|
1
|
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.
Collapse
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
| |
Collapse
|
|
2
|
Steele EJ, Lindley RA. Deaminase-Driven Reverse Transcription Mutagenesis in Oncogenesis: Critical Analysis of Transcriptional Strand Asymmetries of Single Base Substitution Signatures. Int J Mol Sci 2025; 26:989. [PMID: 39940758 PMCID: PMC11817618 DOI: 10.3390/ijms26030989] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/19/2024] [Revised: 01/05/2025] [Accepted: 01/09/2025] [Indexed: 02/16/2025] Open
Abstract
This paper provides a critical analysis of the molecular mechanisms presently used to explain transcriptional strand asymmetries of single base substitution (SBS) signatures observed in cancer genomes curated at the Catalogue of Somatic Mutations in Cancer (COSMIC) database (Wellcome Trust Sanger Institute). The analysis is based on a deaminase-driven reverse transcriptase (DRT) mutagenesis model of cancer oncogenesis involving both the cytosine (AID/APOBEC) and adenosine (ADAR) mutagenic deaminases. In this analysis we apply what is known, or can reasonably be inferred, of the immunoglobulin somatic hypermutation (Ig SHM) mechanism to the analysis of the transcriptional stand asymmetries of the COSMIC SBS signatures that are observed in cancer genomes. The underlying assumption is that somatic mutations arising in cancer genomes are driven by dysregulated off-target Ig SHM-like mutagenic processes at non-Ig loci. It is reasoned that most SBS signatures whether of "unknown etiology" or assigned-molecular causation, can be readily understood in terms of the DRT-paradigm. These include the major age-related "clock-like" SBS5 signature observed in all cancer genomes sequenced and many other common subset signatures including SBS1, SBS3, SBS2/13, SBS6, SBS12, SBS16, SBS17a/17b, SBS19, SBS21, as well as signatures clearly arising from exogenous causation. We conclude that the DRT-model provides a plausible molecular framework that augments our current understanding of immunogenetic mechanisms driving oncogenesis. It accommodates both what is known about AID/APOBEC and ADAR somatic mutation strand asymmetries and provides a fully integrated understanding into the molecular origins of common COSMIC SBS signatures. The DRT-paradigm thus provides scientists and clinicians with additional molecular insights into the causal links between deaminase-associated genomic signatures and oncogenic processes.
Collapse
Affiliation(s)
- Edward J. Steele
- Melville Analytics Pty Ltd. and Immunomics, Kangaroo Point, Brisbane 4169, Australia
| | - Robyn A. Lindley
- Department Clinical Pathology, Victorian Comprehensive Cancer Centre (VCCC), University of Melbourne, Melbourne 3052, Australia;
| |
Collapse
|
|
3
|
Gai Y, Duan S, Wang S, Liu K, Yu X, Yang C, Li G, Zhou Y, Yu B, Wu J, Wang C, Yu X. Design of Vif-Derived Peptide Inhibitors with Anti-HIV-1 Activity by Interrupting Vif-CBFβ Interaction. Viruses 2024; 16:490. [PMID: 38675833 PMCID: PMC11053914 DOI: 10.3390/v16040490] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/23/2023] [Revised: 03/16/2024] [Accepted: 03/20/2024] [Indexed: 04/28/2024] Open
Abstract
One of the major functions of the accessory protein Vif of human immunodeficiency virus type 1 (HIV-1) is to induce the degradation of APOBEC3 (A3) family proteins by recruiting a Cullin5-ElonginB/C-CBFβ E3 ubiquitin ligase complex to facilitate viral replication. Therefore, the interactions between Vif and the E3 complex proteins are promising targets for the development of novel anti-HIV-1 drugs. Here, peptides are designed for the Vif-CBFβ interaction based on the sequences of Vif mutants with higher affinity for CBFβ screened by a yeast surface display platform. We identified two peptides, VMP-63 and VMP-108, that could reduce the infectivity of HIV-1 produced from A3G-positive cells with IC50 values of 49.4 μM and 55.1 μM, respectively. They protected intracellular A3G from Vif-mediated degradation in HEK293T cells, consequently increasing A3G encapsulation into the progeny virions. The peptides could rapidly enter cells after addition to HEK293T cells and competitively inhibit the binding of Vif to CBFβ. Homology modeling analysis demonstrated the binding advantages of VMP-63 and VMP-108 with CBFβ over their corresponding wild-type peptides. However, only VMP-108 effectively restricted long-term HIV-1 replication and protected A3 functions in non-permissive T lymphocytes. Our findings suggest that competitive Vif-derived peptides targeting the Vif-CBFβ interaction are promising for the development of novel therapeutic strategies for acquired immune deficiency syndrome.
Collapse
Affiliation(s)
- Yanxin Gai
- National Engineering Laboratory for AIDS Vaccine, School of Life Sciences, Jilin University, Changchun 130012, China; (Y.G.); (S.D.); (S.W.); (X.Y.); (C.Y.); (G.L.); (Y.Z.); (B.Y.); (J.W.)
| | - Sizhu Duan
- National Engineering Laboratory for AIDS Vaccine, School of Life Sciences, Jilin University, Changchun 130012, China; (Y.G.); (S.D.); (S.W.); (X.Y.); (C.Y.); (G.L.); (Y.Z.); (B.Y.); (J.W.)
| | - Shiqi Wang
- National Engineering Laboratory for AIDS Vaccine, School of Life Sciences, Jilin University, Changchun 130012, China; (Y.G.); (S.D.); (S.W.); (X.Y.); (C.Y.); (G.L.); (Y.Z.); (B.Y.); (J.W.)
| | - Kaifeng Liu
- Key Laboratory for Molecular Enzymology and Engineering, The Ministry of Education, School of Life Sciences, Jilin University, Changchun 130012, China;
| | - Xin Yu
- National Engineering Laboratory for AIDS Vaccine, School of Life Sciences, Jilin University, Changchun 130012, China; (Y.G.); (S.D.); (S.W.); (X.Y.); (C.Y.); (G.L.); (Y.Z.); (B.Y.); (J.W.)
| | - Chumeng Yang
- National Engineering Laboratory for AIDS Vaccine, School of Life Sciences, Jilin University, Changchun 130012, China; (Y.G.); (S.D.); (S.W.); (X.Y.); (C.Y.); (G.L.); (Y.Z.); (B.Y.); (J.W.)
| | - Guoqing Li
- National Engineering Laboratory for AIDS Vaccine, School of Life Sciences, Jilin University, Changchun 130012, China; (Y.G.); (S.D.); (S.W.); (X.Y.); (C.Y.); (G.L.); (Y.Z.); (B.Y.); (J.W.)
| | - Yan Zhou
- National Engineering Laboratory for AIDS Vaccine, School of Life Sciences, Jilin University, Changchun 130012, China; (Y.G.); (S.D.); (S.W.); (X.Y.); (C.Y.); (G.L.); (Y.Z.); (B.Y.); (J.W.)
| | - Bin Yu
- National Engineering Laboratory for AIDS Vaccine, School of Life Sciences, Jilin University, Changchun 130012, China; (Y.G.); (S.D.); (S.W.); (X.Y.); (C.Y.); (G.L.); (Y.Z.); (B.Y.); (J.W.)
| | - Jiaxin Wu
- National Engineering Laboratory for AIDS Vaccine, School of Life Sciences, Jilin University, Changchun 130012, China; (Y.G.); (S.D.); (S.W.); (X.Y.); (C.Y.); (G.L.); (Y.Z.); (B.Y.); (J.W.)
| | - Chu Wang
- National Engineering Laboratory for AIDS Vaccine, School of Life Sciences, Jilin University, Changchun 130012, China; (Y.G.); (S.D.); (S.W.); (X.Y.); (C.Y.); (G.L.); (Y.Z.); (B.Y.); (J.W.)
| | - Xianghui Yu
- National Engineering Laboratory for AIDS Vaccine, School of Life Sciences, Jilin University, Changchun 130012, China; (Y.G.); (S.D.); (S.W.); (X.Y.); (C.Y.); (G.L.); (Y.Z.); (B.Y.); (J.W.)
- Key Laboratory for Molecular Enzymology and Engineering, The Ministry of Education, School of Life Sciences, Jilin University, Changchun 130012, China;
| |
Collapse
|
|
4
|
Ito F, Alvarez-Cabrera AL, Kim K, Zhou ZH, Chen XS. Structural basis of HIV-1 Vif-mediated E3 ligase targeting of host APOBEC3H. Nat Commun 2023; 14:5241. [PMID: 37640699 PMCID: PMC10462622 DOI: 10.1038/s41467-023-40955-x] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/14/2023] [Accepted: 08/17/2023] [Indexed: 08/31/2023] Open
Abstract
Human APOBEC3 (A3) cytidine deaminases are antiviral factors that are particularly potent against retroviruses. As a countermeasure, HIV-1 uses a viral infectivity factor (Vif) to target specific human A3s for proteasomal degradation. Vif recruits cellular transcription cofactor CBF-β and Cullin-5 (CUL5) RING E3 ubiquitin ligase to bind different A3s distinctively, but how this is accomplished remains unclear in the absence of the atomic structure of the complex. Here, we present the cryo-EM structures of HIV-1 Vif in complex with human A3H, CBF-β and components of CUL5 ubiquitin ligase (CUL5, ELOB, and ELOC). Vif nucleates the entire complex by directly binding four human proteins, A3H, CBF-β, CUL5, and ELOC. The structures reveal a large interface area between A3H and Vif, primarily mediated by an α-helical side of A3H and a five-stranded β-sheet of Vif. This A3H-Vif interface unveils the basis for sensitivity-modulating polymorphism of both proteins, including a previously reported gain-of-function mutation in Vif isolated from HIV/AIDS patients. Our structural and functional results provide insights into the remarkable interplay between HIV and humans and would inform development efforts for anti-HIV therapeutics.
Collapse
Affiliation(s)
- Fumiaki Ito
- Molecular and Computational Biology, Department of Biological Sciences, University of Southern California, Los Angeles, CA, 90089, USA
- Department of Microbiology, Immunology and Molecular Genetics, University of California, Los Angeles, CA90095, USA
- California NanoSystems Institute, University of California, Los Angeles, CA90095, USA
| | - Ana L Alvarez-Cabrera
- Department of Microbiology, Immunology and Molecular Genetics, University of California, Los Angeles, CA90095, USA
- California NanoSystems Institute, University of California, Los Angeles, CA90095, USA
| | - Kyumin Kim
- Molecular and Computational Biology, Department of Biological Sciences, University of Southern California, Los Angeles, CA, 90089, USA
| | - Z Hong Zhou
- Department of Microbiology, Immunology and Molecular Genetics, University of California, Los Angeles, CA90095, USA
- California NanoSystems Institute, University of California, Los Angeles, CA90095, USA
| | - Xiaojiang S Chen
- Molecular and Computational Biology, Department of Biological Sciences, University of Southern California, Los Angeles, CA, 90089, USA.
- Genetic, Molecular and Cellular Biology Program, Keck School of Medicine, University of Southern California, Los Angeles, CA90089, USA.
- Norris Comprehensive Cancer Center, University of Southern California, Los Angeles, CA90089, USA.
- Center of Excellence in NanoBiophysics, University of Southern California, Los Angeles, CA90089, USA.
| |
Collapse
|
|
5
|
Butler K, Banday AR. APOBEC3-mediated mutagenesis in cancer: causes, clinical significance and therapeutic potential. J Hematol Oncol 2023; 16:31. [PMID: 36978147 PMCID: PMC10044795 DOI: 10.1186/s13045-023-01425-5] [Citation(s) in RCA: 17] [Impact Index Per Article: 8.5] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/11/2023] [Accepted: 03/14/2023] [Indexed: 03/30/2023] Open
Abstract
Apolipoprotein B mRNA-editing enzyme, catalytic polypeptides (APOBECs) are cytosine deaminases involved in innate and adaptive immunity. However, some APOBEC family members can also deaminate host genomes to generate oncogenic mutations. The resulting mutations, primarily signatures 2 and 13, occur in many tumor types and are among the most common mutational signatures in cancer. This review summarizes the current evidence implicating APOBEC3s as major mutators and outlines the exogenous and endogenous triggers of APOBEC3 expression and mutational activity. The review also discusses how APOBEC3-mediated mutagenesis impacts tumor evolution through both mutagenic and non-mutagenic pathways, including by inducing driver mutations and modulating the tumor immune microenvironment. Moving from molecular biology to clinical outcomes, the review concludes by summarizing the divergent prognostic significance of APOBEC3s across cancer types and their therapeutic potential in the current and future clinical landscapes.
Collapse
Affiliation(s)
- Kelly Butler
- Genitourinary Malignancies Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD, 20892, USA
| | - A Rouf Banday
- Genitourinary Malignancies Branch, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD, 20892, USA.
| |
Collapse
|
|
6
|
Stability of APOBEC3F in the Presence of the APOBEC3 Antagonist HIV-1 Vif Increases at the Expense of Co-Expressed APOBEC3H Haplotype I. Viruses 2023; 15:v15020463. [PMID: 36851677 PMCID: PMC9960753 DOI: 10.3390/v15020463] [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: 12/29/2022] [Revised: 02/03/2023] [Accepted: 02/04/2023] [Indexed: 02/10/2023] Open
Abstract
The seven human APOBEC3 enzymes (APOBEC3A through H, excluding E) are host restriction factors. Most of the APOBEC3 enzymes can restrict HIV-1 replication with different efficiencies. The HIV-1 Vif protein combats APOBEC3-mediated restriction by inducing ubiquitination and degradation in the proteasome. APOBEC3F and APOBEC3G can hetero-oligomerize, which increases their restriction capacity and resistance to Vif. Here we determined if APOBEC3C, APOBEC3F, or APOBEC3G could hetero-oligomerize with APOBEC3H haplotype I. APOBEC3H haplotype I has a short half-life in cells due to ubiquitination and degradation by host proteins, but is also resistant to Vif. We hypothesized that hetero-oligomerization with APOBEC3H haplotype I may result in less Vif-mediated degradation of the interacting APOBEC3 and stabilize APOBEC3H haplotype I, resulting in more efficient HIV-1 restriction. Although we found that all three APOBEC3s could interact with APOBEC3H haplotype I, only APOBEC3F affected APOBEC3H haplotype I by surprisingly accelerating its proteasomal degradation. However, this increased APOBEC3F levels in cells and virions in the absence or presence of Vif and enabled APOBEC3F-mediated restriction of HIV-1 in the presence of Vif. Altogether, the data suggest that APOBEC3 enzymes can co-regulate each other at the protein level and that they cooperate to ensure HIV-1 inactivation rather than evolution.
