-B by ~ 35%. These data suggest that cytidine deamination activity is crucial for efficient restriction of PERV by hA3G, but cytidine deamination cannot fully explain the inactivation of PERV by hA3G. Furthermore, differential DNA denaturation PCR (3D-PCR) products from 293T cells infected with PERV-B expressing hA3G mutants were sequenced. G-to-A hypermutation was detected at a frequency of 4.1% for hA3G, 3.4% for hA3G-E67Q, and 4.7% for poA3F. These results also suggest that hA3G and poA3F inhibit PERV by a deamination-dependent mechanism. To examine the effect of hA3G on the production of PERV DNA, genomic DNA was extracted from 293T cells 12 h after infection with PERV expressing hA3G, and this DNA was used as template for real-time PCR. A 50% decrease in minus strand strong stop (-sss) DNA synthesis/transfer was observed in the presence of hA3G. Based on these results, we conclude that hA3G may restrict PERV by both deamination-dependent mechanisms and inhibition of DNA strand transfer during PERV reverse transcription.
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Guo L, Xu XQ, Zhou L, Zhou RH, Wang X, Li JL, Liu JB, Liu H, Zhang B, Ho WZ. Human Intestinal Epithelial Cells Release Antiviral Factors That Inhibit HIV Infection of Macrophages. Front Immunol 2018. [PMID: 29515574 PMCID: PMC5825896 DOI: 10.3389/fimmu.2018.00247] [Citation(s) in RCA: 30] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/07/2023] Open
Abstract
As a rich source of CD4+ T cells and macrophages, the gastrointestinal (GI) tract is a major target site for HIV infection. The interplay between GI-resident macrophages and intestinal epithelial cells (IECs) constitutes an important element of GI innate immunity against pathogens. In this study, we investigated whether human IECs have the ability to produce antiviral factors that can inhibit HIV infection of macrophages. We demonstrated that IECs possess functional toll-like receptor 3 (TLR3), the activation of which resulted in induction of key interferon (IFN) regulatory factors (IRF3 and IRF7), IFN-β, IFN-λ, and CC chemokines (MIP-1α, MIP-1β, RANTES), the ligands of HIV entry co-receptor CCR5. In addition, TLR3-activated IECs release exosomes that contained the anti-HIV factors, including IFN-stimulated genes (ISGs: ISG15, ISG56, MxB, OAS-1, GBP5, and Viperin) and HIV restriction miRNAs (miRNA-17, miRNA-20, miRNA-28, miRNA-29 family members, and miRNA-125b). Importantly, treatment of macrophages with supernatant (SN) from the activated IEC cultures inhibited HIV replication. Further studies showed that IEC SN could also induce the expression of antiviral ISGs and cellular HIV restriction factors (Tetherin and APOBEC3G/3F) in HIV-infected macrophages. These findings indicated that IECs might act as an important element in GI innate immunity against HIV infection/replication.
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Affiliation(s)
- Le Guo
- Wuhan University School of Basic Medical Sciences, Wuhan, China
| | - Xi-Qiu Xu
- Wuhan University School of Basic Medical Sciences, Wuhan, China
| | - Li Zhou
- Wuhan University School of Basic Medical Sciences, Wuhan, China
| | - Run-Hong Zhou
- Wuhan University School of Basic Medical Sciences, Wuhan, China
| | - Xu Wang
- Department of Pathology and Laboratory Medicine, Lewis Katz School of Medicine, Temple University, Philadelphia, PA, United States
| | - Jie-Liang Li
- Department of Pathology and Laboratory Medicine, Lewis Katz School of Medicine, Temple University, Philadelphia, PA, United States
| | - Jin-Biao Liu
- Wuhan University School of Basic Medical Sciences, Wuhan, China
| | - Hang Liu
- Wuhan University School of Basic Medical Sciences, Wuhan, China
| | - Biao Zhang
- Wuhan University School of Basic Medical Sciences, Wuhan, China
| | - Wen-Zhe Ho
- Wuhan University School of Basic Medical Sciences, Wuhan, China.,Department of Pathology and Laboratory Medicine, Lewis Katz School of Medicine, Temple University, Philadelphia, PA, United States
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29
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Pollpeter D, Parsons M, Sobala AE, Coxhead S, Lang RD, Bruns AM, Papaioannou S, McDonnell JM, Apolonia L, Chowdhury JA, Horvath CM, Malim MH. Deep sequencing of HIV-1 reverse transcripts reveals the multifaceted antiviral functions of APOBEC3G. Nat Microbiol 2018; 3:220-233. [PMID: 29158605 PMCID: PMC6014619 DOI: 10.1038/s41564-017-0063-9] [Citation(s) in RCA: 78] [Impact Index Per Article: 11.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/09/2017] [Accepted: 10/10/2017] [Indexed: 12/15/2022]
Abstract
Following cell entry, the RNA genome of HIV-1 is reverse transcribed into double-stranded DNA that ultimately integrates into the host-cell genome to establish the provirus. These early phases of infection are notably vulnerable to suppression by a collection of cellular antiviral effectors, called restriction or resistance factors. The host antiviral protein APOBEC3G (A3G) antagonizes the early steps of HIV-1 infection through the combined effects of inhibiting viral cDNA production and cytidine-to-uridine-driven hypermutation of this cDNA. In seeking to address the underlying molecular mechanism for inhibited cDNA synthesis, we developed a deep sequencing strategy to characterize nascent reverse transcription products and their precise 3'-termini in HIV-1 infected T cells. Our results demonstrate site- and sequence-independent interference with reverse transcription, which requires the specific interaction of A3G with reverse transcriptase itself. This approach also established, contrary to current ideas, that cellular uracil base excision repair (UBER) enzymes target and cleave A3G-edited uridine-containing viral cDNA. Together, these findings yield further insights into the regulatory interplay between reverse transcriptase, A3G and cellular DNA repair machinery, and identify the suppression of HIV-1 reverse transcriptase by a directly interacting host protein as a new cell-mediated antiviral mechanism.
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Affiliation(s)
- Darja Pollpeter
- Department of Infectious Diseases, King's College London, London, UK
| | - Maddy Parsons
- Randall Division of Cell and Molecular Biophysics, King's College London, London, UK
| | - Andrew E Sobala
- Department of Infectious Diseases, King's College London, London, UK
| | - Sashika Coxhead
- Department of Infectious Diseases, King's College London, London, UK
| | - Rupert D Lang
- Department of Infectious Diseases, King's College London, London, UK
| | - Annie M Bruns
- Department of Molecular Biosciences, Northwestern University, Evanston, IL, USA
| | | | - James M McDonnell
- Randall Division of Cell and Molecular Biophysics, King's College London, London, UK
| | - Luis Apolonia
- Department of Infectious Diseases, King's College London, London, UK
| | - Jamil A Chowdhury
- Department of Infectious Diseases, King's College London, London, UK
| | - Curt M Horvath
- Department of Molecular Biosciences, Northwestern University, Evanston, IL, USA
| | - Michael H Malim
- Department of Infectious Diseases, King's College London, London, UK.
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30
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Multiple Inhibitory Factors Act in the Late Phase of HIV-1 Replication: a Systematic Review of the Literature. Microbiol Mol Biol Rev 2018; 82:82/1/e00051-17. [PMID: 29321222 DOI: 10.1128/mmbr.00051-17] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
The use of lentiviral vectors for therapeutic purposes has shown promising results in clinical trials. The ability to produce a clinical-grade vector at high yields remains a critical issue. One possible obstacle could be cellular factors known to inhibit human immunodeficiency virus (HIV). To date, five HIV restriction factors have been identified, although it is likely that more factors are involved in the complex HIV-cell interaction. Inhibitory factors that have an adverse effect but do not abolish virus production are much less well described. Therefore, a gap exists in the knowledge of inhibitory factors acting late in the HIV life cycle (from transcription to infection of a new cell), which are relevant to the lentiviral vector production process. The objective was to review the HIV literature to identify cellular factors previously implicated as inhibitors of the late stages of lentivirus production. A search for publications was conducted on MEDLINE via the PubMed interface, using the keyword sequence "HIV restriction factor" or "HIV restriction" or "inhibit HIV" or "repress HIV" or "restrict HIV" or "suppress HIV" or "block HIV," with a publication date up to 31 December 2016. Cited papers from the identified records were investigated, and additional database searches were performed. A total of 260 candidate inhibitory factors were identified. These factors have been identified in the literature as having a negative impact on HIV replication. This study identified hundreds of candidate inhibitory factors for which the impact of modulating their expression in lentiviral vector production could be beneficial.
