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1
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Jackson-Jones KA, McKnight Á, Sloan RD. The innate immune factor RPRD2/REAF and its role in the Lv2 restriction of HIV. mBio 2023; 14:e0257221. [PMID: 37882563 PMCID: PMC10746242 DOI: 10.1128/mbio.02572-21] [Citation(s) in RCA: 4] [Impact Index Per Article: 2.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/27/2023] Open
Abstract
Intracellular innate immunity involves co-evolved antiviral restriction factors that specifically inhibit infecting viruses. Studying these restrictions has increased our understanding of viral replication, host-pathogen interactions, and pathogenesis, and represent potential targets for novel antiviral therapies. Lentiviral restriction 2 (Lv2) was identified as an unmapped early-phase restriction of HIV-2 and later shown to also restrict HIV-1 and simian immunodeficiency virus. The viral determinants of Lv2 susceptibility have been mapped to the envelope and capsid proteins in both HIV-1 and HIV-2, and also viral protein R (Vpr) in HIV-1, and appears dependent on cellular entry mechanism. A genome-wide screen identified several likely contributing host factors including members of the polymerase-associated factor 1 (PAF1) and human silencing hub (HUSH) complexes, and the newly characterized regulation of nuclear pre-mRNA domain containing 2 (RPRD2). Subsequently, RPRD2 (or RNA-associated early-stage antiviral factor) has been shown to be upregulated upon T cell activation, is highly expressed in myeloid cells, binds viral reverse transcripts, and potently restricts HIV-1 infection. RPRD2 is also bound by HIV-1 Vpr and targeted for degradation by the proteasome upon reverse transcription, suggesting RPRD2 impedes reverse transcription and Vpr targeting overcomes this block. RPRD2 is mainly localized to the nucleus and binds RNA, DNA, and DNA:RNA hybrids. More recently, RPRD2 has been shown to negatively regulate genome-wide transcription and interact with the HUSH and PAF1 complexes which repress HIV transcription and are implicated in maintenance of HIV latency. In this review, we examine Lv2 restriction and the antiviral role of RPRD2 and consider potential mechanism(s) of action.
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Affiliation(s)
- Kathryn A. Jackson-Jones
- Centre for Inflammation Research, Institute of Regeneration and Repair, The University of Edinburgh, Edinburgh, United Kingdom
- Division of Infectious Diseases & Department of Microbiology-Immunology, Northwestern University Feinberg School of Medicine, Chicago, Illinois, USA
| | - Áine McKnight
- Blizard Institute, Barts and The London School of Medicine and Dentistry, Queen Mary University of London, London, United Kingdom
| | - Richard D. Sloan
- Centre for Inflammation Research, Institute of Regeneration and Repair, The University of Edinburgh, Edinburgh, United Kingdom
- ZJU-UoE Institute, Zhejiang University, Haining, China
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2
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Yang B, Fang L, Gao Q, Xu C, Xu J, Chen ZX, Wang Y, Yang P. Species-specific KRAB-ZFPs function as repressors of retroviruses by targeting PBS regions. Proc Natl Acad Sci U S A 2022; 119:e2119415119. [PMID: 35259018 PMCID: PMC8931336 DOI: 10.1073/pnas.2119415119] [Citation(s) in RCA: 7] [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: 11/03/2021] [Accepted: 02/01/2022] [Indexed: 01/01/2023] Open
Abstract
Eukaryotic genomes harbor sequences derived from the chromosomal integration of ancient viruses, such as endogenous retroviruses (ERVs), which comprise 8% of the human genome. Like exogenous retroviruses, ERVs retain many common functional elements, including the corresponding DNA sequences of transfer RNA (tRNA) primer binding sites (PBSs), which are utilized for reverse transcription initiation by exogenous retroviruses. Here, through a medium-scale analysis of PBS loci positioned within ERVs, coupled with chromatin immunoprecipitation sequencing (ChIP-seq) of Kruppel-associated box zinc finger proteins (KRAB-ZFPs), we identified multiple ZFPs that specifically bind to different PBS loci. Among these, we focused on PBS-Lys, which is utilized by HIV-1, and identified its specific binding proteins to be mouse ZFP961 and human ZNF417/ZNF587. We found that these proteins not only repress ERV transcription but also inhibit retrovirus integration and transcription. Disruption of these ZFPs rendered cells more susceptible to HIV-1 infection. Thus, our research provides a methodology for identifying potential host factors that target retroviruses by ERVs.