Collapse
|
|
7
|
Nakata Y, Ode H, Kubota M, Kasahara T, Matsuoka K, Sugimoto A, Imahashi M, Yokomaku Y, Iwatani Y. Cellular APOBEC3A deaminase drives mutations in the SARS-CoV-2 genome. Nucleic Acids Res 2023; 51:783-795. [PMID: 36610792 PMCID: PMC9881129 DOI: 10.1093/nar/gkac1238] [Citation(s) in RCA: 36] [Impact Index Per Article: 18.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/18/2022] [Revised: 12/07/2022] [Accepted: 12/13/2022] [Indexed: 01/09/2023] Open
Abstract
The number of genetic variations in the SARS-CoV-2 genome has been increasing primarily due to continuous viral mutations. Here, we report that the human APOBEC3A (A3A) cytidine deaminase plays a critical role in the induction of C-to-U substitutions in the SARS-CoV-2 genome. Bioinformatic analysis of the chronological genetic changes in a sequence database indicated that the largest UC-to-UU mutation signature, consistent with APOBEC-recognized nucleotide motifs, was predominant in single-stranded RNA regions of the viral genome. In SARS-CoV-2-infected cells, exogenous expression of A3A but not expression of other APOBEC proteins induced UC-to-UU mutations in viral RNA (vRNA). Additionally, the mutated C bases were often located at the tips in bulge or loop regions in the vRNA secondary structure. Interestingly, A3A mRNA expression was drastically increased by interferons (IFNs) and tumour necrosis factor-α (TNF-α) in epithelial cells derived from the respiratory system, a site of efficient SARS-CoV-2 replication. Moreover, the UC-to-UU mutation rate was increased in SARS-CoV-2 produced from lung epithelial cells treated with IFN-ß and TNF-α, but not from CRISPR/Cas9-based A3A knockout cells. Collectively, these findings demonstrate that A3A is a primary host factor that drives mutations in the SARS-CoV-2 RNA genome via RNA editing.
Collapse
Affiliation(s)
- Yoshihiro Nakata
- Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi 460-0001, Japan,Department of AIDS Research, Division of Basic Medicine, Nagoya University Graduate School of Medicine, Nagoya, Aichi 466-8550, Japan
| | - Hirotaka Ode
- Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi 460-0001, Japan
| | - Mai Kubota
- Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi 460-0001, Japan
| | - Takaaki Kasahara
- Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi 460-0001, Japan,Department of Respiratory Medicine, Division of Internal Medicine, Nagoya University Graduate School of Medicine, Nagoya, Aichi 466-8550, Japan
| | - Kazuhiro Matsuoka
- Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi 460-0001, Japan
| | - Atsuko Sugimoto
- Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi 460-0001, Japan
| | - Mayumi Imahashi
- Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi 460-0001, Japan
| | - Yoshiyuki Yokomaku
- Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi 460-0001, Japan
| | - Yasumasa Iwatani
- To whom correspondence should be addressed. Tel: +81 52 951 1111; Fax: +81 52 963 3970;
| |
Collapse
|
|
8
|
Wong L, Sami A, Chelico L. Competition for DNA binding between the genome protector replication protein A and the genome modifying APOBEC3 single-stranded DNA deaminases. Nucleic Acids Res 2022; 50:12039-12057. [PMID: 36444883 PMCID: PMC9757055 DOI: 10.1093/nar/gkac1121] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/05/2022] [Revised: 10/21/2022] [Accepted: 11/08/2022] [Indexed: 11/30/2022] Open
Abstract
The human APOBEC family of eleven cytosine deaminases use RNA and single-stranded DNA (ssDNA) as substrates to deaminate cytosine to uracil. This deamination event has roles in lipid metabolism by altering mRNA coding, adaptive immunity by causing evolution of antibody genes, and innate immunity through inactivation of viral genomes. These benefits come at a cost where some family members, primarily from the APOBEC3 subfamily (APOBEC3A-H, excluding E), can cause off-target deaminations of cytosine to form uracil on transiently single-stranded genomic DNA, which induces mutations that are associated with cancer evolution. Since uracil is only promutagenic, the mutations observed in cancer genomes originate only when uracil is not removed by uracil DNA glycosylase (UNG) or when the UNG-induced abasic site is erroneously repaired. However, when ssDNA is present, replication protein A (RPA) binds and protects the DNA from nucleases or recruits DNA repair proteins, such as UNG. Thus, APOBEC enzymes must compete with RPA to access their substrate. Certain APOBEC enzymes can displace RPA, bind and scan ssDNA efficiently to search for cytosines, and can become highly overexpressed in tumor cells. Depending on the DNA replication conditions and DNA structure, RPA can either be in excess or deficient. Here we discuss the interplay between these factors and how despite RPA, multiple cancer genomes have a mutation bias at cytosines indicative of APOBEC activity.
Collapse
Affiliation(s)
- Lai Wong
- University of Saskatchewan, College of Medicine, Department of Biochemistry, Microbiology, and Immunology, Saskatoon, Saskatchewan, S7N 5E5, Canada
| | - Alina Sami
- University of Saskatchewan, College of Medicine, Department of Biochemistry, Microbiology, and Immunology, Saskatoon, Saskatchewan, S7N 5E5, Canada
| | - Linda Chelico
- To whom correspondence should be addressed. Tel: +1 306 966 4318; Fax: +1 306 966 4298;
| |
Collapse
|
|
9
|
The current toolbox for APOBEC drug discovery. Trends Pharmacol Sci 2022; 43:362-377. [PMID: 35272863 PMCID: PMC9018551 DOI: 10.1016/j.tips.2022.02.007] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/08/2021] [Revised: 02/03/2022] [Accepted: 02/09/2022] [Indexed: 12/14/2022]
Abstract
Mutational processes driving genome evolution and heterogeneity contribute to immune evasion and therapy resistance in viral infections and cancer. APOBEC3 (A3) enzymes promote such mutations by catalyzing the deamination of cytosines to uracils in single-stranded DNA. Chemical inhibition of A3 enzymes may yield an antimutation therapeutic strategy to improve the durability of current drug therapies that are prone to resistance mutations. A3 small-molecule drug discovery efforts to date have been restricted to a single high-throughput biochemical activity assay; however, the arsenal of discovery assays has significantly expanded in recent years. The assays used to study A3 enzymes are reviewed here with an eye towards their potential for small-molecule discovery efforts.
Collapse
|
|
10
|
Bandarra S, Miyagi E, Ribeiro AC, Gonçalves J, Strebel K, Barahona I. APOBEC3B Potently Restricts HIV-2 but Not HIV-1 in a Vif-Dependent Manner. J Virol 2021; 95:e0117021. [PMID: 34523960 PMCID: PMC8577350 DOI: 10.1128/jvi.01170-21] [Citation(s) in RCA: 4] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/05/2021] [Accepted: 09/03/2021] [Indexed: 11/20/2022] Open
Abstract
Vif is a lentiviral accessory protein that counteracts the antiviral activity of cellular APOBEC3 (A3) cytidine deaminases in infected cells. The exact contribution of each member of the A3 family for the restriction of HIV-2 is still unclear. Thus, the aim of this work was to identify the A3s with anti-HIV-2 activity and compare their restriction potential for HIV-2 and HIV-1. We found that A3G is a strong restriction factor of both types of viruses and A3C restricts neither HIV-1 nor HIV-2. Importantly, A3B exhibited potent antiviral activity against HIV-2, but its effect was negligible against HIV-1. Whereas A3B is packaged with similar efficiency into both viruses in the absence of Vif, HIV-2 and HIV-1 differ in their sensitivity to A3B. HIV-2 Vif targets A3B by reducing its cellular levels and inhibiting its packaging into virions, whereas HIV-1 Vif did not evolve to antagonize A3B. Our observations support the hypothesis that during wild-type HIV-1 and HIV-2 infections, both viruses are able to replicate in host cells expressing A3B but using different mechanisms, probably resulting from a Vif functional adaptation over evolutionary time. Our findings provide new insights into the differences between Vif protein and their cellular partners in the two human viruses. Of note, A3B is highly expressed in some cancer cells and may cause deamination-induced mutations in these cancers. Thus, A3B may represent an important therapeutic target. As such, the ability of HIV-2 Vif to induce A3B degradation could be an effective tool for cancer therapy. IMPORTANCE Primate lentiviruses encode a series of accessory genes that facilitate virus adaptation to its host. Among those, the vif-encoded protein functions primarily by targeting the APOBEC3 (A3) family of cytidine deaminases. All lentiviral Vif proteins have the ability to antagonize A3G; however, antagonizing other members of the A3 family is variable. Here, we report that HIV-2 Vif, unlike HIV-1 Vif, can induce degradation of A3B. Consequently, HIV-2 Vif but not HIV-1 Vif can inhibit the packaging of A3B. Interestingly, while A3B is packaged efficiently into the core of both HIV-1 and HIV-2 virions in the absence of Vif, it only affects the infectivity of HIV-2 particles. Thus, HIV-1 and HIV-2 have evolved two distinct mechanisms to antagonize the antiviral activity of A3B. Aside from its antiviral activity, A3B has been associated with mutations in some cancers. Degradation of A3B by HIV-2 Vif may be useful for cancer therapies.
Collapse
Affiliation(s)
- Susana Bandarra
- Centro de investigação interdisciplinar Egas Moniz (CiiEM), Instituto Universitário Egas Moniz, Quinta da Granja, Caparica, Portugal
- Research Institute for Medicines (iMed.ULisboa), Faculty of Pharmacy, University of Lisbon, Lisbon, Portugal
| | - Eri Miyagi
- Laboratory of Molecular Microbiology, Viral Biochemistry Section, National Institute of Allergy and Infectious Diseases, NIH, Bethesda, Maryland, USA
| | - Ana Clara Ribeiro
- Centro de investigação interdisciplinar Egas Moniz (CiiEM), Instituto Universitário Egas Moniz, Quinta da Granja, Caparica, Portugal
| | - João Gonçalves
- Research Institute for Medicines (iMed.ULisboa), Faculty of Pharmacy, University of Lisbon, Lisbon, Portugal
| | - Klaus Strebel
- Laboratory of Molecular Microbiology, Viral Biochemistry Section, National Institute of Allergy and Infectious Diseases, NIH, Bethesda, Maryland, USA
| | - Isabel Barahona
- Centro de investigação interdisciplinar Egas Moniz (CiiEM), Instituto Universitário Egas Moniz, Quinta da Granja, Caparica, Portugal
| |
Collapse
|
|
11
|
Chen Z, Eggerman TL, Bocharov AV, Baranova IN, Vishnyakova TG, Patterson AP. APOBEC3-induced mutation of the hepatitis virus B DNA genome occurs during its viral RNA reverse transcription into (-)-DNA. J Biol Chem 2021; 297:100889. [PMID: 34181944 PMCID: PMC8321922 DOI: 10.1016/j.jbc.2021.100889] [Citation(s) in RCA: 9] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/24/2021] [Revised: 06/09/2021] [Accepted: 06/16/2021] [Indexed: 11/17/2022] Open
Abstract
APOBEC3s are innate single-stranded DNA cytidine-to-uridine deaminases that catalyze mutations in both pathogen and human genomes with significant roles in human disease. However, how APOBEC3s mutate a single-stranded DNA that is available momentarily during DNA transcription or replication in vivo remains relatively unknown. In this study, utilizing hepatitis B virus (HBV) viral mutations, we evaluated the mutational characteristics of individual APOBEC3s with reference to the HBV replication process through HBV whole single-strand (-)-DNA genome mutation analyses. We found that APOBEC3s induced C-to-T mutations from the HBV reverse transcription start site continuing through the whole (-)-DNA transcript to the termination site with variable efficiency, in an order of A3B >> A3G > A3H-II or A3C. A3B had a 3-fold higher mutation efficiency than A3H-II or A3C with up to 65% of all HBV genomic cytidines being converted into uridines in a single mutation event, consistent with the A3B localized hypermutation signature in cancer, namely, kataegis. On the other hand, A3C expression led to a 3-fold higher number of mutation-positive HBV genome clones, although each individual clone had a lower number of C-to-T mutations. Like A3B, A3C preferred both 5'-TC and 5'-CC sequences, but to a lesser degree. The APOBEC3-induced HBV mutations were predominantly detected in the HBV rcDNA but were not detectable in other intermediates including HBV cccDNA and pgRNA by primer extension of their PCR amplification products. These data demonstrate that APOBEC3-induced HBV genome mutations occur predominantly when the HBV RNA genome was reversely transcribed into (-)-DNA in the viral capsid.
Collapse
Affiliation(s)
- Zhigang Chen
- Department of Laboratory Medicine, Clinical Center, National Institutes of Health, Bethesda, Maryland, USA
| | - Thomas L Eggerman
- Department of Laboratory Medicine, Clinical Center, National Institutes of Health, Bethesda, Maryland, USA; Division of Diabetes, Endocrinology and Metabolic Diseases, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, Maryland, USA
| | - Alexander V Bocharov
- Department of Laboratory Medicine, Clinical Center, National Institutes of Health, Bethesda, Maryland, USA
| | - Irina N Baranova
- Department of Laboratory Medicine, Clinical Center, National Institutes of Health, Bethesda, Maryland, USA
| | - Tatyana G Vishnyakova
- Department of Laboratory Medicine, Clinical Center, National Institutes of Health, Bethesda, Maryland, USA
| | - Amy P Patterson
- Department of Laboratory Medicine, Clinical Center, National Institutes of Health, Bethesda, Maryland, USA; National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, Maryland, USA.
| |
Collapse
|
|
12
|
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.
Collapse
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
| |
Collapse
|
|
13
|
Gaba A, Flath B, Chelico L. Examination of the APOBEC3 Barrier to Cross Species Transmission of Primate Lentiviruses. Viruses 2021; 13:1084. [PMID: 34200141 PMCID: PMC8228377 DOI: 10.3390/v13061084] [Citation(s) in RCA: 17] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/30/2021] [Revised: 05/31/2021] [Accepted: 06/02/2021] [Indexed: 12/12/2022] Open
Abstract
The transmission of viruses from animal hosts into humans have led to the emergence of several diseases. Usually these cross-species transmissions are blocked by host restriction factors, which are proteins that can block virus replication at a specific step. In the natural virus host, the restriction factor activity is usually suppressed by a viral antagonist protein, but this is not the case for restriction factors from an unnatural host. However, due to ongoing viral evolution, sometimes the viral antagonist can evolve to suppress restriction factors in a new host, enabling cross-species transmission. Here we examine the classical case of this paradigm by reviewing research on APOBEC3 restriction factors and how they can suppress human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV). APOBEC3 enzymes are single-stranded DNA cytidine deaminases that can induce mutagenesis of proviral DNA by catalyzing the conversion of cytidine to promutagenic uridine on single-stranded viral (-)DNA if they escape the HIV/SIV antagonist protein, Vif. APOBEC3 degradation is induced by Vif through the proteasome pathway. SIV has been transmitted between Old World Monkeys and to hominids. Here we examine the adaptations that enabled such events and the ongoing impact of the APOBEC3-Vif interface on HIV in humans.