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31
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Iqbal K, Imran M, Ullah S, Jamal M, Waheed Y, Ali Q. Correlation of Apolipoprotein B mRNA-editing Enzyme, Catalytic Polypeptide- like 3G Genetic Variant rs8177832 with HIV-1 Predisposition in Pakistani Population. Curr HIV Res 2018; 16:297-301. [PMID: 30338740 PMCID: PMC6416456 DOI: 10.2174/1570162x16666181018155827] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/21/2018] [Revised: 10/08/2018] [Accepted: 10/12/2018] [Indexed: 02/07/2023]
Abstract
BACKGROUND Human immunodeficiency virus (HIV) infection is a global health burden which ultimately results in acquired immune deficiency syndrome (AIDS). There are multiple host factors which are capable of limiting HIV-1 replication. One of the most important host factors which inhibit HIV-1 DNA synthesis is the apolipoprotein B mRNA-editing enzyme, catalytic polypeptide- like 3G (APOBEC3G). Any genetic variation of this important host factor may influence the host susceptibility to viral infection. OBJECTIVE The aim of the current study was to evaluate any correlation of APOBEC3G genetic variation rs8177832 with HIV-1 infection. METHODS The study involved 142 healthy control and 100 HIV-1 infected subjects. The genetic variation rs8177832 of all studied subjects was determined by allele-specific polymerase chain reaction (AS-PCR). RESULTS The results showed that the distribution of rs8177832 genotypes AA, AG and GG in healthy subjects and HIV-1 subjects was; 42.253%, 42.957%, 14.788% and 66%, 27%, 7% respectively. Statistical analyses of data showed that there was a significant variation in rs8177832 genotype AA in healthy control and HIV-1 infected subjects (42.257% vs 66%; p-value<0.001). CONCLUSION Thus it was concluded that APOBEC3G rs8177832 AA genotype contributes in genetic predisposition to HIV-1 infection in Pakistani population.
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Affiliation(s)
- Khurshid Iqbal
- Address correspondence to this author at the Department of Medical Laboratory Sciences, Imperial College of Business Studies, Lahore, Pakistan; Tel: 00923028051657; E-mail:
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Canonical and Non-Canonical Autophagy in HIV-1 Replication Cycle. Viruses 2017; 9:v9100270. [PMID: 28946621 PMCID: PMC5691622 DOI: 10.3390/v9100270] [Citation(s) in RCA: 31] [Impact Index Per Article: 3.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/22/2017] [Revised: 09/19/2017] [Accepted: 09/21/2017] [Indexed: 12/11/2022] Open
Abstract
Autophagy is a lysosomal-dependent degradative process essential for maintaining cellular homeostasis, and is a key player in innate and adaptive immune responses to intracellular pathogens such as human immunodeficiency virus type 1 (HIV-1). In HIV-1 target cells, autophagy mechanisms can (i) selectively direct viral proteins and viruses for degradation; (ii) participate in the processing and presentation of viral-derived antigens through major histocompatibility complexes; and (iii) contribute to interferon production in response to HIV-1 infection. As a consequence, HIV-1 has evolved different strategies to finely regulate the autophagy pathway to favor its replication and dissemination. HIV-1 notably encodes accessory genes encoding Tat, Nef and Vpu proteins, which are able to perturb and hijack canonical and non-canonical autophagy mechanisms. This review outlines the current knowledge on the complex interplay between autophagy and HIV-1 replication cycle, providing an overview of the autophagy-mediated molecular processes deployed both by infected cells to combat the virus and by HIV-1 to evade antiviral response.
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Dynamics and regulation of nuclear import and nuclear movements of HIV-1 complexes. PLoS Pathog 2017; 13:e1006570. [PMID: 28827840 PMCID: PMC5578721 DOI: 10.1371/journal.ppat.1006570] [Citation(s) in RCA: 93] [Impact Index Per Article: 11.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/23/2017] [Revised: 08/31/2017] [Accepted: 08/05/2017] [Indexed: 11/19/2022] Open
Abstract
The dynamics and regulation of HIV-1 nuclear import and its intranuclear movements after import have not been studied. To elucidate these essential HIV-1 post-entry events, we labeled viral complexes with two fluorescently tagged virion-incorporated proteins (APOBEC3F or integrase), and analyzed the HIV-1 dynamics of nuclear envelope (NE) docking, nuclear import, and intranuclear movements in living cells. We observed that HIV-1 complexes exhibit unusually long NE residence times (1.5±1.6 hrs) compared to most cellular cargos, which are imported into the nuclei within milliseconds. Furthermore, nuclear import requires HIV-1 capsid (CA) and nuclear pore protein Nup358, and results in significant loss of CA, indicating that one of the viral core uncoating steps occurs during nuclear import. Our results showed that the CA-Cyclophilin A interaction regulates the dynamics of nuclear import by delaying the time of NE docking as well as transport through the nuclear pore, but blocking reverse transcription has no effect on the kinetics of nuclear import. We also visualized the translocation of viral complexes docked at the NE into the nucleus and analyzed their nuclear movements and determined that viral complexes exhibited a brief fast phase (<9 min), followed by a long slow phase lasting several hours. A comparison of the movement of viral complexes to those of proviral transcription sites supports the hypothesis that HIV-1 complexes quickly tether to chromatin at or near their sites of integration in both wild-type cells and cells in which LEDGF/p75 was deleted using CRISPR/cas9, indicating that the tethering interactions do not require LEDGF/p75. These studies provide novel insights into the dynamics of viral complex-NE association, regulation of nuclear import, viral core uncoating, and intranuclear movements that precede integration site selection. Although nuclear import of HIV-1 is essential for viral replication, many aspects of this process are currently unknown. Here, we defined the dynamics of HIV-1 nuclear envelope (NE) docking, nuclear import and its relationship to viral core uncoating, and intranuclear movements. We observed that HIV-1 complexes exhibit an unusually long residence time at the NE (∼1.5 hrs) compared to other cellular and viral cargos, and that HIV-1 capsid (CA) and host nuclear pore protein Nup358 are required for NE docking and nuclear import. Soon after import, the viral complexes exhibit a brief fast phase of movement, followed by a long slow phase, during which their movement is similar to that of integrated proviruses, suggesting that they rapidly become tethered to chromatin through interactions that do not require LEDGF/p75. Importantly, we found that NE association and nuclear import is regulated by the CA-cyclophilin A interaction, but not reverse transcription, and that one of the viral core uncoating steps, characterized by substantial loss of CA, occurs concurrently with nuclear import.
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Konstantoulas CJ, Hagen B, Indik S. Moderate sensitivity of mouse mammary tumour virus to inhibition by human APOBEC3G. J Gen Virol 2017; 98:2362-2367. [PMID: 28809145 DOI: 10.1099/jgv.0.000897] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/18/2022] Open
Abstract
Infectivity of the mouse mammary tumour virus (MMTV) is inhibited by mouse APOBEC3 (mA3) which is efficiently packaged into virions. As the inhibition is only partial, the virus can replicate in tissues expressing mA3 and complete its replication cycle. Here, we have examined the sensitivity of MMTV to inhibition by a human orthologue of mA3, A3G. We report that the virus containing A3G is only moderately susceptible to inhibition by the human factor. Whereas the vif-deficient HIV-1 vector produced in human epithelial cells expressing endogenous levels of A3G was efficiently inhibited, an MMTV vector remained fully infectious. Greater A3G expression levels were necessary to restrict infectivity of MMTV, but only when the factor retained its deaminase activity. Furthermore, the spreading kinetic of a replication competent MMTV was only moderately accelerated in cells with downmodulated A3G expression. These data suggest that MMTV has evolved a mechanism to neutralize antiviral activity of APOBEC3 proteins.