Collapse
Affiliation(s)
- Amit Gaba
- Department of Biochemistry, Microbiology, and Immunology, University of Saskatchewan, Saskatoon, SA S7H 0E5, Canada
| | - Ben Flath
- Department of Biochemistry, Microbiology, and Immunology, University of Saskatchewan, Saskatoon, SA S7H 0E5, Canada
| | - Linda Chelico
- Department of Biochemistry, Microbiology, and Immunology, University of Saskatchewan, Saskatoon, SA S7H 0E5, Canada
| |
Collapse
|
|
14
|
Cotroneo CE, Mangano N, Dragani TA, Colombo F. Lung expression of genes putatively involved in SARS-CoV-2 infection is modulated in cis by germline variants. Eur J Hum Genet 2021; 29:1019-1026. [PMID: 33649539 PMCID: PMC7917374 DOI: 10.1038/s41431-021-00831-y] [Citation(s) in RCA: 6] [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: 07/01/2020] [Revised: 12/09/2020] [Accepted: 02/09/2021] [Indexed: 01/10/2023] Open
Abstract
Germline variants in genes involved in SARS-CoV-2 cell entry and in host innate immune responses to viruses may influence the susceptibility to infection. This study used whole-genome analyses of lung tissue to identify polymorphisms acting as expression quantitative trait loci (eQTLs) for 60 genes of relevance to SARS-CoV-2 infection susceptibility. The expression of genes with confirmed or possible roles in viral entry-replication and in host antiviral responses was studied in the non-diseased lung tissue of 408 lung adenocarcinoma patients. No gene was differently expressed by sex, but APOBEC3H levels were higher and PARP12 levels lower in older individuals. A total of 125 cis-eQTLs (false discovery rate < 0.05) was found to modulate mRNA expression of 15 genes (ABO, ANPEP, AP2A2, APOBEC3D, APOBEC3G, BSG, CLEC4G, DDX58, DPP4, FURIN, FYCO1, RAB14, SERINC3, TRIM5, ZCRB1). eQTLs regulating ABO and FYCO1 were found in COVID-19 susceptibility loci. No trans-eQTLs were identified. Genetic control of the expression of these 15 genes, which encode putative virus receptors, proteins required for vesicle trafficking, enzymes that interfere with viral replication, and other restriction factors, may underlie interindividual differences in risk or severity of infection with SARS-CoV-2 or other viruses.
Collapse
Affiliation(s)
- Chiara E Cotroneo
- Department of Research, Fondazione IRCCS Istituto Nazionale dei Tumori, Milan, Italy
| | - Nunzia Mangano
- Department of Research, Fondazione IRCCS Istituto Nazionale dei Tumori, Milan, Italy
| | - Tommaso A Dragani
- Department of Research, Fondazione IRCCS Istituto Nazionale dei Tumori, Milan, Italy.
| | - Francesca Colombo
- Institute of Biomedical Technologies, National Research Council (ITB-CNR), Segrate, MI, Italy
| |
Collapse
|
|
15
|
Hu Y, Knecht KM, Shen Q, Xiong Y. Multifaceted HIV-1 Vif interactions with human E3 ubiquitin ligase and APOBEC3s. FEBS J 2021; 288:3407-3417. [PMID: 32893454 PMCID: PMC8172064 DOI: 10.1111/febs.15550] [Citation(s) in RCA: 8] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/17/2020] [Accepted: 09/01/2020] [Indexed: 12/31/2022]
Abstract
APOBEC3 (A3) proteins are a family of host antiviral restriction factors that potently inhibit various retroviral infections, including human immunodeficiency virus (HIV)-1. To overcome this restriction, HIV-1 virion infectivity factor (Vif) recruits the cellular cofactor CBFβ to assist in targeting A3 proteins to a host E3 ligase complex for polyubiquitination and subsequent proteasomal degradation. Intervention of the Vif-A3 interactions could be a promising therapeutic strategy to facilitate A3-mediated suppression of HIV-1 in patients. In this structural snapshot, we review the structural features of the recently determined structure of human A3F in complex with HIV-1 Vif and its cofactor CBFβ, discuss insights into the molecular principles of Vif-A3 interplay during the arms race between the virus and host, and highlight the therapeutic implications.
Collapse
Affiliation(s)
- Yingxia Hu
- Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT, USA
| | - Kirsten M. Knecht
- Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT, USA
| | - Qi Shen
- Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT, USA
| | - Yong Xiong
- Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, CT, USA
| |
Collapse
|
|
16
|
Rouf Banday A, Onabajo OO, Lin SHY, Obajemu A, Vargas JM, Delviks-Frankenberry KA, Lamy P, Bayanjargal A, Zettelmeyer C, Florez-Vargas O, Pathak VK, Dyrskjøt L, Prokunina-Olsson L. Targeting natural splicing plasticity of APOBEC3B restricts its expression and mutagenic activity. Commun Biol 2021; 4:386. [PMID: 33753867 PMCID: PMC7985488 DOI: 10.1038/s42003-021-01844-5] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/20/2020] [Accepted: 02/11/2021] [Indexed: 12/14/2022] Open
Abstract
APOBEC3A (A3A) and APOBEC3B (A3B) enzymes drive APOBEC-mediated mutagenesis. Identification of factors affecting the activity of these enzymes could help modulate mutagenesis and associated clinical outcomes. Here, we show that canonical and alternatively spliced A3A and A3B isoforms produce corresponding mutagenic and non-mutagenic enzymes. Increased expression of the mutagenic A3B isoform predicted shorter progression-free survival in bladder cancer. We demonstrate that the production of mutagenic vs. non-mutagenic A3B protein isoforms was considerably affected by inclusion/skipping of exon 5 in A3B. Furthermore, exon 5 skipping, resulting in lower levels of mutagenic A3B enzyme, could be increased in vitro. Specifically, we showed the effects of treatment with an SF3B1 inhibitor affecting spliceosome interaction with a branch point site in intron 4, or with splice-switching oligonucleotides targeting exon 5 of A3B. Our results underscore the clinical role of A3B and implicate alternative splicing of A3B as a mechanism that could be targeted to restrict APOBEC-mediated mutagenesis.
Collapse
Affiliation(s)
- A Rouf Banday
- Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, National Cancer Institute, National Institutes of Health, Bethesda, MD, USA
| | - Olusegun O Onabajo
- Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, National Cancer Institute, National Institutes of Health, Bethesda, MD, USA
| | - Seraph Han-Yin Lin
- Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, National Cancer Institute, National Institutes of Health, Bethesda, MD, USA
| | - Adeola Obajemu
- Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, National Cancer Institute, National Institutes of Health, Bethesda, MD, USA
| | - Joselin M Vargas
- Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, National Cancer Institute, National Institutes of Health, Bethesda, MD, USA
| | - Krista A Delviks-Frankenberry
- Viral Mutation Section, HIV Dynamics and Replication Program, Center for Cancer Research, National Cancer Institute, Frederick, MD, USA
| | - Philippe Lamy
- Department of Molecular Medicine, Aarhus University Hospital, Aarhus, Denmark
| | - Ariunaa Bayanjargal
- Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, National Cancer Institute, National Institutes of Health, Bethesda, MD, USA
| | - Clara Zettelmeyer
- Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, National Cancer Institute, National Institutes of Health, Bethesda, MD, USA
| | - Oscar Florez-Vargas
- Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, National Cancer Institute, National Institutes of Health, Bethesda, MD, USA
| | - Vinay K Pathak
- Viral Mutation Section, HIV Dynamics and Replication Program, Center for Cancer Research, National Cancer Institute, Frederick, MD, USA
| | - Lars Dyrskjøt
- Department of Molecular Medicine, Aarhus University Hospital, Aarhus, Denmark
| | - Ludmila Prokunina-Olsson
- Laboratory of Translational Genomics, Division of Cancer Epidemiology and Genetics, National Cancer Institute, National Institutes of Health, Bethesda, MD, USA.
| |
Collapse
|
|
17
|
Insights into the Structures and Multimeric Status of APOBEC Proteins Involved in Viral Restriction and Other Cellular Functions. Viruses 2021; 13:v13030497. [PMID: 33802945 PMCID: PMC8002816 DOI: 10.3390/v13030497] [Citation(s) in RCA: 15] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/15/2021] [Revised: 03/15/2021] [Accepted: 03/15/2021] [Indexed: 12/16/2022] Open
Abstract
Apolipoprotein B mRNA editing catalytic polypeptide-like (APOBEC) proteins belong to a family of deaminase proteins that can catalyze the deamination of cytosine to uracil on single-stranded DNA or/and RNA. APOBEC proteins are involved in diverse biological functions, including adaptive and innate immunity, which are critical for restricting viral infection and endogenous retroelements. Dysregulation of their functions can cause undesired genomic mutations and RNA modification, leading to various associated diseases, such as hyper-IgM syndrome and cancer. This review focuses on the structural and biochemical data on the multimerization status of individual APOBECs and the associated functional implications. Many APOBECs form various multimeric complexes, and multimerization is an important way to regulate functions for some of these proteins at several levels, such as deaminase activity, protein stability, subcellular localization, protein storage and activation, virion packaging, and antiviral activity. The multimerization of some APOBECs is more complicated than others, due to the associated complex RNA binding modes.
Collapse
|
|
18
|
Potential APOBEC-mediated RNA editing of the genomes of SARS-CoV-2 and other coronaviruses and its impact on their longer term evolution. Virology 2021; 556:62-72. [PMID: 33545556 PMCID: PMC7831814 DOI: 10.1016/j.virol.2020.12.018] [Citation(s) in RCA: 77] [Impact Index Per Article: 19.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/21/2020] [Revised: 12/21/2020] [Accepted: 12/21/2020] [Indexed: 12/19/2022]
Abstract
Members of the APOBEC family of cytidine deaminases show antiviral activities in mammalian cells through lethal editing in the genomes of small DNA viruses, herpesviruses and retroviruses, and potentially those of RNA viruses such as coronaviruses. Consistent with the latter, APOBEC-like directional C→U transitions of genomic plus-strand RNA are greatly overrepresented in SARS-CoV-2 genome sequences of variants emerging during the COVID-19 pandemic. A C→U mutational process may leave evolutionary imprints on coronavirus genomes, including extensive homoplasy from editing and reversion at targeted sites and the occurrence of driven amino acid sequence changes in viral proteins. If sustained over longer periods, this process may account for the previously reported marked global depletion of C and excess of U bases in human seasonal coronavirus genomes. This review synthesizes the current knowledge on APOBEC evolution and function and the evidence of their role in APOBEC-mediated genome editing of SARS-CoV-2 and other coronaviruses.
SARS-CoV-2 sequence variants contain an overabundance of C- > U transitions C- > U transitions are the hallmark of the activity of APOBEC cytosine deaminases Further work is needed to determine APOBEC's role in coronavirus evolution
Collapse
|
|
19
|
Wei Y, Silke JR, Aris P, Xia X. Coronavirus genomes carry the signatures of their habitats. PLoS One 2020; 15:e0244025. [PMID: 33351847 PMCID: PMC7755226 DOI: 10.1371/journal.pone.0244025] [Citation(s) in RCA: 21] [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: 08/19/2020] [Accepted: 12/01/2020] [Indexed: 12/15/2022] Open
Abstract
Coronaviruses such as SARS-CoV-2 regularly infect host tissues that express antiviral proteins (AVPs) in abundance. Understanding how they evolve to adapt or evade host immune responses is important in the effort to control the spread of infection. Two AVPs that may shape viral genomes are the zinc finger antiviral protein (ZAP) and the apolipoprotein B mRNA editing enzyme-catalytic polypeptide-like 3 (APOBEC3). The former binds to CpG dinucleotides to facilitate the degradation of viral transcripts while the latter frequently deaminates C into U residues which could generate notable viral sequence variations. We tested the hypothesis that both APOBEC3 and ZAP impose selective pressures that shape the genome of an infecting coronavirus. Our investigation considered a comprehensive number of publicly available genomes for seven coronaviruses (SARS-CoV-2, SARS-CoV, and MERS infecting Homo sapiens, Bovine CoV infecting Bos taurus, MHV infecting Mus musculus, HEV infecting Sus scrofa, and CRCoV infecting Canis lupus familiaris). We show that coronaviruses that regularly infect tissues with abundant AVPs have CpG-deficient and U-rich genomes; whereas those that do not infect tissues with abundant AVPs do not share these sequence hallmarks. Among the coronaviruses surveyed herein, CpG is most deficient in SARS-CoV-2 and a temporal analysis showed a marked increase in C to U mutations over four months of SARS-CoV-2 genome evolution. Furthermore, the preferred motifs in which these C to U mutations occur are the same as those subjected to APOBEC3 editing in HIV-1. These results suggest that both ZAP and APOBEC3 shape the SARS-CoV-2 genome: ZAP imposes a strong CpG avoidance, and APOBEC3 constantly edits C to U. Evolutionary pressures exerted by host immune systems onto viral genomes may motivate novel strategies for SARS-CoV-2 vaccine development.
Collapse
Affiliation(s)
- Yulong Wei
- Department of Biology, University of Ottawa, Ottawa, Ontario, Canada
| | - Jordan R. Silke
- Department of Biology, University of Ottawa, Ottawa, Ontario, Canada
| | - Parisa Aris
- Department of Biology, University of Ottawa, Ottawa, Ontario, Canada
| | - Xuhua Xia
- Department of Biology, University of Ottawa, Ottawa, Ontario, Canada
- Ottawa Institute of Systems Biology, University of Ottawa, Ottawa, Ontario, Canada
| |
Collapse
|
|
20
|
Retroviral Restriction Factors and Their Viral Targets: Restriction Strategies and Evolutionary Adaptations. Microorganisms 2020; 8:microorganisms8121965. [PMID: 33322320 PMCID: PMC7764263 DOI: 10.3390/microorganisms8121965] [Citation(s) in RCA: 19] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/30/2020] [Revised: 11/30/2020] [Accepted: 12/08/2020] [Indexed: 12/17/2022] Open
Abstract
The evolutionary conflict between retroviruses and their vertebrate hosts over millions of years has led to the emergence of cellular innate immune proteins termed restriction factors as well as their viral antagonists. Evidence accumulated in the last two decades has substantially increased our understanding of the elaborate mechanisms utilized by these restriction factors to inhibit retroviral replication, mechanisms that either directly block viral proteins or interfere with the cellular pathways hijacked by the viruses. Analyses of these complex interactions describe patterns of accelerated evolution for these restriction factors as well as the acquisition and evolution of their virus-encoded antagonists. Evidence is also mounting that many restriction factors identified for their inhibition of specific retroviruses have broader antiviral activity against additional retroviruses as well as against other viruses, and that exposure to these multiple virus challenges has shaped their adaptive evolution. In this review, we provide an overview of the restriction factors that interfere with different steps of the retroviral life cycle, describing their mechanisms of action, adaptive evolution, viral targets and the viral antagonists that evolved to counter these factors.