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Affiliation(s)
| | - Benedikt Hagen
- Institute of Virology, University of Veterinary Medicine, Veterinaerplatz 1, 1210 Vienna, Austria
| | - Stanislav Indik
- Institute of Virology, University of Veterinary Medicine, Veterinaerplatz 1, 1210 Vienna, Austria
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35
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Jaguva Vasudevan AA, Hofmann H, Willbold D, Häussinger D, Koenig BW, Münk C. Enhancing the Catalytic Deamination Activity of APOBEC3C Is Insufficient to Inhibit Vif-Deficient HIV-1. J Mol Biol 2017; 429:1171-1191. [DOI: 10.1016/j.jmb.2017.03.015] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/16/2016] [Revised: 03/08/2017] [Accepted: 03/08/2017] [Indexed: 12/17/2022]
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36
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Ara A, Love RP, Follack TB, Ahmed KA, Adolph MB, Chelico L. Mechanism of Enhanced HIV Restriction by Virion Coencapsidated Cytidine Deaminases APOBEC3F and APOBEC3G. J Virol 2017; 91:e02230-16. [PMID: 27881650 PMCID: PMC5244329 DOI: 10.1128/jvi.02230-16] [Citation(s) in RCA: 27] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/15/2016] [Accepted: 11/16/2016] [Indexed: 12/21/2022] Open
Abstract
The APOBEC3 (A3) enzymes, A3G and A3F, are coordinately expressed in CD4+ T cells and can become coencapsidated into HIV-1 virions, primarily in the absence of the viral infectivity factor (Vif). A3F and A3G are deoxycytidine deaminases that inhibit HIV-1 replication by inducing guanine-to-adenine hypermutation through deamination of cytosine to form uracil in minus-strand DNA. The effect of the simultaneous presence of both A3G and A3F on HIV-1 restriction ability is not clear. Here, we used a single-cycle infectivity assay and biochemical analyses to determine if coencapsidated A3G and A3F differ in their restriction capacity from A3G or A3F alone. Proviral DNA sequencing demonstrated that compared to each A3 enzyme alone, A3G and A3F, when combined, had a coordinate effect on hypermutation. Using size exclusion chromatography, rotational anisotropy, and in vitro deamination assays, we demonstrate that A3F promotes A3G deamination activity by forming an A3F/G hetero-oligomer in the absence of RNA which is more efficient at deaminating cytosines. Further, A3F caused the accumulation of shorter reverse transcripts due to decreasing reverse transcriptase efficiency, which would leave single-stranded minus-strand DNA exposed for longer periods of time, enabling more deamination events to occur. Although A3G and A3F are known to function alongside each other, these data provide evidence for an A3F/G hetero-oligomeric A3 with unique properties compared to each individual counterpart. IMPORTANCE The APOBEC3 enzymes APOBEC3F and APOBEC3G act as a barrier to HIV-1 replication in the absence of the HIV-1 Vif protein. After APOBEC3 enzymes are encapsidated into virions, they deaminate cytosines in minus-strand DNA, which forms promutagenic uracils that induce transition mutations or proviral DNA degradation. Even in the presence of Vif, footprints of APOBEC3-catalyzed deaminations are found, demonstrating that APOBEC3s still have discernible activity against HIV-1 in infected individuals. We undertook a study to better understand the activity of coexpressed APOBEC3F and APOBEC3G. The data demonstrate that an APOBEC3F/APOBEC3G hetero-oligomer can form that has unique properties compared to each APOBEC3 alone. This hetero-oligomer has increased efficiency of virus hypermutation, raising the idea that we still may not fully realize the antiviral mechanisms of endogenous APOBEC3 enzymes. Hetero-oligomerization may be a mechanism to increase their antiviral activity in the presence of Vif.
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Affiliation(s)
- Anjuman Ara
- University of Saskatchewan, Microbiology and Immunology, College of Medicine, Saskatoon, Saskatchewan, Canada
| | - Robin P Love
- University of Saskatchewan, Microbiology and Immunology, College of Medicine, Saskatoon, Saskatchewan, Canada
| | - Tyson B Follack
- University of Saskatchewan, Microbiology and Immunology, College of Medicine, Saskatoon, Saskatchewan, Canada
| | - Khawaja A Ahmed
- University of Saskatchewan, Microbiology and Immunology, College of Medicine, Saskatoon, Saskatchewan, Canada
| | - Madison B Adolph
- University of Saskatchewan, Microbiology and Immunology, College of Medicine, Saskatoon, Saskatchewan, Canada
| | - Linda Chelico
- University of Saskatchewan, Microbiology and Immunology, College of Medicine, Saskatoon, Saskatchewan, Canada
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Translational regulation of APOBEC3G mRNA by Vif requires its 5'UTR and contributes to restoring HIV-1 infectivity. Sci Rep 2016; 6:39507. [PMID: 27996044 PMCID: PMC5171582 DOI: 10.1038/srep39507] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/22/2016] [Accepted: 11/23/2016] [Indexed: 12/12/2022] Open
Abstract
The essential HIV-1 viral infectivity factor (Vif) allows productive infection of non-permissive cells expressing cytidine deaminases APOBEC3G (A3G) and A3F by decreasing their cellular level, and preventing their incorporation into virions. Unlike the Vif-induced degradation of A3G, the functional role of the inhibition of A3G translation by Vif remained unclear. Here, we show that two stem-loop structures within the 5′-untranslated region of A3G mRNA are crucial for translation inhibition by Vif in cells, and most Vif alleles neutralize A3G translation efficiently. Interestingly, K26R mutation in Vif abolishes degradation of A3G by the proteasome but has no effect at the translational level, indicating these two pathways are independent. These two mechanisms, proteasomal degradation and translational inhibition, similarly contribute to decrease the cellular level of A3G by Vif and to prevent its incorporation into virions. Importantly, inhibition of A3G translation is sufficient to partially restore viral infectivity in the absence of proteosomal degradation. These findings demonstrate that HIV-1 has evolved redundant mechanisms to specifically inhibit the potent antiviral activity of A3G.
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Okada A, Iwatani Y. APOBEC3G-Mediated G-to-A Hypermutation of the HIV-1 Genome: The Missing Link in Antiviral Molecular Mechanisms. Front Microbiol 2016; 7:2027. [PMID: 28066353 PMCID: PMC5165236 DOI: 10.3389/fmicb.2016.02027] [Citation(s) in RCA: 38] [Impact Index Per Article: 4.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/03/2016] [Accepted: 12/02/2016] [Indexed: 12/20/2022] Open
Abstract
APOBEC3G (A3G) is a member of the cellular polynucleotide cytidine deaminases, which catalyze the deamination of cytosine (dC) to uracil (dU) in single-stranded DNA. These enzymes potently inhibit the replication of a variety of retroviruses and retrotransposons, including HIV-1. A3G is incorporated into vif-deficient HIV-1 virions and targets viral reverse transcripts, particularly minus-stranded DNA products, in newly infected cells. It is well established that the enzymatic activity of A3G is closely correlated with the potential to greatly inhibit HIV-1 replication in the absence of Vif. However, the details of the underlying molecular mechanisms are not fully understood. One potential mechanism of A3G antiviral activity is that the A3G-dependent deamination may trigger degradation of the dU-containing reverse transcripts by cellular uracil DNA glycosylases (UDGs). More recently, another mechanism has been suggested, in which the virion-incorporated A3G generates lethal levels of the G-to-A hypermutation in the viral DNA genome, thus potentially driving the viruses into “error catastrophe” mode. In this mini review article, we summarize the deaminase-dependent and deaminase-independent molecular mechanisms of A3G and discuss how A3G-mediated deamination is linked to antiviral mechanisms.
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Affiliation(s)
- Ayaka Okada
- Department of Microbiology and Immunology, Laboratory of Infectious Diseases, Clinical Research Center, National Hospital Organization Nagoya Medical Center Nagoya, Japan
| | - Yasumasa Iwatani
- Department of Microbiology and Immunology, Laboratory of Infectious Diseases, Clinical Research Center, National Hospital Organization Nagoya Medical CenterNagoya, Japan; Department of AIDS Research, Nagoya University Graduate School of MedicineNagoya, Japan
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Feline Immunodeficiency Virus Vif N-Terminal Residues Selectively Counteract Feline APOBEC3s. J Virol 2016; 90:10545-10557. [PMID: 27630243 DOI: 10.1128/jvi.01593-16] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/11/2016] [Accepted: 09/10/2016] [Indexed: 01/14/2023] Open
Abstract
Feline immunodeficiency virus (FIV) Vif protein counteracts feline APOBEC3s (FcaA3s) restriction factors by inducing their proteasomal degradation. The functional domains in FIV Vif for interaction with FcaA3s are poorly understood. Here, we have identified several motifs in FIV Vif that are important for selective degradation of different FcaA3s. Cats (Felis catus) express three types of A3s: single-domain A3Z2, single-domain A3Z3, and double-domain A3Z2Z3. We proposed that FIV Vif would selectively interact with the Z2 and the Z3 A3s. Indeed, we identified two N-terminal Vif motifs (12LF13 and 18GG19) that specifically interacted with the FcaA3Z2 protein but not with A3Z3. In contrast, the exclusive degradation of FcaA3Z3 was regulated by a region of three residues (M24, L25, and I27). Only a FIV Vif carrying a combination of mutations from both interaction sites lost the capacity to degrade and counteract FcaA3Z2Z3. However, alterations in the specific A3s interaction sites did not affect the cellular localization of the FIV Vif protein and binding to feline A3s. Pulldown experiments demonstrated that the A3 binding region localized to FIV Vif residues 50 to 80, outside the specific A3 interaction domain. Finally, we found that the Vif sites specific to individual A3s are conserved in several FIV lineages of domestic cat and nondomestic cats, while being absent in the FIV Vif of pumas. Our data support a complex model of multiple Vif-A3 interactions in which the specific region for selective A3 counteraction is discrete from a general A3 binding domain. IMPORTANCE Both human immunodeficiency virus (HIV) and feline immunodeficiency virus (FIV) Vif proteins counteract their host's APOBEC3 restriction factors. However, these two Vif proteins have limited sequence homology. The molecular interaction between FIV Vif and feline APOBEC3s are not well understood. Here, we identified N-terminal FIV Vif sites that regulate the selective interaction of Vif with either feline APOBEC3Z2 or APOBEC3Z3. These specific Vif sites are conserved in several FIV lineages of domestic cat and nondomestic cats, while being absent in FIV Vif from puma. Our findings provide important insights for future experiments describing the FIV Vif interaction with feline APOBEC3s and also indicate that the conserved feline APOBEC3s interaction sites of FIV Vif allow FIV transmissions in Felidae.
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Vif Proteins from Diverse Human Immunodeficiency Virus/Simian Immunodeficiency Virus Lineages Have Distinct Binding Sites in A3C. J Virol 2016; 90:10193-10208. [PMID: 27581978 DOI: 10.1128/jvi.01497-16] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/27/2016] [Accepted: 08/25/2016] [Indexed: 01/01/2023] Open
Abstract
Lentiviruses have evolved the Vif protein to counteract APOBEC3 (A3) restriction factors by targeting them for proteasomal degradation. Previous studies have identified important residues in the interface of human immunodeficiency virus type 1 (HIV-1) Vif and human APOBEC3C (hA3C) or human APOBEC3F (hA3F). However, the interaction between primate A3C proteins and HIV-1 Vif or natural HIV-1 Vif variants is still poorly understood. Here, we report that HIV-1 Vif is inactive against A3Cs of rhesus macaques (rhA3C), sooty mangabey monkeys (smmA3C), and African green monkeys (agmA3C), while HIV-2, African green monkey simian immunodeficiency virus (SIVagm), and SIVmac Vif proteins efficiently mediate the depletion of all tested A3Cs. We identified that residues N/H130 and Q133 in rhA3C and smmA3C are determinants for this HIV-1 Vif-triggered counteraction. We also found that the HIV-1 Vif interaction sites in helix 4 of hA3C and hA3F differ. Vif alleles from diverse HIV-1 subtypes were tested for degradation activities related to hA3C. The subtype F-1 Vif was identified to be inactive for degradation of hA3C and hA3F. The residues that determined F-1 Vif inactivity in the degradation of A3C/A3F were located in the C-terminal region (K167 and D182). Structural analysis of F-1 Vif revealed that impairing the internal salt bridge of E171-K167 restored reduction capacities to A3C/A3F. Furthermore, we found that D101 could also form an internal interaction with K167. Replacing D101 with glycine and R167 with lysine in NL4-3 Vif impaired its counteractivity to A3F and A3C. This finding indicates that internal interactions outside the A3 binding region in HIV-1 Vif influence the capacity to induce degradation of A3C/A3F. IMPORTANCE The APOBEC3 restriction factors can serve as potential barriers to lentiviral cross-species transmissions. Vif proteins from lentiviruses counteract APOBEC3 by proteasomal degradation. In this study, we found that monkey-derived A3C, rhA3C and smmA3C, were resistant to HIV-1 Vif. This was determined by A3C residues N/H130 and Q133. However, HIV-2, SIVagm, and SIVmac Vif proteins were found to be able to mediate the depletion of all tested primate A3C proteins. In addition, we identified a natural HIV-1 Vif (F-1 Vif) that was inactive in the degradation of hA3C/hA3F. Here, we provide for the first time a model that explains how an internal salt bridge of E171-K167-D101 influences Vif-mediated degradation of hA3C/hA3F. This finding provides a novel way to develop HIV-1 inhibitors by targeting the internal interactions of the Vif protein.