Collapse
|
|
21
|
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.
Collapse
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.)
| |
Collapse
|
|
22
|
Chesarino NM, Emerman M. Polymorphisms in Human APOBEC3H Differentially Regulate Ubiquitination and Antiviral Activity. Viruses 2020; 12:E378. [PMID: 32235597 PMCID: PMC7232234 DOI: 10.3390/v12040378] [Citation(s) in RCA: 12] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/07/2020] [Revised: 03/25/2020] [Accepted: 03/27/2020] [Indexed: 12/14/2022] Open
Abstract
The APOBEC3 family of cytidine deaminases are an important part of the host innate immune defense against endogenous retroelements and retroviruses like Human Immunodeficiency Virus (HIV). APOBEC3H (A3H) is the most polymorphic of the human APOBEC3 genes, with four major haplotypes circulating in the population. Haplotype II is the only antivirally-active variant of A3H, while the majority of the population possess independently destabilizing polymorphisms present in haplotype I (R105G) and haplotypes III and IV (N15del). In this paper, we show that instability introduced by either polymorphism is positively correlated with degradative ubiquitination, while haplotype II is protected from this modification. Inhibiting ubiquitination by mutating all of the A3H lysines increased the expression of haplotypes III and IV, but these stabilized forms of haplotype III and IV had a strict nuclear localization, and did not incorporate into virions, nor exhibit antiviral activity. Fusion chimeras with haplotype II allowed for stabilization, cytoplasmic retention, and packaging of the N15del-containing haplotype III, but the haplotype III component of these chimeras was unable to restrict HIV-1 on its own. Thus, the evolutionary loss of A3H activity in many humans involves functional deficiencies independent of protein stability.
Collapse
Affiliation(s)
| | - Michael Emerman
- Divisions of Human Biology and Basic Sciences, Fred Hutchinson Cancer Research Center, Seattle, WA 98109, USA;
| |
Collapse
|
|
23
|
Azimi FC, Lee JE. Structural perspectives on HIV-1 Vif and APOBEC3 restriction factor interactions. Protein Sci 2020; 29:391-406. [PMID: 31518043 PMCID: PMC6954718 DOI: 10.1002/pro.3729] [Citation(s) in RCA: 16] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/16/2019] [Revised: 09/08/2019] [Accepted: 09/09/2019] [Indexed: 11/06/2022]
Abstract
Human immunodeficiency virus (HIV) is a retroviral pathogen that targets human immune cells such as CD4+ T cells, macrophages, and dendritic cells. The human apolipoprotein B mRNA- editing catalytic polypeptide 3 (APOBEC3 or A3) cytidine deaminases are a key class of intrinsic restriction factors that inhibit replication of HIV. When HIV-1 enters the cell, the immune system responds by inducing the activation of the A3 family proteins, which convert cytosines to uracils in single-stranded DNA replication intermediates, neutralizing the virus. HIV counteracts this intrinsic immune response by encoding a protein termed viral infectivity factor (Vif). Vif targets A3 to an E3 ubiquitin ligase complex for poly-ubiquitination and proteasomal degradation. Vif is unique in that it can recognize and counteract multiple A3 restriction factor substrates. Structural biology studies have provided significant insights into the overall architectures and functions of Vif and A3 proteins; however, a structure of the Vif-A3 complex has remained elusive. In this review, we summarize and reanalyze experimental data from recent structural, biochemical, and functional studies to provide key perspectives on the residues involved in Vif-A3 protein-protein interactions.
Collapse
Affiliation(s)
- Farshad C. Azimi
- Department of Laboratory Medicine and Pathobiology, Faculty of MedicineUniversity of TorontoTorontoOntarioCanada
| | - Jeffrey E. Lee
- Department of Laboratory Medicine and Pathobiology, Faculty of MedicineUniversity of TorontoTorontoOntarioCanada
| |
Collapse
|
|
24
|
Wang J, Becker JT, Shi K, Lauer KV, Salamango DJ, Aihara H, Shaban NM, Harris RS. The Role of RNA in HIV-1 Vif-Mediated Degradation of APOBEC3H. J Mol Biol 2019; 431:5019-5031. [PMID: 31628948 DOI: 10.1016/j.jmb.2019.09.014] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/10/2019] [Revised: 09/04/2019] [Accepted: 09/16/2019] [Indexed: 11/17/2022]
Abstract
As many as five members of the APOBEC3 family of DNA cytosine deaminases are capable of inhibiting HIV-1 replication by deaminating viral cDNA cytosines and interfering with reverse transcription. HIV-1 counteracts restriction with the virally encoded Vif protein, which forms a hybrid ubiquitin ligase complex that directly binds APOBEC3 enzymes and targets them for proteasomal degradation. APOBEC3H (A3H) is unique among family members by dimerization through cellular and viral duplex RNA species. RNA binding is required for localization of A3H to the cytoplasmic compartment, for efficient packaging into nascent HIV-1 particles and ultimately for effective virus restriction activity. Here we compared wild-type human A3H and RNA binding-defective mutants to ask whether RNA may be a factor in the functional interaction with HIV-1 Vif. We used structural modeling, immunoblotting, live cell imaging, and split green fluorescence protein (GFP) reconstitution approaches to assess the capability of HIV-1 Vif to promote the degradation of wild-type A3H in comparison to RNA binding-defective mutants. The results combined to show that RNA is not strictly required for Vif-mediated degradation of A3H, and that RNA and Vif are likely to bind this single-domain DNA cytosine deaminase on physically distinct surfaces. However, a subset of the results also indicated that the A3H degradation process may be affected by A3H protein structure, subcellular localization, and differences in the constellation of A3H interaction partners, suggesting additional factors may also influence the fate and functionality of this host-pathogen interaction.
Collapse
Affiliation(s)
- Jiayi Wang
- Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, MN 55455, USA; Masonic Cancer Center, University of Minnesota, Minneapolis, MN 55455, USA; Institute for Molecular Virology, University of Minnesota, Minneapolis, MN 55455, USA; Center for Genome Engineering, University of Minnesota, Minneapolis, MN 55455, USA
| | - Jordan T Becker
- Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, MN 55455, USA; Masonic Cancer Center, University of Minnesota, Minneapolis, MN 55455, USA; Institute for Molecular Virology, University of Minnesota, Minneapolis, MN 55455, USA; Center for Genome Engineering, University of Minnesota, Minneapolis, MN 55455, USA
| | - Ke Shi
- Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, MN 55455, USA; Masonic Cancer Center, University of Minnesota, Minneapolis, MN 55455, USA; Institute for Molecular Virology, University of Minnesota, Minneapolis, MN 55455, USA
| | - Kate V Lauer
- Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, MN 55455, USA; Masonic Cancer Center, University of Minnesota, Minneapolis, MN 55455, USA; Institute for Molecular Virology, University of Minnesota, Minneapolis, MN 55455, USA; Center for Genome Engineering, University of Minnesota, Minneapolis, MN 55455, USA
| | - Daniel J Salamango
- Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, MN 55455, USA; Masonic Cancer Center, University of Minnesota, Minneapolis, MN 55455, USA; Institute for Molecular Virology, University of Minnesota, Minneapolis, MN 55455, USA; Center for Genome Engineering, University of Minnesota, Minneapolis, MN 55455, USA
| | - Hideki Aihara
- Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, MN 55455, USA; Masonic Cancer Center, University of Minnesota, Minneapolis, MN 55455, USA; Institute for Molecular Virology, University of Minnesota, Minneapolis, MN 55455, USA
| | - Nadine M Shaban
- Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, MN 55455, USA; Masonic Cancer Center, University of Minnesota, Minneapolis, MN 55455, USA; Institute for Molecular Virology, University of Minnesota, Minneapolis, MN 55455, USA; Center for Genome Engineering, University of Minnesota, Minneapolis, MN 55455, USA.
| | - Reuben S Harris
- Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, MN 55455, USA; Masonic Cancer Center, University of Minnesota, Minneapolis, MN 55455, USA; Institute for Molecular Virology, University of Minnesota, Minneapolis, MN 55455, USA; Center for Genome Engineering, University of Minnesota, Minneapolis, MN 55455, USA; Howard Hughes Medical Institute, University of Minnesota, Minneapolis, MN 55455, USA.
| |
Collapse
|
|
25
|
Different antiviral activities of natural APOBEC3C, APOBEC3G, and APOBEC3H variants against hepatitis B virus. Biochem Biophys Res Commun 2019; 518:26-31. [PMID: 31400856 DOI: 10.1016/j.bbrc.2019.08.003] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/11/2019] [Accepted: 08/01/2019] [Indexed: 12/29/2022]
Abstract
Some APOBEC3 family members have antiviral activity against retroviruses and DNA viruses. Hepatitis B virus (HBV) is a DNA virus that is the major causative factor of severe liver diseases such as cirrhosis and hepatocellular carcinoma. To determine whether APOBEC3 variants in humans have different anti-HBV activities, we evaluated natural variants of APOBEC3C, APOBEC3G, and APOBEC3H using an HBV-replicating cell culture model. Our data demonstrate that the APOBEC3C variant S188I had increased restriction activity and hypermutation frequency against HBV DNA. In contrast, the APOBEC3G variant H186R did not alter the anti-HBV and hypermutation activities. Among APOBEC3H polymorphisms (hap I-VII) and splicing variants (SV-200, SV-183, SV-182, and SV-154), hap II SV-183 showed the strongest restriction activity. These data suggest that the genetic variations in APOBEC3 genes may affect the efficiency of HBV elimination in humans.
Collapse
|
|
26
|
Matsuoka T, Nagae T, Ode H, Awazu H, Kurosawa T, Hamano A, Matsuoka K, Hachiya A, Imahashi M, Yokomaku Y, Watanabe N, Iwatani Y. Structural basis of chimpanzee APOBEC3H dimerization stabilized by double-stranded RNA. Nucleic Acids Res 2019; 46:10368-10379. [PMID: 30060196 PMCID: PMC6212771 DOI: 10.1093/nar/gky676] [Citation(s) in RCA: 20] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/01/2018] [Accepted: 07/17/2018] [Indexed: 12/22/2022] Open
Abstract
APOBEC3H (A3H) is a mammal-specific cytidine deaminase that potently restricts the replication of retroviruses. Primate A3Hs are known to exert key selective pressures against the cross-species transmission of primate immunodeficiency viruses from chimpanzees to humans. Despite recent advances, the molecular structures underlying the functional mechanisms of primate A3Hs have not been fully understood. Here, we reveal the 2.20-Å crystal structure of the chimpanzee A3H (cpzA3H) dimer bound to a short double-stranded RNA (dsRNA), which appears to be similar to two recently reported structures of pig-tailed macaque A3H and human A3H. In the structure, the dsRNA-binding interface forms a specialized architecture with unique features. The analysis of the dsRNA nucleotides in the cpzA3H complex revealed the GC-rich palindrome-like sequence preference for dsRNA interaction, which is largely determined by arginine residues in loop 1. In cells, alterations of the cpzA3H residues critical for the dsRNA interaction severely reduce intracellular protein stability due to proteasomal degradation. This suggests that cpzA3H stability is regulated by the dsRNA-mediated dimerization as well as by unknown cellular machinery through proteasomal degradation in cells. Taken together, these findings highlight unique structural features of primate A3Hs that are important to further understand their cellular functions and regulation.
Collapse
Affiliation(s)
- Tatsuya Matsuoka
- Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi 460-0001, Japan.,Department of Biotechnology, Nagoya University Graduate School of Engineering, Nagoya, Aichi 464-8603, Japan
| | - Takayuki Nagae
- Synchrotron Radiation Research Center, Nagoya University, Nagoya, Aichi 464-8603, Japan
| | - Hirotaka Ode
- Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi 460-0001, Japan
| | - Hiroaki Awazu
- Department of Biotechnology, Nagoya University Graduate School of Engineering, Nagoya, Aichi 464-8603, Japan
| | - Teppei Kurosawa
- Department of Biotechnology, Nagoya University Graduate School of Engineering, Nagoya, Aichi 464-8603, Japan
| | - Akiko Hamano
- Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi 460-0001, Japan
| | - Kazuhiro Matsuoka
- Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi 460-0001, Japan
| | - Atsuko Hachiya
- Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi 460-0001, Japan
| | - Mayumi Imahashi
- Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi 460-0001, Japan
| | - Yoshiyuki Yokomaku
- Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi 460-0001, Japan
| | - Nobuhisa Watanabe
- Department of Biotechnology, Nagoya University Graduate School of Engineering, Nagoya, Aichi 464-8603, Japan.,Synchrotron Radiation Research Center, Nagoya University, Nagoya, Aichi 464-8603, Japan
| | - Yasumasa Iwatani
- Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi 460-0001, Japan.,Program in Integrated Molecular Medicine, Nagoya University Graduate School of Medicine, Nagoya, Aichi 466-8550, Japan
| |
Collapse
|
|
27
|
Role of co-expressed APOBEC3F and APOBEC3G in inducing HIV-1 drug resistance. Heliyon 2019; 5:e01498. [PMID: 31025011 PMCID: PMC6475876 DOI: 10.1016/j.heliyon.2019.e01498] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/09/2019] [Revised: 03/24/2019] [Accepted: 04/05/2019] [Indexed: 01/04/2023] Open
Abstract
The APOBEC3 enzymes can induce mutagenesis of HIV-1 proviral DNA through the deamination of cytosine. HIV-1 overcomes this restriction through the viral protein Vif that induces APOBEC3 proteasomal degradation. Within this dynamic host-pathogen relationship, the APOBEC3 enzymes have been found to be beneficial, neutral, or detrimental to HIV-1 biology. Here, we assessed the ability of co-expressed APOBEC3F and APOBEC3G to induce HIV-1 resistance to antiviral drugs. We found that co-expression of APOBEC3F and APOBEC3G enabled partial resistance of APOBEC3F to Vif-mediated degradation with a corresponding increase in APOBEC3F-induced deaminations in the presence of Vif, in addition to APOBEC3G-induced deaminations. We recovered HIV-1 drug resistant variants resulting from APOBEC3-induced mutagenesis, but these variants were less able to replicate than drug resistant viruses derived from RT-induced mutations alone. The data support a model in which APOBEC3 enzymes cooperate to restrict HIV-1, promoting viral inactivation over evolution to drug resistance.