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Zhang Z, Gu Q, Jaguva Vasudevan AA, Hain A, Kloke BP, Hasheminasab S, Mulnaes D, Sato K, Cichutek K, Häussinger D, Bravo IG, Smits SHJ, Gohlke H, Münk C. Determinants of FIV and HIV Vif sensitivity of feline APOBEC3 restriction factors. Retrovirology 2016; 13:46. [PMID: 27368163 PMCID: PMC4930625 DOI: 10.1186/s12977-016-0274-9] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/08/2016] [Accepted: 06/09/2016] [Indexed: 02/07/2023] Open
Abstract
Background Feline immunodeficiency virus (FIV) is a global pathogen of Felidae species and a model system for Human immunodeficiency virus (HIV)-induced AIDS. In felids such as the domestic cat (Felis catus), APOBEC3 (A3) genes encode for single-domain A3Z2s, A3Z3 and double-domain A3Z2Z3 anti-viral cytidine deaminases. The feline A3Z2Z3 is expressed following read-through transcription and alternative splicing, introducing a previously untranslated exon in frame, encoding a domain insertion called linker. Only A3Z3 and A3Z2Z3 inhibit Vif-deficient FIV. Feline A3s also are restriction factors for HIV and Simian immunodeficiency viruses (SIV). Surprisingly, HIV-2/SIV Vifs can counteract feline A3Z2Z3. Results To identify residues in feline A3s that Vifs need for interaction and degradation, chimeric human–feline A3s were tested. Here we describe the molecular direct interaction of feline A3s with Vif proteins from cat FIV and present the first structural A3 model locating these interaction regions. In the Z3 domain we have identified residues involved in binding of FIV Vif, and their mutation blocked Vif-induced A3Z3 degradation. We further identified additional essential residues for FIV Vif interaction in the A3Z2 domain, allowing the generation of FIV Vif resistant A3Z2Z3. Mutated feline A3s also showed resistance to the Vif of a lion-specific FIV, indicating an evolutionary conserved Vif–A3 binding. Comparative modelling of feline A3Z2Z3 suggests that the residues interacting with FIV Vif have, unlike Vif-interacting residues in human A3s, a unique location at the domain interface of Z2 and Z3 and that the linker forms a homeobox-like domain protruding of the Z2Z3 core. HIV-2/SIV Vifs efficiently degrade feline A3Z2Z3 by possible targeting the linker stretch connecting both Z-domains. Conclusions Here we identified in feline A3s residues important for binding of FIV Vif and a unique protein domain insertion (linker). To understand Vif evolution, a structural model of the feline A3 was developed. Our results show that HIV Vif binds human A3s differently than FIV Vif feline A3s. The linker insertion is suggested to form a homeo-box domain, which is unique to A3s of cats and related species, and not found in human and mouse A3s. Together, these findings indicate a specific and different A3 evolution in cats and human. Electronic supplementary material The online version of this article (doi:10.1186/s12977-016-0274-9) contains supplementary material, which is available to authorized users.
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Affiliation(s)
- Zeli Zhang
- Clinic for Gastroenterology, Hepatology, and Infectiology, Medical Faculty, Heinrich-Heine-University Düsseldorf, Building 23.12.U1.82, Moorenstr. 5, 40225, Düsseldorf, Germany
| | - Qinyong Gu
- Clinic for Gastroenterology, Hepatology, and Infectiology, Medical Faculty, Heinrich-Heine-University Düsseldorf, Building 23.12.U1.82, Moorenstr. 5, 40225, Düsseldorf, Germany
| | - Ananda Ayyappan Jaguva Vasudevan
- Clinic for Gastroenterology, Hepatology, and Infectiology, Medical Faculty, Heinrich-Heine-University Düsseldorf, Building 23.12.U1.82, Moorenstr. 5, 40225, Düsseldorf, Germany
| | - Anika Hain
- Clinic for Gastroenterology, Hepatology, and Infectiology, Medical Faculty, Heinrich-Heine-University Düsseldorf, Building 23.12.U1.82, Moorenstr. 5, 40225, Düsseldorf, Germany
| | - Björn-Philipp Kloke
- Department of Medical Biotechnology, Paul-Ehrlich-Institute, Paul-Ehrlich-Str. 51-59, 63225, Langen, Germany.,BioNTech RNA Pharmaceuticals GmbH, An der Goldgrube 12, 55131, Mainz, Germany
| | - Sascha Hasheminasab
- Clinic for Gastroenterology, Hepatology, and Infectiology, Medical Faculty, Heinrich-Heine-University Düsseldorf, Building 23.12.U1.82, Moorenstr. 5, 40225, Düsseldorf, Germany
| | - Daniel Mulnaes
- Institute of Pharmaceutical and Medicinal Chemistry, Heinrich Heine University Düsseldorf, Universitätsstr. 1, 40225, Düsseldorf, Germany
| | - Kei Sato
- Laboratory of Viral Pathogenesis, Institute for Virus Research, Kyoto University, Kyoto, 6068507, Japan.,CREST, Japan Science and Technology Agency, Saitama, 3220012, Japan
| | - Klaus Cichutek
- Department of Medical Biotechnology, Paul-Ehrlich-Institute, Paul-Ehrlich-Str. 51-59, 63225, Langen, Germany
| | - Dieter Häussinger
- Clinic for Gastroenterology, Hepatology, and Infectiology, Medical Faculty, Heinrich-Heine-University Düsseldorf, Building 23.12.U1.82, Moorenstr. 5, 40225, Düsseldorf, Germany
| | - Ignacio G Bravo
- MIVEGEC (UMR CNRS 5290, IRD 224, UM), National Center of Scientific Research (CNRS), 34394, Montpellier, France
| | - Sander H J Smits
- Institute of Biochemistry, Heinrich Heine University Düsseldorf, Universitätsstr. 1, 40225, Düsseldorf, Germany
| | - Holger Gohlke
- Institute of Pharmaceutical and Medicinal Chemistry, Heinrich Heine University Düsseldorf, Universitätsstr. 1, 40225, Düsseldorf, Germany
| | - Carsten Münk
- Clinic for Gastroenterology, Hepatology, and Infectiology, Medical Faculty, Heinrich-Heine-University Düsseldorf, Building 23.12.U1.82, Moorenstr. 5, 40225, Düsseldorf, Germany.
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Delviks-Frankenberry KA, Nikolaitchik OA, Burdick RC, Gorelick RJ, Keele BF, Hu WS, Pathak VK. Minimal Contribution of APOBEC3-Induced G-to-A Hypermutation to HIV-1 Recombination and Genetic Variation. PLoS Pathog 2016; 12:e1005646. [PMID: 27186986 PMCID: PMC4871359 DOI: 10.1371/journal.ppat.1005646] [Citation(s) in RCA: 38] [Impact Index Per Article: 4.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/26/2016] [Accepted: 04/28/2016] [Indexed: 11/19/2022] Open
Abstract
Although the predominant effect of host restriction APOBEC3 proteins on HIV-1 infection is to block viral replication, they might inadvertently increase retroviral genetic variation by inducing G-to-A hypermutation. Numerous studies have disagreed on the contribution of hypermutation to viral genetic diversity and evolution. Confounding factors contributing to the debate include the extent of lethal (stop codon) and sublethal hypermutation induced by different APOBEC3 proteins, the inability to distinguish between G-to-A mutations induced by APOBEC3 proteins and error-prone viral replication, the potential impact of hypermutation on the frequency of retroviral recombination, and the extent to which viral recombination occurs in vivo, which can reassort mutations in hypermutated genomes. Here, we determined the effects of hypermutation on the HIV-1 recombination rate and its contribution to genetic variation through recombination to generate progeny genomes containing portions of hypermutated genomes without lethal mutations. We found that hypermutation did not significantly affect the rate of recombination, and recombination between hypermutated and wild-type genomes only increased the viral mutation rate by 3.9 × 10-5 mutations/bp/replication cycle in heterozygous virions, which is similar to the HIV-1 mutation rate. Since copackaging of hypermutated and wild-type genomes occurs very rarely in vivo, recombination between hypermutated and wild-type genomes does not significantly contribute to the genetic variation of replicating HIV-1. We also analyzed previously reported hypermutated sequences from infected patients and determined that the frequency of sublethal mutagenesis for A3G and A3F is negligible (4 × 10-21 and1 × 10-11, respectively) and its contribution to viral mutations is far below mutations generated during error-prone reverse transcription. Taken together, we conclude that the contribution of APOBEC3-induced hypermutation to HIV-1 genetic variation is substantially lower than that from mutations during error-prone replication.