Collapse
|
|
28
|
Mussil B, Suspène R, Caval V, Durandy A, Wain-Hobson S, Vartanian JP. Genotoxic stress increases cytoplasmic mitochondrial DNA editing by human APOBEC3 mutator enzymes at a single cell level. Sci Rep 2019; 9:3109. [PMID: 30816165 PMCID: PMC6395610 DOI: 10.1038/s41598-019-39245-8] [Citation(s) in RCA: 12] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/11/2017] [Accepted: 11/23/2018] [Indexed: 01/23/2023] Open
Abstract
Human cells are stressed by numerous mechanisms that can lead to leakage of mitochondrial DNA (mtDNA) to the cytoplasm and ultimately apoptosis. This agonist DNA constitutes a danger to the cell and is counteracted by cytoplasmic DNases and APOBEC3 cytidine deamination of DNA. To investigate APOBEC3 editing of leaked mtDNA to the cytoplasm, we performed a PCR analysis of APOBEC3 edited cytoplasmic mtDNA (cymtDNA) at the single cell level for primary CD4+ T cells and the established P2 EBV blast cell line. Up to 17% of primary CD4+ T cells showed signs of APOBEC3 edited cymtDNA with ~50% of all mtDNA sequences showing signs of APOBEC3 editing - between 1500-5000 molecules. Although the P2 cell line showed a much lower frequency of stressed cells, the number of edited mtDNA molecules in such cells was of the same order. Addition of the genotoxic molecules, etoposide or actinomycin D increased the number of cells showing APOBEC3 edited cymtDNA to around 40%. These findings reveal a very dynamic image of the mitochondrial network, which changes considerably under stress. APOBEC3 deaminases are involved in the catabolism of mitochondrial DNA to circumvent chronic immune stimulation triggered by released mitochondrial DNA from damaged cells.
Collapse
Affiliation(s)
- Bianka Mussil
- Molecular Retrovirology Unit, Institut Pasteur, CNRS, UMR 3569, 28 rue du Dr. Roux, F-75724, Paris cedex 15, France
- Unit of Infection Models, German Primate Centre, Kellnerweg 4, D-37077, Goettingen, Germany
| | - Rodolphe Suspène
- Molecular Retrovirology Unit, Institut Pasteur, CNRS, UMR 3569, 28 rue du Dr. Roux, F-75724, Paris cedex 15, France
| | - Vincent Caval
- Molecular Retrovirology Unit, Institut Pasteur, CNRS, UMR 3569, 28 rue du Dr. Roux, F-75724, Paris cedex 15, France
| | - Anne Durandy
- INSERM UMR 1163, The Human Lymphohematopoiesis Laboratory, Institut Imagine, 24 boulevard du Montparnasse, F-75015, Paris, France
| | - Simon Wain-Hobson
- Molecular Retrovirology Unit, Institut Pasteur, CNRS, UMR 3569, 28 rue du Dr. Roux, F-75724, Paris cedex 15, France
| | - Jean-Pierre Vartanian
- Molecular Retrovirology Unit, Institut Pasteur, CNRS, UMR 3569, 28 rue du Dr. Roux, F-75724, Paris cedex 15, France.
| |
Collapse
|
|
29
|
Adrian J, Bonsignore P, Hammer S, Frickey T, Hauck CR. Adaptation to Host-Specific Bacterial Pathogens Drives Rapid Evolution of a Human Innate Immune Receptor. Curr Biol 2019; 29:616-630.e5. [PMID: 30744974 DOI: 10.1016/j.cub.2019.01.058] [Citation(s) in RCA: 40] [Impact Index Per Article: 6.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/26/2018] [Revised: 12/12/2018] [Accepted: 01/22/2019] [Indexed: 12/25/2022]
Abstract
The selective pressure by infectious agents is a major driving force in the evolution of humans and other mammals. Members of the carcinoembryonic antigen-related cell adhesion molecule (CEACAM) family serve as receptors for bacterial pathogens of the genera Haemophilus, Helicobacter, Neisseria, and Moraxella, which engage CEACAMs via distinct surface adhesins. While microbial attachment to epithelial CEACAMs facilitates host colonization, recognition by CEACAM3, a phagocytic receptor expressed by granulocytes, eliminates CEACAM-binding bacteria. Sequence analysis of primate CEACAM3 orthologs reveals that this innate immune receptor is one of the most rapidly evolving human proteins. In particular, the pathogen-binding extracellular domain of CEACAM3 shows a high degree of non-synonymous versus synonymous nucleotide exchanges, indicating an exceptionally strong positive selection. Using CEACAM3 domains derived from different primates, we find that the amino acid alterations found in CEACAM3 translate into characteristic binding patterns for bacterial adhesins. One such amino acid residue is F62 in human and chimp CEACAM3, which is not present in other primates and which is critical for binding the OMP P1 adhesin of Haemophilus aegyptius. Incorporation of the F62-containing motif into gorilla CEACAM3 results in a gain-of-function phenotype with regard to phagocytosis of H. aegyptius. Moreover, CEACAM3 polymorphisms found in human subpopulations widen the spectrum of recognized bacterial adhesins, suggesting an ongoing multivariate selection acting on this innate immune receptor. The species-specific detection of diverse bacterial adhesins helps to explain the exceptionally fast evolution of CEACAM3 within the primate lineage and provides an example of Red Queen dynamics in the human genome.
Collapse
Affiliation(s)
- Jonas Adrian
- Lehrstuhl für Zellbiologie, Fachbereich Biologie, Universität Konstanz, Universitätsstraße 10, 78457 Konstanz, Germany
| | - Patrizia Bonsignore
- Lehrstuhl für Zellbiologie, Fachbereich Biologie, Universität Konstanz, Universitätsstraße 10, 78457 Konstanz, Germany
| | - Sebastian Hammer
- Lehrstuhl für Zellbiologie, Fachbereich Biologie, Universität Konstanz, Universitätsstraße 10, 78457 Konstanz, Germany
| | - Tancred Frickey
- Forest Industry Informatics, Scion, Te Papa Tipu Innovation Park, 49 Sala Street, 3015 Rotorua, New Zealand; Konstanz Research School-Chemical Biology, Universität Konstanz, Universitätsstraße 10, 78457 Konstanz, Germany
| | - Christof R Hauck
- Lehrstuhl für Zellbiologie, Fachbereich Biologie, Universität Konstanz, Universitätsstraße 10, 78457 Konstanz, Germany; Konstanz Research School-Chemical Biology, Universität Konstanz, Universitätsstraße 10, 78457 Konstanz, Germany.
| |
Collapse
|
|
30
|
Matume ND, Tebit DM, Gray LR, Turner SD, Rekosh D, Bessong PO, Hammarskjöld ML. Characterization of APOBEC3 variation in a population of HIV-1 infected individuals in northern South Africa. BMC MEDICAL GENETICS 2019; 20:21. [PMID: 30660178 PMCID: PMC6339282 DOI: 10.1186/s12881-018-0740-4] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Subscribe] [Scholar Register] [Received: 07/02/2018] [Accepted: 12/21/2018] [Indexed: 11/10/2022]
Abstract
BACKGROUND The apolipoprotein B mRNA-editing enzyme, catalytic polypeptide-like 3 (APOBEC3) genes A3D, A3F, A3G and A3H have all been implicated in the restriction of human immunodeficiency virus type 1 (HIV-1) replication. Polymorphisms in these genes are likely to impact viral replication and fitness, contributing to viral diversity. Currently, only a few studies indicate that polymorphisms in the A3 genes may be correlated with infection risk and disease progression. METHODS To characterize polymorphisms in the coding regions of these APOBEC3 genes in an HIV-1 infected population from the Limpopo Province of South Africa, APOBEC3 gene fragments were amplified from genomic DNA of 192 HIV-1 infected subjects and sequenced on an Illumina MiSeq platform. SNPs were confirmed and compared to SNPs in other populations reported in the 1000 Genome Phase III and HapMap databases, as well as in the ExAC exome database. Hardy-Weinberg Equilibrium was calculated and haplotypes were inferred using the LDlink 3.0 web tool. Linkage Disequilibrium (LD) for these SNPS were calculated in the total 1000 genome and AFR populations using the same tool. RESULTS Known variants compared to the GRCh37 consensus genome sequence were detected at relatively high frequencies (> 5%) in all of the APOBEC3 genes. A3H showed the most variation, with several of the variants present in both alleles in almost all of the patients. Several minor allele variants (< 5%) were also detected in A3D, A3F and A3G. In addition, novel R6K, L221R and T238I variants in A3D and I117I in A3F were observed. Four, five, four, and three haplotypes were identified for A3D, A3F, A3G, and A3H respectively. CONCLUSIONS The study showed significant polymorphisms in the APOBEC3D, 3F, 3G and 3H genes in our South African HIV1-infected cohort. In the case of all of these genes, the polymorphisms were generally present at higher frequencies than reported in other 1000 genome populations and in the ExAC exome consortium database .
Collapse
Affiliation(s)
- Nontokozo D Matume
- Myles H. Thaler Center for AIDS and Human Retrovirus Research, Department of Microbiology, Immunology and Cancer Biology, University of Virginia, Charlottesville, VA, USA.,HIV/AIDS & Global Health Research Programme and Department of Microbiology, University of Venda, Thohoyandou, South Africa
| | - Denis M Tebit
- Myles H. Thaler Center for AIDS and Human Retrovirus Research, Department of Microbiology, Immunology and Cancer Biology, University of Virginia, Charlottesville, VA, USA.,HIV/AIDS & Global Health Research Programme and Department of Microbiology, University of Venda, Thohoyandou, South Africa.,Global Biomed Scientific LLC, PO Box 2368, Forest, VA, 24551, USA
| | - Laurie R Gray
- Myles H. Thaler Center for AIDS and Human Retrovirus Research, Department of Microbiology, Immunology and Cancer Biology, University of Virginia, Charlottesville, VA, USA
| | - Stephen D Turner
- Department of Public Health Sciences and Bioinformatics Core, University of Virginia School of Medicine, Charlottesville, VA, 22908, USA
| | - David Rekosh
- Myles H. Thaler Center for AIDS and Human Retrovirus Research, Department of Microbiology, Immunology and Cancer Biology, University of Virginia, Charlottesville, VA, USA.,HIV/AIDS & Global Health Research Programme and Department of Microbiology, University of Venda, Thohoyandou, South Africa
| | - Pascal O Bessong
- HIV/AIDS & Global Health Research Programme and Department of Microbiology, University of Venda, Thohoyandou, South Africa.
| | - Marie-Louise Hammarskjöld
- Myles H. Thaler Center for AIDS and Human Retrovirus Research, Department of Microbiology, Immunology and Cancer Biology, University of Virginia, Charlottesville, VA, USA. .,HIV/AIDS & Global Health Research Programme and Department of Microbiology, University of Venda, Thohoyandou, South Africa.
| |
Collapse
|
|
31
|
APOBEC3H Subcellular Localization Determinants Define Zipcode for Targeting HIV-1 for Restriction. Mol Cell Biol 2018; 38:MCB.00356-18. [PMID: 30224517 DOI: 10.1128/mcb.00356-18] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/14/2018] [Accepted: 09/11/2018] [Indexed: 12/22/2022] Open
Abstract
APOBEC enzymes are DNA cytosine deaminases that normally serve as virus restriction factors, but several members, including APOBEC3H, also contribute to cancer mutagenesis. Despite their importance in multiple fields, little is known about cellular processes that regulate these DNA mutating enzymes. We show that APOBEC3H exists in two distinct subcellular compartments, cytoplasm and nucleolus, and that the structural determinants for each mechanism are genetically separable. First, native and fluorescently tagged APOBEC3Hs localize to these two compartments in multiple cell types. Second, a series of genetic, pharmacologic, and cell biological studies demonstrate active cytoplasmic and nucleolar retention mechanisms, whereas nuclear import and export occur through passive diffusion. Third, APOBEC3H cytoplasmic retention determinants relocalize APOBEC3A from a passive cell-wide state to the cytosol and, additionally, endow potent HIV-1 restriction activity. These results indicate that APOBEC3H has a structural zipcode for subcellular localization and selecting viral substrates for restriction.
Collapse
|
|
32
|
RNA-Mediated Dimerization of the Human Deoxycytidine Deaminase APOBEC3H Influences Enzyme Activity and Interaction with Nucleic Acids. J Mol Biol 2018; 430:4891-4907. [PMID: 30414963 DOI: 10.1016/j.jmb.2018.11.006] [Citation(s) in RCA: 14] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/04/2018] [Revised: 11/04/2018] [Accepted: 11/04/2018] [Indexed: 12/11/2022]
Abstract
Human APOBEC3H is a single-stranded (ss)DNA deoxycytidine deaminase that inhibits replication of retroelements and HIV-1 in CD4+ T cells. When aberrantly expressed in lung or breast tissue, APOBEC3H can contribute to cancer mutagenesis. These different activities are carried out by different haplotypes of APOBEC3H. Here we studied APOBEC3H haplotype II, which is able to restrict HIV-1 replication and retroelements. We determined how the dimerization mechanism, which is mediated by a double-stranded RNA molecule, influenced interactions with and activity on ssDNA. The data demonstrate that the cellular RNA bound by APOBEC3H does not completely inhibit enzyme activity, in contrast to other APOBEC family members. Despite degradation of the cellular RNA, an approximately 12-nt RNA remains bound to the enzyme, even in the presence of ssDNA. The RNA-mediated dimer is disrupted by mutating W115 on loop 7 or R175 and R176 on helix 6, but this also disrupts protein stability. In contrast, mutation of Y112 and Y113 on loop 7 also destabilizes RNA-mediated dimerization but results in a stable enzyme. Mutants unable to bind cellular RNA are unable to bind RNA oligonucleotides, oligomerize, and deaminate ssDNA in vitro, but ssDNA binding is retained. Comparison of A3H wild type and Y112A/Y113A by fluorescence polarization, single-molecule optical tweezer, and atomic force microscopy experiments demonstrates that RNA-mediated dimerization alters the interactions of A3H with ssDNA and other RNA molecules. Altogether, the biochemical analysis demonstrates that RNA binding is integral to APOBEC3H function.
Collapse
|
|
33
|
Ebrahimi D, Richards CM, Carpenter MA, Wang J, Ikeda T, Becker JT, Cheng AZ, McCann JL, Shaban NM, Salamango DJ, Starrett GJ, Lingappa JR, Yong J, Brown WL, Harris RS. Genetic and mechanistic basis for APOBEC3H alternative splicing, retrovirus restriction, and counteraction by HIV-1 protease. Nat Commun 2018; 9:4137. [PMID: 30297863 PMCID: PMC6175962 DOI: 10.1038/s41467-018-06594-3] [Citation(s) in RCA: 21] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/20/2018] [Accepted: 09/13/2018] [Indexed: 12/11/2022] Open
Abstract
Human APOBEC3H (A3H) is a single-stranded DNA cytosine deaminase that inhibits HIV-1. Seven haplotypes (I–VII) and four splice variants (SV154/182/183/200) with differing antiviral activities and geographic distributions have been described, but the genetic and mechanistic basis for variant expression and function remains unclear. Using a combined bioinformatic/experimental analysis, we find that SV200 expression is specific to haplotype II, which is primarily found in sub-Saharan Africa. The underlying genetic mechanism for differential mRNA splicing is an ancient intronic deletion [del(ctc)] within A3H haplotype II sequence. We show that SV200 is at least fourfold more HIV-1 restrictive than other A3H splice variants. To counteract this elevated antiviral activity, HIV-1 protease cleaves SV200 into a shorter, less restrictive isoform. Our analyses indicate that, in addition to Vif-mediated degradation, HIV-1 may use protease as a counter-defense mechanism against A3H in >80% of sub-Saharan African populations. Human APOBEC3H has several haplotypes and splice variants with distinct anti-HIV-1 activities, but the genetics underlying the expression of these variants are unclear. Here, the authors identify an intronic deletion in A3H haplotype II resulting in production of the most active splice variant, which is counteracted by HIV-1 protease.
Collapse
Affiliation(s)
- Diako Ebrahimi
- Department of Biochemistry, Molecular Biology and Biophysics, Masonic Cancer Center, Institute for Molecular Virology, Center for Genome Engineering, University of Minnesota, Minneapolis, MN, 55455, USA
| | - Christopher M Richards
- Department of Biochemistry, Molecular Biology and Biophysics, Masonic Cancer Center, Institute for Molecular Virology, Center for Genome Engineering, University of Minnesota, Minneapolis, MN, 55455, USA
| | - Michael A Carpenter
- Department of Biochemistry, Molecular Biology and Biophysics, Masonic Cancer Center, Institute for Molecular Virology, Center for Genome Engineering, University of Minnesota, Minneapolis, MN, 55455, USA.,Howard Hughes Medical Institute, University of Minnesota, Minneapolis, MN, 55455, USA
| | - Jiayi Wang
- Department of Biochemistry, Molecular Biology and Biophysics, Masonic Cancer Center, Institute for Molecular Virology, Center for Genome Engineering, University of Minnesota, Minneapolis, MN, 55455, USA
| | - Terumasa Ikeda
- Department of Biochemistry, Molecular Biology and Biophysics, Masonic Cancer Center, Institute for Molecular Virology, Center for Genome Engineering, University of Minnesota, Minneapolis, MN, 55455, USA.,Howard Hughes Medical Institute, University of Minnesota, Minneapolis, MN, 55455, USA
| | - Jordan T Becker
- Department of Biochemistry, Molecular Biology and Biophysics, Masonic Cancer Center, Institute for Molecular Virology, Center for Genome Engineering, University of Minnesota, Minneapolis, MN, 55455, USA
| | - Adam Z Cheng
- Department of Biochemistry, Molecular Biology and Biophysics, Masonic Cancer Center, Institute for Molecular Virology, Center for Genome Engineering, University of Minnesota, Minneapolis, MN, 55455, USA
| | - Jennifer L McCann
- Department of Biochemistry, Molecular Biology and Biophysics, Masonic Cancer Center, Institute for Molecular Virology, Center for Genome Engineering, University of Minnesota, Minneapolis, MN, 55455, USA
| | - Nadine M Shaban
- Department of Biochemistry, Molecular Biology and Biophysics, Masonic Cancer Center, Institute for Molecular Virology, Center for Genome Engineering, University of Minnesota, Minneapolis, MN, 55455, USA
| | - Daniel J Salamango
- Department of Biochemistry, Molecular Biology and Biophysics, Masonic Cancer Center, Institute for Molecular Virology, Center for Genome Engineering, University of Minnesota, Minneapolis, MN, 55455, USA
| | - Gabriel J Starrett
- Department of Biochemistry, Molecular Biology and Biophysics, Masonic Cancer Center, Institute for Molecular Virology, Center for Genome Engineering, University of Minnesota, Minneapolis, MN, 55455, USA.,Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD, 20892, USA
| | - Jairam R Lingappa
- Departments of Global Health, Medicine and Pediatrics, University of Washington, Seattle, WA, 98104, USA
| | - Jeongsik Yong
- Department of Biochemistry, Molecular Biology and Biophysics, Masonic Cancer Center, Institute for Molecular Virology, Center for Genome Engineering, University of Minnesota, Minneapolis, MN, 55455, USA
| | - William L Brown
- Department of Biochemistry, Molecular Biology and Biophysics, Masonic Cancer Center, Institute for Molecular Virology, Center for Genome Engineering, University of Minnesota, Minneapolis, MN, 55455, USA
| | - Reuben S Harris
- Department of Biochemistry, Molecular Biology and Biophysics, Masonic Cancer Center, Institute for Molecular Virology, Center for Genome Engineering, University of Minnesota, Minneapolis, MN, 55455, USA. .,Howard Hughes Medical Institute, University of Minnesota, Minneapolis, MN, 55455, USA.
| |
Collapse
|
|
34
|
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.
Collapse
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
| |
Collapse
|
|
35
|
Recurrent Loss of APOBEC3H Activity during Primate Evolution. J Virol 2018; 92:JVI.00971-18. [PMID: 29925657 DOI: 10.1128/jvi.00971-18] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/04/2018] [Accepted: 06/08/2018] [Indexed: 01/01/2023] Open
Abstract
Genes in the APOBEC3 family encode cytidine deaminases that provide a barrier against viral infection and retrotransposition. Of all the APOBEC3 genes in humans, APOBEC3H (A3H) is the most polymorphic: some genes encode stable and active A3H proteins, while others are unstable and poorly antiviral. Such variation in human A3H affects interactions with the lentiviral antagonist Vif, which counteracts A3H via proteasomal degradation. In order to broaden our understanding of A3H-Vif interactions, as well as its evolution in Old World monkeys, we characterized A3H variation within four African green monkey (AGM) subspecies. We found that A3H is highly polymorphic in AGMs and has lost antiviral activity in multiple Old World monkeys. This loss of function was partially related to protein expression levels but was also influenced by amino acid mutations in the N terminus. Moreover, we demonstrate that the evolution of A3H in the primate lineages leading to AGMs was not driven by Vif. Our work suggests that the activity of A3H is evolutionarily dynamic and may have a negative effect on host fitness, resulting in its recurrent loss in primates.IMPORTANCE Adaptation of viruses to their hosts is critical for viral transmission between different species. Previous studies had identified changes in a protein from the APOBEC3 family that influenced the species specificity of simian immunodeficiency viruses (SIVs) in African green monkeys. We studied the evolution of a related protein in the same system, APOBEC3H, which has experienced a loss of function in humans. This evolutionary approach revealed that recurrent loss of APOBEC3H activity has taken place during primate evolution, suggesting that APOBEC3H places a fitness cost on hosts. The variability of APOBEC3H activity between different primates highlights the differential selective pressures on the APOBEC3 gene family.
Collapse
|
|
36
|
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.
Collapse
|
|
37
|
Ito F, Yang H, Xiao X, Li SX, Wolfe A, Zirkle B, Arutiunian V, Chen XS. Understanding the Structure, Multimerization, Subcellular Localization and mC Selectivity of a Genomic Mutator and Anti-HIV Factor APOBEC3H. Sci Rep 2018; 8:3763. [PMID: 29491387 PMCID: PMC5830531 DOI: 10.1038/s41598-018-21955-0] [Citation(s) in RCA: 34] [Impact Index Per Article: 4.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/05/2017] [Accepted: 02/12/2018] [Indexed: 12/27/2022] Open
Abstract
APOBEC3H (A3H) is a member of the APOBEC3 subfamily of DNA cytosine deaminases that are important for innate immune defense and have been implicated in cancer biogenesis. To understand the structural basis for A3H biochemical function, we determined a high-resolution structure of human A3H and performed extensive biochemical analysis. The 2.49 Å crystal structure reveals a uniquely long C-terminal helix 6 (h6), a disrupted β5 strand of the canonical five-stranded β-sheet core, and a long loop 1 around the Zn-active center. Mutation of a loop 7 residue, W115, disrupted the RNA-mediated dimerization of A3H yielding an RNA-free monomeric form that still possessed nucleic acid binding and deaminase activity. A3H expressed in HEK293T cells showed RNA dependent HMW complex formation and RNase A-dependent deaminase activity. A3H has a highly positively charged surface surrounding the Zn-active center, and multiple positively charged residues within this charged surface play an important role in the RNA-mediated HMW formation and deaminase inhibition. Furthermore, these positively charged residues affect subcellular localization of A3H between the nucleus and cytosol. Finally, we have identified multiple residues of loop 1 and 7 that contribute to the overall deaminase activity and the methylcytosine selectivity.
Collapse
Affiliation(s)
- Fumiaki Ito
- Molecular and Computational Biology, Departments of Biological Sciences and Chemistry, University of Southern California, Los Angeles, CA, 90089, USA
| | - Hanjing Yang
- Molecular and Computational Biology, Departments of Biological Sciences and Chemistry, University of Southern California, Los Angeles, CA, 90089, USA
| | - Xiao Xiao
- Molecular and Computational Biology, Departments of Biological Sciences and Chemistry, University of Southern California, Los Angeles, CA, 90089, USA.,Genetic, Molecular and Cellular Biology Program, Keck School of Medicine, University of Southern California, Los Angeles, CA, 90089, USA.,Department of Infectious Diseases and Vaccines Research, Merck Research Laboratories, Merck & Co., Inc, West Point, PA, USA
| | - Shu-Xing Li
- Molecular and Computational Biology, 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
| | - Aaron Wolfe
- Molecular and Computational Biology, Departments of Biological Sciences and Chemistry, University of Southern California, Los Angeles, CA, 90089, USA.,Genetic, Molecular and Cellular Biology Program, Keck School of Medicine, University of Southern California, Los Angeles, CA, 90089, USA
| | - Brett Zirkle
- Molecular and Computational Biology, Departments of Biological Sciences and Chemistry, University of Southern California, Los Angeles, CA, 90089, USA.,Genetic, Molecular and Cellular Biology Program, Keck School of Medicine, University of Southern California, Los Angeles, CA, 90089, USA
| | - Vagan Arutiunian
- Molecular and Computational Biology, Departments of Biological Sciences and Chemistry, University of Southern California, Los Angeles, CA, 90089, USA.,Department of Internal Medicine, Meharry Medical College, Nashville, TN, USA
| | - Xiaojiang S Chen
- Molecular and Computational Biology, Departments of Biological Sciences and Chemistry, University of Southern California, Los Angeles, CA, 90089, USA. .,Genetic, Molecular and Cellular Biology Program, Keck School of Medicine, 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.
| |
Collapse
|
|
38
|
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.
Collapse
|
|
39
|
Benito JM, Hillung J, Restrepo C, Cuevas JM, León A, Ruiz-Mateos E, Palacios-Muñoz R, Górgolas M, Sanjuán R, Rallón N. Role of APOBEC3H in the Viral Control of HIV Elite Controller Patients. Int J Med Sci 2018; 15:95-100. [PMID: 29333092 PMCID: PMC5765721 DOI: 10.7150/ijms.22317] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 08/10/2017] [Accepted: 10/12/2017] [Indexed: 11/05/2022] Open
Abstract
Background APOBEC3H (A3H) gene presents variation at 2 positions (rs139297 and rs79323350) leading to a non-functional protein. So far, there is no information on the role played by A3H in spontaneous control of HIV. The aim of this study was to evaluate the A3H polymorphisms distribution in a well-characterized group of Elite Controller (EC) subjects. Methods We analyzed the genotype distribution of two different SNPs (rs139297 and rs79323350) of A3H in 30 EC patients and compared with 11 non-controller (NC) HIV patients. Genotyping was performed by PCR, cloning and Sanger sequencing. Both polymorphisms were analyzed jointly in order to adequately attribute the active or inactive status of A3H protein. Results EC subjects included in this study were able to maintain a long-term sustained spontaneous HIV-viral control and optimal CD4-T-cell counts; however, haplotypes leading to an active protein were very poorly represented in these patients. We found that the majority of EC subjects (23/30; 77%) presented allelic combinations leading to an inactive A3H protein, a frequency slightly lower than that observed for NC studied patients (10/11; 91%). Conclusions The high prevalence of non-functional protein coding-genotypes in EC subjects seems to indicate that other innate restriction factors different from APOBEC3H could be implicated in the replication control exhibited by these subjects.
Collapse
Affiliation(s)
- José M Benito
- Instituto de Investigación Sanitaria-Fundación Jiménez Díaz, Universidad Autónoma de Madrid (IIS-FJD, UAM), Spain.,Hospital Universitario Rey Juan Carlos, Móstoles, Spain
| | - Julia Hillung
- Institute for Integrative Systems Biology (I2SysBio), Universitat de València and Consejo Superior de Investigaciones Científicas, València, Spain
| | - Clara Restrepo
- Instituto de Investigación Sanitaria-Fundación Jiménez Díaz, Universidad Autónoma de Madrid (IIS-FJD, UAM), Spain.,Hospital Universitario Rey Juan Carlos, Móstoles, Spain
| | - José M Cuevas
- Institute for Integrative Systems Biology (I2SysBio), Universitat de València and Consejo Superior de Investigaciones Científicas, València, Spain.,Departament de Genètica, Universitat de València, València, Spain
| | - Agathe León
- Hospital Clínic of Barcelona, IDIBAPS, Barcelona, Spain
| | | | | | - Miguel Górgolas
- Hospital Universitario Fundación Jiménez Díaz, Madrid, Spain
| | - Rafael Sanjuán
- Institute for Integrative Systems Biology (I2SysBio), Universitat de València and Consejo Superior de Investigaciones Científicas, València, Spain.,Departament de Genètica, Universitat de València, València, Spain
| | - Norma Rallón
- Instituto de Investigación Sanitaria-Fundación Jiménez Díaz, Universidad Autónoma de Madrid (IIS-FJD, UAM), Spain.,Hospital Universitario Rey Juan Carlos, Móstoles, Spain
| |
Collapse
|
|
40
|
The Antiviral and Cancer Genomic DNA Deaminase APOBEC3H Is Regulated by an RNA-Mediated Dimerization Mechanism. Mol Cell 2017; 69:75-86.e9. [PMID: 29290613 DOI: 10.1016/j.molcel.2017.12.010] [Citation(s) in RCA: 58] [Impact Index Per Article: 7.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/15/2017] [Revised: 10/25/2017] [Accepted: 12/13/2017] [Indexed: 01/23/2023]
Abstract
Human APOBEC3H and homologous single-stranded DNA cytosine deaminases are unique to mammals. These DNA-editing enzymes function in innate immunity by restricting the replication of viruses and transposons. APOBEC3H also contributes to cancer mutagenesis. Here, we address the fundamental nature of RNA in regulating human APOBEC3H activities. APOBEC3H co-purifies with RNA as an inactive protein, and RNase A treatment enables strong DNA deaminase activity. RNA-binding-defective mutants demonstrate clear separation of function by becoming DNA hypermutators. Biochemical and crystallographic data demonstrate a mechanism in which double-stranded RNA mediates enzyme dimerization. Additionally, APOBEC3H separation-of-function mutants show that RNA binding is required for cytoplasmic localization, packaging into HIV-1 particles, and antiviral activity. Overall, these results support a model in which structured RNA negatively regulates the potentially harmful DNA deamination activity of APOBEC3H while, at the same time, positively regulating its antiviral activity.