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Affiliation(s)
- Krista A. Delviks-Frankenberry
- Viral Mutation Section, HIV Dynamics and Replication Program, National Cancer Institute at Frederick, Frederick, Maryland, United States of America
| | - Olga A. Nikolaitchik
- Viral Recombination Section, HIV Dynamics and Replication Program, National Cancer Institute at Frederick, Frederick, Maryland, United States of America
| | - Ryan C. Burdick
- Viral Mutation Section, HIV Dynamics and Replication Program, National Cancer Institute at Frederick, Frederick, Maryland, United States of America
| | - Robert J. Gorelick
- AIDS and Cancer Virus Program, Leidos Biomedical Research, Inc., Frederick National Lab, Frederick, Maryland, United States of America
| | - Brandon F. Keele
- AIDS and Cancer Virus Program, Leidos Biomedical Research, Inc., Frederick National Lab, Frederick, Maryland, United States of America
| | - Wei-Shau Hu
- Viral Recombination Section, HIV Dynamics and Replication Program, National Cancer Institute at Frederick, Frederick, Maryland, United States of America
| | - Vinay K. Pathak
- Viral Mutation Section, HIV Dynamics and Replication Program, National Cancer Institute at Frederick, Frederick, Maryland, United States of America
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APOBEC3G and APOBEC3F Act in Concert To Extinguish HIV-1 Replication. J Virol 2016; 90:4681-4695. [PMID: 26912618 DOI: 10.1128/jvi.03275-15] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/02/2016] [Accepted: 02/18/2016] [Indexed: 11/20/2022] Open
Abstract
UNLABELLED The multifunctional HIV-1 accessory protein Vif counters the antiviral activities of APOBEC3G (A3G) and APOBEC3F (A3F), and some Vifs counter stable alleles of APOBEC3H (A3H). Studies in humanized mice have shown that HIV-1 lacking Vif expression is not viable. Here, we look at the relative contributions of the three APOBEC3s to viral extinction. Inoculation of bone marrow/liver/thymus (BLT) mice with CCR5-tropic HIV-1JRCSF(JRCSF) expressing a vif gene inactive for A3G but not A3F degradation activity (JRCSFvifH42/43D) displayed either no or delayed replication. JRCSF expressing a vif gene mutated to inactivate A3F degradation but not A3G degradation (JRCSFvifW79S) always replicated to high viral loads with variable delays. JRCSF with vif mutated to lack both A3G and A3F degradation activities (JRCSFvifH42/43DW79S) failed to replicate, mimicking JRCSF without Vif expression (JRCSFΔvif). JRCSF and JRCSFvifH42/43D, but not JRCSFvifW79S or JRCSFvifH42/43DW79S, degraded APOBEC3D. With one exception, JRCSFs expressing mutant Vifs that replicated acquired enforced vif mutations. These mutations partially restored A3G or A3F degradation activity and fully replaced JRCSFvifH42/43D or JRCSFvifW79S by 10 weeks. Surprisingly, induced mutations temporally lagged behind high levels of virus in blood. In the exceptional case, JRCSFvifH42/43D replicated after a prolonged delay with no mutations in vif but instead a V27I mutation in the RNase H coding sequence. JRCSFvifH42/43D infections exhibited massive GG/AG mutations in pol viral DNA, but in viral RNA, there were no fixed mutations in the Gag or reverse transcriptase coding sequence. A3H did not contribute to viral extinction but, in combination with A3F, could delay JRCSF replication. A3H was also found to hypermutate viral DNA. IMPORTANCE Vif degradation of A3G and A3F enhances viral fitness, as virus with even a partially restored capacity for degradation outgrows JRCSFvifH42/43D and JRCSFvifW79S. Unexpectedly, fixation of mutations that replaced H42/43D or W79S in viral RNA lagged behind the appearance of high viral loads. In one exceptional JRCSFvifH42/43D infection, vif was unchanged but replication proceeded after a long delay. These results suggest that Vif binds and inhibits the non-cytosine deaminase activities of intact A3G and intact A3F, allowing JRCSFvifH42/43D and JRCSFvifW79S to replicate with reduced fitness. Subsequently, enhanced Vif function is acquired by enforced mutations. In infected cells, JRCSFΔvif and JRCSFvifH42/43DW79S are exposed to active A3F and A3G and fail to replicate. JRCSFvifH42/43D Vif degrades A3F and, in some cases, overcomes A3G mutagenic activity to replicate. Vif may have evolved to inhibit A3F and A3G by stoichiometric binding and subsequently acquired the ability to target these proteins to proteasomes.
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Chen Q, Xiao X, Wolfe A, Chen XS. The in vitro Biochemical Characterization of an HIV-1 Restriction Factor APOBEC3F: Importance of Loop 7 on Both CD1 and CD2 for DNA Binding and Deamination. J Mol Biol 2016; 428:2661-70. [PMID: 27063502 DOI: 10.1016/j.jmb.2016.03.031] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/11/2015] [Revised: 03/27/2016] [Accepted: 03/28/2016] [Indexed: 10/22/2022]
Abstract
APOBEC3F (A3F) is a member of the apolipoprotein B mRNA-editing enzyme catalytic polypeptide-like (APOBEC) family of proteins that can deaminate cytosine (C) to uracil (U) on nucleic acids. A3F is one of the four APOBEC members with two Zn-coordinated homologous cytosine deaminase (CD) domains, with the others being A3G, A3D, and A3B. Here we report the in vitro characterization of DNA binding and deaminase activities using purified wild-type and various mutant proteins of A3F from an Escherichia coli expression system. We show that even though CD1 is catalytically inactive and CD2 is the active deaminase domain, presence of CD1 on the N-terminus of CD2 enhances the deaminase activity by over an order of magnitude. This enhancement of CD2 catalytic activity is mainly through the increase of substrate single-stranded (ss) DNA binding by the N-terminal CD1 domain. We further show that the loop 7 of both CD1 and CD2 of A3F plays an important role for ssDNA binding for each individual domain, as well as for the deaminase activity of CD2 domain in the full-length A3F.
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Affiliation(s)
- Qihan Chen
- Molecular and Computational Biology Program, Departments of Biological Sciences and Chemistry, University of Southern California, Los Angeles, CA 90089, USA
| | - Xiao Xiao
- Genetic, Molecular and Cellular Biology, Department of Biological Sciences, University of Southern California, Los Angeles, CA 90089, USA
| | - Aaron Wolfe
- Genetic, Molecular and Cellular Biology, Department of Biological Sciences, University of Southern California, Los Angeles, CA 90089, USA
| | - Xiaojiang S Chen
- Molecular and Computational Biology Program, Departments of Biological Sciences and Chemistry, University of Southern California, Los Angeles, CA 90089, USA; Center of Excellence in NanoBiophysics, University of Southern California, Los Angeles, CA 90089, USA; Norris Comprehensive Cancer Center; University of Southern California, Los Angeles, CA 90089, USA.
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Ross KF, Herzberg MC. Autonomous immunity in mucosal epithelial cells: fortifying the barrier against infection. Microbes Infect 2016; 18:387-398. [PMID: 27005450 DOI: 10.1016/j.micinf.2016.03.008] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/17/2016] [Revised: 03/09/2016] [Accepted: 03/10/2016] [Indexed: 12/19/2022]
Abstract
Mucosal epithelial cells express an autonomous innate immune response that controls the overgrowth of invaded bacteria, mitigates the harmful effects of the bacteria carried within, and does not rely on other external arms of the immune response. Epithelial cell autonomous innate immunity "respects" the social biology of invading bacteria to achieve symbiosis, and is the primary protective mechanism against pathogens.
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Affiliation(s)
- Karen F Ross
- Department of Diagnostic and Biological Sciences, School of Dentistry, University of Minnesota, USA
| | - Mark C Herzberg
- Department of Diagnostic and Biological Sciences, School of Dentistry, University of Minnesota, USA.