Collapse
|
|
41
|
Zhang Z, Gu Q, de Manuel Montero M, Bravo IG, Marques-Bonet T, Häussinger D, Münk C. Stably expressed APOBEC3H forms a barrier for cross-species transmission of simian immunodeficiency virus of chimpanzee to humans. PLoS Pathog 2017; 13:e1006746. [PMID: 29267382 PMCID: PMC5739507 DOI: 10.1371/journal.ppat.1006746] [Citation(s) in RCA: 24] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/05/2017] [Accepted: 11/12/2017] [Indexed: 02/07/2023] Open
Abstract
APOBEC3s (A3s) are potent restriction factors of human immunodeficiency virus type 1/simian immunodeficiency viruses (HIV-1/SIV), and can repress cross-species transmissions of lentiviruses. HIV-1 originated from a zoonotic infection of SIV of chimpanzee (SIVcpz) to humans. However, the impact of human A3s on the replication of SIVcpz remains unclear. By using novel SIVcpz reporter viruses, we identified that human APOBEC3B (A3B) and APOBEC3H (A3H) haplotype II strongly reduced the infectivity of SIVcpz, because both of them are resistant to SIVcpz Vifs. We further demonstrated that human A3H inhibited SIVcpz by deaminase dependent as well independent mechanisms. In addition, other stably expressed human A3H haplotypes and splice variants showed strong antiviral activity against SIVcpz. Moreover, most SIV and HIV lineage Vif proteins could degrade chimpanzee A3H, but no Vifs from SIVcpz and SIV of gorilla (SIVgor) lineages antagonized human A3H haplotype II. Expression of human A3H hapII in human T cells efficiently blocked the spreading replication of SIVcpz. The spreading replication of SIVcpz was also restricted by stable A3H in human PBMCs. Thus, we speculate that stably expressed human A3H protects humans against the cross-species transmission of SIVcpz and that SIVcpz spillover to humans may have started in individuals that harbor haplotypes of unstable A3H proteins.
Collapse
Affiliation(s)
- Zeli Zhang
- Clinic for Gastroenterology, Hepatology, and Infectiology, Medical Faculty, Heinrich-Heine-Universität Düsseldorf, Düsseldorf, Germany
| | - Qinyong Gu
- Clinic for Gastroenterology, Hepatology, and Infectiology, Medical Faculty, Heinrich-Heine-Universität Düsseldorf, Düsseldorf, Germany
| | | | | | - Tomas Marques-Bonet
- Institut Biologia Evolutiva (Universitat Pompeu Fabra/CSIC) ICREA, Barcelona, Spain
| | - Dieter Häussinger
- Clinic for Gastroenterology, Hepatology, and Infectiology, Medical Faculty, Heinrich-Heine-Universität Düsseldorf, Düsseldorf, Germany
| | - Carsten Münk
- Clinic for Gastroenterology, Hepatology, and Infectiology, Medical Faculty, Heinrich-Heine-Universität Düsseldorf, Düsseldorf, Germany
| |
Collapse
|
|
42
|
Characterization of Ovine A3Z1 Restriction Properties against Small Ruminant Lentiviruses (SRLVs). Viruses 2017; 9:v9110345. [PMID: 29149056 PMCID: PMC5707552 DOI: 10.3390/v9110345] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/30/2017] [Revised: 11/14/2017] [Accepted: 11/15/2017] [Indexed: 12/22/2022] Open
Abstract
Intrinsic factors of the innate immune system include the apolipoprotein B editing enzyme catalytic polypeptide-like 3 (APOBEC3) protein family. APOBEC3 inhibits replication of different virus families by cytosine deamination of viral DNA and a not fully characterized cytosine deamination-independent mechanism. Sheep are susceptible to small ruminant lentivirus (SRLVs) infection and contain three APOBEC3 genes encoding four proteins (A3Z1, Z2, Z3 and Z2-Z3) with yet not deeply described antiviral properties. Using sheep blood monocytes and in vitro-derived macrophages, we found that A3Z1 expression is associated with lower viral replication in this cellular type. A3Z1 transcripts may also contain spliced variants (A3Z1Tr) lacking the cytidine deaminase motif. A3Z1 exogenous expression in fully permissive fibroblast-like cells restricted SRLVs infection while A3Z1Tr allowed infection. A3Z1Tr was induced after SRLVs infection or stimulation of blood-derived macrophages with interferon gamma (IFN-γ). Interaction between truncated isoform and native A3Z1 protein was detected as well as incorporation of both proteins into virions. A3Z1 and A3Z1Tr interacted with SRLVs Vif, but this interaction was not associated with degradative properties. Similar A3Z1 truncated isoforms were also present in human and monkey cells suggesting a conserved alternative splicing regulation in primates. A3Z1-mediated retroviral restriction could be constrained by different means, including gene expression and specific alternative splicing regulation, leading to truncated protein isoforms lacking a cytidine-deaminase motif.
Collapse
|
|
43
|
Bohn JA, Thummar K, York A, Raymond A, Brown WC, Bieniasz PD, Hatziioannou T, Smith JL. APOBEC3H structure reveals an unusual mechanism of interaction with duplex RNA. Nat Commun 2017; 8:1021. [PMID: 29044109 PMCID: PMC5647330 DOI: 10.1038/s41467-017-01309-6] [Citation(s) in RCA: 36] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/26/2017] [Accepted: 09/06/2017] [Indexed: 11/08/2022] Open
Abstract
The APOBEC3 family of cytidine deaminases cause lethal hypermutation of retroviruses via deamination of newly reverse-transcribed viral DNA. Their ability to bind RNA is essential for virion infiltration and antiviral activity, yet the mechanisms of viral RNA recognition are unknown. By screening naturally occurring, polymorphic, non-human primate APOBEC3H variants for biological and crystallization properties, we obtained a 2.24-Å crystal structure of pig-tailed macaque APOBEC3H with bound RNA. Here, we report that APOBEC3H forms a dimer around a short RNA duplex and, despite the bound RNA, has potent cytidine deaminase activity. The structure reveals an unusual RNA-binding mode in which two APOBEC3H molecules at opposite ends of a seven-base-pair duplex interact extensively with both RNA strands, but form no protein-protein contacts. CLIP-seq analysis revealed that APOBEC3H preferentially binds to sequences in the viral genome predicted to contain duplexes, a property that may facilitate both virion incorporation and catalytic activity.
Collapse
Affiliation(s)
- Jennifer A Bohn
- Life Sciences Institute, University of Michigan, Ann Arbor, MI, 48109, USA
- Department of Biological Chemistry, University of Michigan, Ann Arbor, MI, 48109, USA
| | - Keyur Thummar
- Laboratory of Retrovirology, The Rockefeller University, New York, NY, 10065, USA
- Howard Hughes Medical Institute, The Rockefeller University, New York, NY, 10065, USA
| | - Ashley York
- Laboratory of Retrovirology, The Rockefeller University, New York, NY, 10065, USA
- Howard Hughes Medical Institute, The Rockefeller University, New York, NY, 10065, USA
| | - Alice Raymond
- Laboratory of Retrovirology, The Rockefeller University, New York, NY, 10065, USA
- Howard Hughes Medical Institute, The Rockefeller University, New York, NY, 10065, USA
| | - W Clay Brown
- Life Sciences Institute, University of Michigan, Ann Arbor, MI, 48109, USA
| | - Paul D Bieniasz
- Laboratory of Retrovirology, The Rockefeller University, New York, NY, 10065, USA
- Howard Hughes Medical Institute, The Rockefeller University, New York, NY, 10065, USA
| | | | - Janet L Smith
- Life Sciences Institute, University of Michigan, Ann Arbor, MI, 48109, USA.
- Department of Biological Chemistry, University of Michigan, Ann Arbor, MI, 48109, USA.
| |
Collapse
|
|
44
|
Feng Y, Goubran MH, Follack TB, Chelico L. Deamination-independent restriction of LINE-1 retrotransposition by APOBEC3H. Sci Rep 2017; 7:10881. [PMID: 28883657 PMCID: PMC5589869 DOI: 10.1038/s41598-017-11344-4] [Citation(s) in RCA: 30] [Impact Index Per Article: 3.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/31/2017] [Accepted: 08/18/2017] [Indexed: 12/16/2022] Open
Abstract
The APOBEC3 family of cytosine deaminase enzymes are able to restrict replication of retroelements, such as LINE-1. However, each of the seven APOBEC3 enzymes have been reported to act differentially to prevent LINE-1 retrotransposition and the mechanisms of APOBEC3-mediated LINE-1 inhibition has not been well understood. The prevailing view for many years was that APOBEC3-mediated LINE-1 inhibition was deamination-independent and relied on APOBEC3s blocking the LINE-1 reverse transcriptase DNA polymerization or transport of the LINE-1 RNA into the nucleus. However, recently it was shown that APOBEC3A can deaminate cytosine, to form uracil, on transiently exposed single-stranded LINE-1 cDNA and this leads to LINE-1 cDNA degradation. In this study, we confirmed that APOBEC3A is a potent deamination-dependent inhibitor of LINE-1 retrotransposition, but show that in contrast, A3H haplotype II and haplotype V restrict LINE-1 activity using a deamination-independent mechanism. Our study supports the model that different APOBEC3 proteins have evolved to inhibit LINE-1 retrotransposition through distinct mechanisms.
Collapse
Affiliation(s)
- Yuqing Feng
- Department of Microbiology & Immunology, University of Saskatchewan, Saskatoon, Saskatchewan, S7N 5E5, Canada
| | - Mariam H Goubran
- Department of Microbiology & Immunology, University of Saskatchewan, Saskatoon, Saskatchewan, S7N 5E5, Canada
| | - Tyson B Follack
- Department of Microbiology & Immunology, University of Saskatchewan, Saskatoon, Saskatchewan, S7N 5E5, Canada
| | - Linda Chelico
- Department of Microbiology & Immunology, University of Saskatchewan, Saskatoon, Saskatchewan, S7N 5E5, Canada.
| |
Collapse
|
|
45
|
Rawson JMO, Gohl DM, Landman SR, Roth ME, Meissner ME, Peterson TS, Hodges JS, Beckman KB, Mansky LM. Single-Strand Consensus Sequencing Reveals that HIV Type but not Subtype Significantly Impacts Viral Mutation Frequencies and Spectra. J Mol Biol 2017; 429:2290-2307. [PMID: 28502791 DOI: 10.1016/j.jmb.2017.05.010] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/22/2016] [Revised: 05/07/2017] [Accepted: 05/08/2017] [Indexed: 10/19/2022]
Abstract
A long-standing question of human immunodeficiency virus (HIV) genetic variation and evolution has been whether differences exist in mutation rate and/or mutation spectra among HIV types (i.e., HIV-1 versus HIV-2) and among HIV groups (i.e., HIV-1 groups M-P and HIV-2 groups A-H) and HIV-1 Group M subtypes (i.e., subtypes A-D, F-H, and J-K). To address this, we developed a new single-strand consensus sequencing assay for the determination of HIV mutation frequencies and spectra using the Illumina sequencing platform. This assay enables parallel and standardized comparison of HIV mutagenesis among various viral vectors with lower background error than traditional methods of Illumina library preparation. We found significant differences in viral mutagenesis between HIV types but intriguingly no significant differences among HIV-1 Group M subtypes. More specifically, HIV-1 exhibited higher transition frequencies than HIV-2, due mostly to single G-to-A mutations and (to a lesser extent) G-to-A hypermutation. These data suggest that HIV-2 RT exhibits higher fidelity during viral replication, and taken together, these findings demonstrate that HIV type but not subtype significantly affects viral mutation frequencies and spectra. These differences may inform antiviral and vaccine strategies.
Collapse
Affiliation(s)
- Jonathan M O Rawson
- Molecular, Cellular, Developmental Biology & Genetics Graduate Program, University of Minnesota-Twin Cities, Minneapolis, MN, 55455, USA; Institute for Molecular Virology, University of Minnesota-Twin Cities, Minneapolis, MN, 55455, USA
| | - Daryl M Gohl
- University of Minnesota Genomics Center, University of Minnesota-Twin Cities, Minneapolis, MN, 55455, USA
| | - Sean R Landman
- Department of Computer Science and Engineering, University of Minnesota-Twin Cities, Minneapolis, MN, 55455, USA
| | - Megan E Roth
- Institute for Molecular Virology, University of Minnesota-Twin Cities, Minneapolis, MN, 55455, USA
| | - Morgan E Meissner
- Molecular, Cellular, Developmental Biology & Genetics Graduate Program, University of Minnesota-Twin Cities, Minneapolis, MN, 55455, USA; Institute for Molecular Virology, University of Minnesota-Twin Cities, Minneapolis, MN, 55455, USA
| | - Tara S Peterson
- Institute for Molecular Virology, University of Minnesota-Twin Cities, Minneapolis, MN, 55455, USA
| | - James S Hodges
- Division of Biostatistics, School of Public Health, University of Minnesota-Twin Cities, Minneapolis, MN, 55455, USA
| | - Kenneth B Beckman
- University of Minnesota Genomics Center, University of Minnesota-Twin Cities, Minneapolis, MN, 55455, USA
| | - Louis M Mansky
- Molecular, Cellular, Developmental Biology & Genetics Graduate Program, University of Minnesota-Twin Cities, Minneapolis, MN, 55455, USA; Institute for Molecular Virology, University of Minnesota-Twin Cities, Minneapolis, MN, 55455, USA; Division of Basic Sciences, School of Dentistry, University of Minnesota-Twin Cities, Minneapolis, MN, 55455, USA; Department of Microbiology & Immunology, University of Minnesota-Twin Cities, Minneapolis, MN, 55455, USA.
| |
Collapse
|
|
46
|
Ito F, Fu Y, Kao SCA, Yang H, Chen XS. Family-Wide Comparative Analysis of Cytidine and Methylcytidine Deamination by Eleven Human APOBEC Proteins. J Mol Biol 2017; 429:1787-1799. [PMID: 28479091 DOI: 10.1016/j.jmb.2017.04.021] [Citation(s) in RCA: 68] [Impact Index Per Article: 8.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/19/2017] [Revised: 04/28/2017] [Accepted: 04/29/2017] [Indexed: 01/17/2023]
Abstract
Apolipoprotein B mRNA editing enzyme, catalytic polypeptide-like (APOBEC) proteins are a family of cytidine deaminases involved in various important biological processes such as antibody diversification/maturation, restriction of viral infection, and generation of somatic mutations. Catalytically active APOBEC proteins execute their biological functions mostly through deaminating cytosine (C) to uracil on single-stranded DNA/RNA. Activation-induced cytidine deaminase, one of the APOBEC members, was reported to deaminate methylated cytosine (mC) on DNA, and this mC deamination was proposed to be involved in the demethylation of mC for epigenetic regulation. The mC deamination activity is later demonstrated for APOBEC3A (A3A) and more recently for APOBEC3B and APOBEC3H (A3H). Despite extensive studies on APOBEC proteins, questions regarding whether the rest of APOBEC members have any mC deaminase activity and what are the relative deaminase activities for each APOBEC member remain unclear. Here, we performed a family-wide analysis of deaminase activities on C and mC by using purified recombinant proteins for 11 known human APOBEC proteins under similar conditions. Our comprehensive analyses revealed that each APOBEC has unique deaminase activity and selectivity for mC. A3A and A3H showed distinctively high deaminase activities on C and mC with relatively high selectivity for mC, whereas six other APOBEC members showed relatively low deaminase activity and selectivity for mC. Our mutational analysis showed that loop-1 of A3A is responsible for its high deaminase activity and selectivity for mC. These findings extend our understanding of APOBEC family proteins that have important roles in diverse biological functions and in genetic mutations.