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46
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An P, Penugonda S, Thorball CW, Bartha I, Goedert JJ, Donfield S, Buchbinder S, Binns-Roemer E, Kirk GD, Zhang W, Fellay J, Yu XF, Winkler CA. Role of APOBEC3F Gene Variation in HIV-1 Disease Progression and Pneumocystis Pneumonia. PLoS Genet 2016; 12:e1005921. [PMID: 26942578 PMCID: PMC4778847 DOI: 10.1371/journal.pgen.1005921] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/11/2015] [Accepted: 02/16/2016] [Indexed: 12/26/2022] Open
Abstract
Human APOBEC3 cytidine deaminases are intrinsic resistance factors to HIV-1. However, HIV-1 encodes a viral infectivity factor (Vif) that degrades APOBEC3 proteins. In vitro APOBEC3F (A3F) anti-HIV-1 activity is weaker than A3G but is partially resistant to Vif degradation unlike A3G. It is unknown whether A3F protein affects HIV-1 disease in vivo. To assess the effect of A3F gene on host susceptibility to HIV- acquisition and disease progression, we performed a genetic association study in six well-characterized HIV-1 natural cohorts. A common six-Single Nucleotide Polymorphism (SNP) haplotype of A3F tagged by a codon-changing variant (p. I231V, with allele (V) frequency of 48% in European Americans) was associated with significantly lower set-point viral load and slower rate of progression to AIDS (Relative Hazards (RH) = 0.71, 95% CI: 0.56, 0.91) and delayed development of pneumocystis pneumonia (PCP) (RH = 0.53, 95% CI: 0.37-0.76). A validation study in the International Collaboration for the Genomics of HIV (ICGH) showed a consistent association with lower set-point viral load. An in vitro assay revealed that the A3F I231V variant may influence Vif mediated A3F degradation. Our results provide genetic epidemiological evidence that A3F modulates HIV-1/AIDS disease progression.
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Affiliation(s)
- Ping An
- Basic Research Laboratory, Center for Cancer Research, National Cancer Institute, Leidos Biomedical Research, Inc., Frederick National Laboratory for Cancer Research, Frederick, Maryland, United States of America
- * E-mail: (PA); (CAW)
| | - Sudhir Penugonda
- Department of Medicine, Northwestern University Feinberg School of Medicine, Chicago, Illinois, United States of America
| | - Christian W. Thorball
- School of Life Sciences, École Polytechnique Fédérale de Lausanne, Lausanne, Switzerland
- Swiss Institute of Bioinformatics, Lausanne, Switzerland
| | - Istvan Bartha
- School of Life Sciences, École Polytechnique Fédérale de Lausanne, Lausanne, Switzerland
- Swiss Institute of Bioinformatics, Lausanne, Switzerland
| | - James J. Goedert
- Infections and Immunoepidemiology Branch, Division of Cancer Epidemiology and Genetics, National Cancer Institute, Bethesda, Maryland, United States of America
| | - Sharyne Donfield
- Rho, Inc., Chapel Hill, North Carolina, United States of America
| | - Susan Buchbinder
- San Francisco Department of Public Health, San Francisco, California, United States of America
| | - Elizabeth Binns-Roemer
- Basic Research Laboratory, Center for Cancer Research, National Cancer Institute, Leidos Biomedical Research, Inc., Frederick National Laboratory for Cancer Research, Frederick, Maryland, United States of America
| | - Gregory D. Kirk
- Department of Epidemiology, Johns Hopkins School of Public Health, Baltimore, Maryland, United States of America
| | - Wenyan Zhang
- Institute of Virology and AIDS Research, First Hospital of Jilin University, Changchun, China
| | - Jacques Fellay
- School of Life Sciences, École Polytechnique Fédérale de Lausanne, Lausanne, Switzerland
- Swiss Institute of Bioinformatics, Lausanne, Switzerland
| | - Xiao-Fang Yu
- Institute of Virology and AIDS Research, First Hospital of Jilin University, Changchun, China
- Department of Molecular Microbiology and Immunology, Johns Hopkins School of Public Health, Baltimore, Maryland, United States of America
| | - Cheryl A. Winkler
- Basic Research Laboratory, Center for Cancer Research, National Cancer Institute, Leidos Biomedical Research, Inc., Frederick National Laboratory for Cancer Research, Frederick, Maryland, United States of America
- * E-mail: (PA); (CAW)
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47
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Knisbacher BA, Levanon EY. DNA Editing of LTR Retrotransposons Reveals the Impact of APOBECs on Vertebrate Genomes. Mol Biol Evol 2015; 33:554-67. [PMID: 26541172 PMCID: PMC4866542 DOI: 10.1093/molbev/msv239] [Citation(s) in RCA: 20] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/31/2022] Open
Abstract
Long terminal repeat retrotransposons (LTR) are widespread in vertebrates and their dynamism facilitates genome evolution. However, these endogenous retroviruses (ERVs) must be restricted to maintain genomic stability. The APOBECs, a protein family that can edit C-to-U in DNA, do so by interfering with reverse transcription and hypermutating retrotransposon DNA. In some cases, a retrotransposon may integrate into the genome despite being hypermutated. Such an event introduces a unique sequence into the genome, increasing retrotransposon diversity and the probability of developing new function at the locus of insertion. The prevalence of this phenomenon and its effects on vertebrate genomes are still unclear. In this study, we screened ERV sequences in the genomes of 123 diverse species and identified hundreds of thousands of edited sites in multiple vertebrate lineages, including placental mammals, marsupials, and birds. Numerous edited ERVs carry high mutation loads, some with greater than 350 edited sites, profoundly damaging their open-reading frames. For many of the species studied, this is the first evidence that APOBECs are active players in their innate immune system. Unexpectedly, some birds and especially zebra finch and medium ground-finch (one of Darwin’s finches) are exceptionally enriched in DNA editing. We demonstrate that edited retrotransposons may be preferentially retained in active genomic regions, as reflected from their enrichment in genes, exons, promoters, and transcription start sites, thereby raising the probability of their exaptation for novel function. In conclusion, DNA editing of retrotransposons by APOBECs has a substantial role in vertebrate innate immunity and may boost genome evolution.
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Affiliation(s)
- Binyamin A Knisbacher
- The Mina & Everard Goodman Faculty of Life Sciences, Bar-Ilan University, Ramat Gan, 52900, Israel
| | - Erez Y Levanon
- The Mina & Everard Goodman Faculty of Life Sciences, Bar-Ilan University, Ramat Gan, 52900, Israel
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Chang MO, Suzuki T, Kitajima M, Takaku H. Baculovirus Infection of Human Monocyte-Derived Dendritic Cells Restricts HIV-1 Replication. AIDS Res Hum Retroviruses 2015; 31:1023-31. [PMID: 26178669 DOI: 10.1089/aid.2015.0060] [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: 11/12/2022] Open
Abstract
Acquired immune deficiency syndrome (AIDS) is mainly caused by infection with human immunodeficiency virus-1 (HIV-1) and still poses a global threat for which we lack a protective or therapeutic vaccine. Dendritic cells (DCs) play a major role in the onset of HIV infection, providing one of the primary sites of HIV replication, and also act as viral reservoirs in vivo. Previous studies have shown that baculovirus (BV) induces strong host immune responses against infections and malignancies. In this study, we infected human monocyte-derived DCs with recombinant BV (AcCAG-gag) and showed that AcCAG-gag-infected human DCs underwent maturation and produced interferon alpha and other proinflammatory cytokines accompanied by increases in the mRNA and protein expression levels of APOBEC3 (A3A, A3F, and A3G), proteins associated with the inhibition of HIV-1 replication. Surprisingly, HIV-1 inhibition is also observed in human DCs infected with a wild-type BV, as determined by the production of inflammatory cytokines, the expression of A3, and a reduction in the p24 level. Our findings outline the mechanism underlying the inhibition of HIV-1 in BV-infected human DCs and pave the way for the use of BV as an effective tool for immunotherapy against HIV-1.
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Affiliation(s)
- Myint Oo Chang
- 1 High Technology Research Centre, Chiba Institute of Technology , Chiba, Japan
| | - Tomoyuki Suzuki
- 2 Department of Life and Environmental Sciences, Chiba Institute of Technology , Chiba, Japan
| | - Masayuki Kitajima
- 2 Department of Life and Environmental Sciences, Chiba Institute of Technology , Chiba, Japan
- 3 Department of Immunology and Pathology, Research Institute National Center for Global Health and Medicine , Chiba, Japan
| | - Hiroshi Takaku
- 1 High Technology Research Centre, Chiba Institute of Technology , Chiba, Japan
- 2 Department of Life and Environmental Sciences, Chiba Institute of Technology , Chiba, Japan
- 4 Research Institute, Chiba Institute of Technology , Chiba, Japan
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49
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Valera MS, de Armas-Rillo L, Barroso-González J, Ziglio S, Batisse J, Dubois N, Marrero-Hernández S, Borel S, García-Expósito L, Biard-Piechaczyk M, Paillart JC, Valenzuela-Fernández A. The HDAC6/APOBEC3G complex regulates HIV-1 infectiveness by inducing Vif autophagic degradation. Retrovirology 2015; 12:53. [PMID: 26105074 PMCID: PMC4479245 DOI: 10.1186/s12977-015-0181-5] [Citation(s) in RCA: 47] [Impact Index Per Article: 4.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/06/2014] [Accepted: 06/10/2015] [Indexed: 02/06/2023] Open
Abstract
BACKGROUND Human immunodeficiency virus type 1 (HIV-1) has evolved a complex strategy to overcome the immune barriers it encounters throughout an organism thanks to its viral infectivity factor (Vif), a key protein for HIV-1 infectivity and in vivo pathogenesis. Vif interacts with and promotes "apolipoprotein B mRNA-editing enzyme-catalytic, polypeptide-like 3G" (A3G) ubiquitination and subsequent degradation by the proteasome, thus eluding A3G restriction activity against HIV-1. RESULTS We found that cellular histone deacetylase 6 (HDAC6) directly interacts with A3G through its C-terminal BUZ domain (residues 841-1,215) to undergo a cellular co-distribution along microtubules and cytoplasm. The HDAC6/A3G complex occurs in the absence or presence of Vif, competes for Vif-mediated A3G degradation, and accounts for A3G steady-state expression level. In fact, HDAC6 directly interacts with and promotes Vif autophagic clearance, thanks to its C-terminal BUZ domain, a process requiring the deacetylase activity of HDAC6. HDAC6 degrades Vif without affecting the core binding factor β (CBF-β), a Vif-associated partner reported to be key for Vif- mediated A3G degradation. Thus HDAC6 antagonizes the proviral activity of Vif/CBF-β-associated complex by targeting Vif and stabilizing A3G. Finally, in cells producing virions, we observed a clear-cut correlation between the ability of HDAC6 to degrade Vif and to restore A3G expression, suggesting that HDAC6 controls the amount of Vif incorporated into nascent virions and the ability of HIV-1 particles of being infectious. This effect seems independent on the presence of A3G inside virions and on viral tropism. CONCLUSIONS Our study identifies for the first time a new cellular complex, HDAC6/A3G, involved in the autophagic degradation of Vif, and suggests that HDAC6 represents a new antiviral factor capable of controlling HIV-1 infectiveness by counteracting Vif and its functions.
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Affiliation(s)
- María-Soledad Valera
- Laboratorio de Inmunología Celular y Viral, Unidad de Farmacología, Departamento de Medicina Física y Farmacología, Facultad de Medicina, Universidad de La Laguna (ULL), Campus de Ofra s/n, 38071, La Laguna, Tenerife, Spain.
| | - Laura de Armas-Rillo
- Laboratorio de Inmunología Celular y Viral, Unidad de Farmacología, Departamento de Medicina Física y Farmacología, Facultad de Medicina, Universidad de La Laguna (ULL), Campus de Ofra s/n, 38071, La Laguna, Tenerife, Spain.
| | - Jonathan Barroso-González
- Laboratorio de Inmunología Celular y Viral, Unidad de Farmacología, Departamento de Medicina Física y Farmacología, Facultad de Medicina, Universidad de La Laguna (ULL), Campus de Ofra s/n, 38071, La Laguna, Tenerife, Spain.
| | - Serena Ziglio
- Laboratorio de Inmunología Celular y Viral, Unidad de Farmacología, Departamento de Medicina Física y Farmacología, Facultad de Medicina, Universidad de La Laguna (ULL), Campus de Ofra s/n, 38071, La Laguna, Tenerife, Spain.
| | - Julien Batisse
- Architecture et Réactivité de l'ARN, CNRS, Institut de Biologie Moléculaire et Cellulaire, Université de Strasbourg, 15 rue René Descartes, 67084, Strasbourg, France.
| | - Noé Dubois
- Architecture et Réactivité de l'ARN, CNRS, Institut de Biologie Moléculaire et Cellulaire, Université de Strasbourg, 15 rue René Descartes, 67084, Strasbourg, France.
| | - Sara Marrero-Hernández
- Laboratorio de Inmunología Celular y Viral, Unidad de Farmacología, Departamento de Medicina Física y Farmacología, Facultad de Medicina, Universidad de La Laguna (ULL), Campus de Ofra s/n, 38071, La Laguna, Tenerife, Spain.
| | - Sophie Borel
- Centre d'études d'agents Pathogènes et Biotechnologies pour la Santé (CPBS) UMR5236 CNRS UMSF, 1919 route de Mende, 34293, Montpellier Cedex 5, France.
| | - Laura García-Expósito
- Laboratorio de Inmunología Celular y Viral, Unidad de Farmacología, Departamento de Medicina Física y Farmacología, Facultad de Medicina, Universidad de La Laguna (ULL), Campus de Ofra s/n, 38071, La Laguna, Tenerife, Spain.
| | - Martine Biard-Piechaczyk
- Centre d'études d'agents Pathogènes et Biotechnologies pour la Santé (CPBS) UMR5236 CNRS UMSF, 1919 route de Mende, 34293, Montpellier Cedex 5, France.
| | - Jean-Christophe Paillart
- Architecture et Réactivité de l'ARN, CNRS, Institut de Biologie Moléculaire et Cellulaire, Université de Strasbourg, 15 rue René Descartes, 67084, Strasbourg, France.
| | - Agustín Valenzuela-Fernández
- Laboratorio de Inmunología Celular y Viral, Unidad de Farmacología, Departamento de Medicina Física y Farmacología, Facultad de Medicina, Universidad de La Laguna (ULL), Campus de Ofra s/n, 38071, La Laguna, Tenerife, Spain.
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50
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High Expression of Antiviral Proteins in Mucosa from Individuals Exhibiting Resistance to Human Immunodeficiency Virus. PLoS One 2015; 10:e0131139. [PMID: 26091527 PMCID: PMC4474690 DOI: 10.1371/journal.pone.0131139] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/20/2015] [Accepted: 05/27/2015] [Indexed: 11/19/2022] Open
Abstract
BACKGROUND Several soluble factors have been reported to have the capacity of inhibiting HIV replication at different steps of the virus life cycle, without eliminating infected cells and through enhancement of specific cellular mechanisms. Yet, it is unclear if these antiviral factors play a role in the protection from HIV infection or in the control of viral replication. Here we evaluated two cohorts: i) one of 58 HIV-exposed seronegative individuals (HESNs) who were compared with 59 healthy controls (HCs), and ii) another of 13 HIV-controllers who were compared with 20 HIV-progressors. Peripheral blood, oral and genital mucosa and gut-associated lymphoid tissue (GALT) samples were obtained to analyze the mRNA expression of ELAFIN, APOBEC3G, SAMHD1, TRIM5α, RNase 7 and SerpinA1 using real-time PCR. RESULTS HESNs exhibited higher expression of all antiviral factors in peripheral blood mononuclear cells (PBMCs), oral or genital mucosa when compared with HCs. Furthermore, HIV-controllers exhibited higher levels of SerpinA1 in GALT. CONCLUSIONS These findings suggest that the activity of these factors is compartmentalized and that these proteins have a predominant role depending on the tissue to avoid the infection, reduce the viral load and modulate the susceptibility to HIV infection.
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