Collapse
Affiliation(s)
- Fumiaki Ito
- Molecular and Computational Biology Program, Department of Biological Sciences, University of Southern California, Los Angeles, CA 90089, USA
| | - Yang Fu
- Molecular and Computational Biology Program, Department of Biological Sciences, University of Southern California, Los Angeles, CA 90089, USA
| | - Shen-Chi A Kao
- Molecular and Computational Biology Program, Department of Biological Sciences, University of Southern California, Los Angeles, CA 90089, USA
| | - Hanjing Yang
- Molecular and Computational Biology Program, Department of Biological Sciences, University of Southern California, Los Angeles, CA 90089, USA
| | - Xiaojiang S Chen
- Molecular and Computational Biology Program, Department of Biological Sciences, University of Southern California, Los Angeles, CA 90089, USA; Department of Chemistry, Norris Comprehensive Cancer Center, University of Southern California, Los Angeles, CA 90089, USA; Center of Excellence in NanoBiophysics, Norris Comprehensive Cancer Center, University of Southern California, Los Angeles, CA 90089, USA.
| |
Collapse
|
|
47
|
Nakano Y, Misawa N, Juarez-Fernandez G, Moriwaki M, Nakaoka S, Funo T, Yamada E, Soper A, Yoshikawa R, Ebrahimi D, Tachiki Y, Iwami S, Harris RS, Koyanagi Y, Sato K. HIV-1 competition experiments in humanized mice show that APOBEC3H imposes selective pressure and promotes virus adaptation. PLoS Pathog 2017; 13:e1006348. [PMID: 28475648 PMCID: PMC5435363 DOI: 10.1371/journal.ppat.1006348] [Citation(s) in RCA: 29] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/05/2017] [Revised: 05/17/2017] [Accepted: 04/12/2017] [Indexed: 01/14/2023] Open
Abstract
APOBEC3 (A3) family proteins are DNA cytosine deaminases recognized for contributing to HIV-1 restriction and mutation. Prior studies have demonstrated that A3D, A3F, and A3G enzymes elicit a robust anti-HIV-1 effect in cell cultures and in humanized mouse models. Human A3H is polymorphic and can be categorized into three phenotypes: stable, intermediate, and unstable. However, the anti-viral effect of endogenous A3H in vivo has yet to be examined. Here we utilize a hematopoietic stem cell-transplanted humanized mouse model and demonstrate that stable A3H robustly affects HIV-1 fitness in vivo. In contrast, the selection pressure mediated by intermediate A3H is relaxed. Intriguingly, viral genomic RNA sequencing reveled that HIV-1 frequently adapts to better counteract stable A3H during replication in humanized mice. Molecular phylogenetic analyses and mathematical modeling suggest that stable A3H may be a critical factor in human-to-human viral transmission. Taken together, this study provides evidence that stable variants of A3H impose selective pressure on HIV-1.
Collapse
Affiliation(s)
- Yusuke Nakano
- Laboratory of Systems Virology, Department of Biosystems Science, Institute for Frontier Life and Medical Sciences, Kyoto University, Kyoto, Japan
| | - Naoko Misawa
- Laboratory of Systems Virology, Department of Biosystems Science, Institute for Frontier Life and Medical Sciences, Kyoto University, Kyoto, Japan
| | - Guillermo Juarez-Fernandez
- Laboratory of Systems Virology, Department of Biosystems Science, Institute for Frontier Life and Medical Sciences, Kyoto University, Kyoto, Japan
| | - Miyu Moriwaki
- Laboratory of Systems Virology, Department of Biosystems Science, Institute for Frontier Life and Medical Sciences, Kyoto University, Kyoto, Japan
- Graduate School of Biostudies, Kyoto University, Kyoto, Japan
| | - Shinji Nakaoka
- Institute of Industrial Sciences, The University of Tokyo, Meguro-ku, Tokyo, Japan
- PRESTO, Japan Science and Technology Agency, Kawaguchi, Saitama, Japan
| | - Takaaki Funo
- Mathematical Biology Laboratory, Department of Biology, Faculty of Sciences, Kyushu University, Fukuoka, Japan
| | - Eri Yamada
- Laboratory of Systems Virology, Department of Biosystems Science, Institute for Frontier Life and Medical Sciences, Kyoto University, Kyoto, Japan
| | - Andrew Soper
- Laboratory of Systems Virology, Department of Biosystems Science, Institute for Frontier Life and Medical Sciences, Kyoto University, Kyoto, Japan
| | - Rokusuke Yoshikawa
- Laboratory of Systems Virology, Department of Biosystems Science, Institute for Frontier Life and Medical Sciences, Kyoto University, Kyoto, Japan
| | - Diako Ebrahimi
- Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, Minnesota, United States of America
- Masonic Cancer Center, University of Minnesota, Minneapolis, Minnesota, United States of America
- Institute for Molecular Virology, University of Minnesota, Minneapolis, Minnesota, United States of America
- Center for Genome Engineering, University of Minnesota, Minneapolis, Minnesota, United States of America
| | - Yuuya Tachiki
- Laboratory of Systems Virology, Department of Biosystems Science, Institute for Frontier Life and Medical Sciences, Kyoto University, Kyoto, Japan
- Mathematical Biology Laboratory, Department of Biology, Faculty of Sciences, Kyushu University, Fukuoka, Japan
- CREST, Japan Science and Technology Agency, Kawaguchi, Saitama, Japan
| | - Shingo Iwami
- PRESTO, Japan Science and Technology Agency, Kawaguchi, Saitama, Japan
- Mathematical Biology Laboratory, Department of Biology, Faculty of Sciences, Kyushu University, Fukuoka, Japan
- CREST, Japan Science and Technology Agency, Kawaguchi, Saitama, Japan
| | - Reuben S. Harris
- Department of Biochemistry, Molecular Biology and Biophysics, University of Minnesota, Minneapolis, Minnesota, United States of America
- Masonic Cancer Center, University of Minnesota, Minneapolis, Minnesota, United States of America
- Institute for Molecular Virology, University of Minnesota, Minneapolis, Minnesota, United States of America
- Center for Genome Engineering, University of Minnesota, Minneapolis, Minnesota, United States of America
- Howard Hughes Medical Institute, University of Minnesota, Minneapolis, Minnesota, United States of America
| | - Yoshio Koyanagi
- Laboratory of Systems Virology, Department of Biosystems Science, Institute for Frontier Life and Medical Sciences, Kyoto University, Kyoto, Japan
| | - Kei Sato
- Laboratory of Systems Virology, Department of Biosystems Science, Institute for Frontier Life and Medical Sciences, Kyoto University, Kyoto, Japan
- CREST, Japan Science and Technology Agency, Kawaguchi, Saitama, Japan
| |
Collapse
|
|
48
|
Nakashima M, Tsuzuki S, Awazu H, Hamano A, Okada A, Ode H, Maejima M, Hachiya A, Yokomaku Y, Watanabe N, Akari H, Iwatani Y. Mapping Region of Human Restriction Factor APOBEC3H Critical for Interaction with HIV-1 Vif. J Mol Biol 2017; 429:1262-1276. [PMID: 28336404 DOI: 10.1016/j.jmb.2017.03.019] [Citation(s) in RCA: 16] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/22/2017] [Revised: 03/14/2017] [Accepted: 03/15/2017] [Indexed: 11/30/2022]
Abstract
The APOBEC3 (A3) family of cellular cytidine deaminases comprises seven members (A, B, C, D, F, G, and H) that potently inhibit retroviral replication. Human immunodeficiency virus type 1 (HIV-1) Vif is a small pleiotropic protein that specifically inactivates these enzymes, targeting them for ubiquitin-mediated proteasomal degradation. A3 Vif-interaction sites are presumed to fall into three distinct types: A3C/D/F, A3G, and A3H. To date, two types of A3G and A3C/D/F sites have been well characterized, whereas the A3H Vif-binding site remains poorly defined. Here, we explore the residues critical for the A3H-type Vif interaction. To avoid technical difficulties in performing experiments with human A3H haplotype II (hapII), which is relatively resistant to HIV-1 Vif, we employed its ortholog chimpanzee A3H (cA3H), which displays high Vif sensitivity, for a comparison of sensitivity with that of A3H hapII. The Vif susceptibility of A3H hapII-cA3H chimeras and their substitution mutants revealed a single residue at position 97 as a major determinant for the difference in their Vif sensitivities. We further surveyed critical residues by structure-guided mutagenesis using an A3H structural model and thus identified eight additional residues important for Vif sensitivity, which mapped to the α3 and α4 helices of A3H. Interestingly, this area is located on a surface adjacent to the A3G and A3C/D/F interfaces and is composed of negatively charged and hydrophobic patches. These findings suggest that HIV-1 Vif has evolved to utilize three dispersed surfaces for recognizing three types of interfaces on A3 proteins under certain structural constraints.
Collapse
Affiliation(s)
- Masaaki Nakashima
- Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi 460-0001, Japan
| | - Shinya Tsuzuki
- Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi 460-0001, Japan; Department of Biotechnology, Nagoya University Graduate School of Engineering, Nagoya, Aichi 464-8603, Japan
| | - Hiroaki Awazu
- Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi 460-0001, Japan; Department of Biotechnology, Nagoya University Graduate School of Engineering, Nagoya, Aichi 464-8603, Japan
| | - Akiko Hamano
- Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi 460-0001, Japan
| | - Ayaka Okada
- Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi 460-0001, Japan
| | - Hirotaka Ode
- Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi 460-0001, Japan
| | - Masami Maejima
- Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi 460-0001, Japan
| | - Atsuko Hachiya
- Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi 460-0001, Japan
| | - Yoshiyuki Yokomaku
- Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi 460-0001, Japan
| | - Nobuhisa Watanabe
- Department of Biotechnology, Nagoya University Graduate School of Engineering, Nagoya, Aichi 464-8603, Japan; Synchrotron Radiation Research Center, Nagoya University, Nagoya, Aichi, 489-0965, Japan
| | - Hirofumi Akari
- Primate Research Institute, Kyoto University, Inuyama, Aichi 484-8506, Japan; Institute for Virus Research, Kyoto University, Kyoto, Kyoto 606-8507, Japan
| | - Yasumasa Iwatani
- Clinical Research Center, National Hospital Organization Nagoya Medical Center, Nagoya, Aichi 460-0001, Japan; Division of Basic Medicine, Nagoya University Graduate School of Medicine, Nagoya, Aichi 466-8550, Japan.
| |
Collapse
|
|
49
|
The Structural Interface between HIV-1 Vif and Human APOBEC3H. J Virol 2017; 91:JVI.02289-16. [PMID: 28031368 DOI: 10.1128/jvi.02289-16] [Citation(s) in RCA: 19] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/26/2016] [Accepted: 12/21/2016] [Indexed: 11/20/2022] Open
Abstract
Human APOBEC3H (A3H) is a cytidine deaminase that inhibits HIV-1 replication. To evade this restriction, the HIV-1 Vif protein binds A3H and mediates its proteasomal degradation. To date, little information on the Vif-A3H interface has been available. To decipher how both proteins interact, we first mapped the Vif-binding site on A3H by functionally testing a large set of A3H mutants in single-cycle infectivity and replication assays. Our data show that the two A3H α-helixes α3 and α4 represent the Vif-binding site of A3H. We next used viral adaptation and a set of Vif mutants to identify novel, reciprocal Vif variants that rescued viral infectivity in the presence of two Vif-resistant A3H mutants. These A3H-Vif interaction points were used to generate the first A3H-Vif structure model, which revealed that the A3H helixes α3 and α4 interact with the Vif β-sheet (β2-β5). This model is in good agreement with previously reported Vif and A3H amino acids important for interaction. Based on the predicted A3H-Vif interface, we tested additional points of contact, which validated our model. Moreover, these experiments showed that the A3H and A3G binding sites on HIV-1 Vif are largely distinct, with both host proteins interacting with Vif β-strand 2. Taken together, this virus-host interface model explains previously reported data and will help to identify novel drug targets to combat HIV-1 infection.IMPORTANCE HIV-1 needs to overcome several intracellular restriction factors in order to replicate efficiently. The human APOBEC3 locus encodes seven proteins, of which A3D, A3F, A3G, and A3H restrict HIV-1. HIV encodes the Vif protein, which binds to the APOBEC3 proteins and leads to their proteasomal degradation. No HIV-1 Vif-APOBEC3 costructure exists to date despite extensive research. We and others previously generated HIV-1 Vif costructure models with A3G and A3F by mapping specific contact points between both proteins. Here, we applied a similar approach to HIV-1 Vif and A3H and successfully generated a Vif-A3H interaction model. Importantly, we find that the HIV-1 Vif-A3H interface is distinct from the Vif-A3G and Vif-A3F interfaces, with a small Vif region being important for recognition of both A3G and A3H. Our Vif-A3H structure model informs on how both proteins interact and could guide toward approaches to block the Vif-A3H interface to target HIV replication.
Collapse
|
|
50
|
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.
Collapse
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
| |
Collapse
|