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Affiliation(s)
- Bo Yang
- Shanghai Key Laboratory of Maternal and Fetal Medicine, Clinical and Translational Research Center of Shanghai First Maternity & Infant Hospital, Frontier Science Center for Stem Cell Research, School of Life Sciences and Technology, Tongji University, Shanghai 200092, China
| | - Lu Fang
- Translational Medical Center for Stem Cell Therapy, Institute for Regenerative Medicine of Shanghai East Hospital, Frontier Science Center for Stem Cell Research, School of Life Sciences and Technology, Tongji University, Shanghai 200092, China
| | - Qianqian Gao
- Shanghai Key Laboratory of Maternal and Fetal Medicine, Clinical and Translational Research Center of Shanghai First Maternity & Infant Hospital, Frontier Science Center for Stem Cell Research, School of Life Sciences and Technology, Tongji University, Shanghai 200092, China
| | - Ce Xu
- Shanghai Key Laboratory of Maternal and Fetal Medicine, Clinical and Translational Research Center of Shanghai First Maternity & Infant Hospital, Frontier Science Center for Stem Cell Research, School of Life Sciences and Technology, Tongji University, Shanghai 200092, China
| | - Junqin Xu
- Shanghai Key Laboratory of Maternal and Fetal Medicine, Clinical and Translational Research Center of Shanghai First Maternity & Infant Hospital, Frontier Science Center for Stem Cell Research, School of Life Sciences and Technology, Tongji University, Shanghai 200092, China
| | - Zhen-Xia Chen
- Hubei Hongshan Laboratory, College of Biomedicine and Health, Huazhong Agricultural University, Wuhan 430070, China
- Hubei Key Laboratory of Agricultural Bioinformatics, College of Life Science and Technology, Huazhong Agricultural University, Wuhan 430070, China
| | - Yixuan Wang
- Translational Medical Center for Stem Cell Therapy, Institute for Regenerative Medicine of Shanghai East Hospital, Frontier Science Center for Stem Cell Research, School of Life Sciences and Technology, Tongji University, Shanghai 200092, China
| | - Peng Yang
- Shanghai Key Laboratory of Maternal and Fetal Medicine, Clinical and Translational Research Center of Shanghai First Maternity & Infant Hospital, Frontier Science Center for Stem Cell Research, School of Life Sciences and Technology, Tongji University, Shanghai 200092, China
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Abstract
The development of therapies to eliminate the latent HIV-1 reservoir is hampered by our incomplete understanding of the biomolecular mechanism governing HIV-1 latency. To further complicate matters, recent single cell RNA-seq studies reported extensive heterogeneity between latently HIV-1-infected primary T cells, implying that latent HIV-1 infection can persist in greatly differing host cell environments. We here show that transcriptomic heterogeneity is also found between latently infected T cell lines, which allowed us to study the underlying mechanisms of intercell heterogeneity at high signal resolution. Latently infected T cells exhibited a de-differentiated phenotype, characterized by the loss of T cell-specific markers and gene regulation profiles reminiscent of hematopoietic stem cells (HSC). These changes had functional consequences. As reported for stem cells, latently HIV-1 infected T cells efficiently forced lentiviral superinfections into a latent state and favored glycolysis. As a result, metabolic reprogramming or cell re-differentiation destabilized latent infection. Guided by these findings, data-mining of single cell RNA-seq data of latently HIV-1 infected primary T cells from patients revealed the presence of similar dedifferentiation motifs. >20% of the highly detectable genes that were differentially regulated in latently infected cells were associated with hematopoietic lineage development (e.g. HUWE1, IRF4, PRDM1, BATF3, TOX, ID2, IKZF3, CDK6) or were hematopoietic markers (SRGN; hematopoietic proteoglycan core protein). The data add to evidence that the biomolecular phenotype of latently HIV-1 infected cells differs from normal T cells and strategies to address their differential phenotype need to be considered in the design of therapeutic cure interventions. IMPORTANCE HIV-1 persists in a latent reservoir in memory CD4 T cells for the lifetime of a patient. Understanding the biomolecular mechanisms used by the host cells to suppress viral expression will provide essential insights required to develop curative therapeutic interventions. Unfortunately, our current understanding of these control mechanisms is still limited. By studying gene expression profiles, we demonstrated that latently HIV-1-infected T cells have a de-differentiated T cell phenotype. Software-based data integration allowed for the identification of drug targets that would re-differentiate viral host cells and, in extension, destabilize latent HIV-1 infection events. The importance of the presented data lies within the clear demonstration that HIV-1 latency is a host cell phenomenon. As such, therapeutic strategies must first restore proper host cell functionality to accomplish efficient HIV-1 reactivation.
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Yuan P, Yan J, Wang S, Guo Y, Xi X, Han S, Yin J, Peng B, He X, Bodem J, Liu W. Trim28 acts as restriction factor of prototype foamy virus replication by modulating H3K9me3 marks and destabilizing the viral transactivator Tas. Retrovirology 2021; 18:38. [PMID: 34903241 PMCID: PMC8670036 DOI: 10.1186/s12977-021-00584-y] [Citation(s) in RCA: 14] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/21/2021] [Accepted: 11/26/2021] [Indexed: 12/17/2022] Open
Abstract
Background Prototype foamy virus (PFV) is nonpathogenic complex retroviruses that express a transcriptional transactivator Tas, which is essential for the activity of viral long terminal repeat (LTR) promoter and internal promoter (IP). Tripartite motif-containing protein 28 (Trim28) is well known as a scaffold protein normally enriched in gene promoter region to repress transcription. We sought to determine if whether Trim28 could be enriched in PFV promoter region to participate the establishment of PFV latency infection. Results In this study, we show that Trim28 restricts Tas-dependent transactivation activity of PFV promoter and negatively regulates PFV replication. Trim28 was found to be enriched in LTR instead of IP promoter regions of PFV genome and contribute to the maintenance of histone H3K9me3 marks on the LTR promoter. Furthermore, Trim28 interacts with Tas and colocalizes with Tas in the nucleus. Besides, we found that Trim28, an E3 ubiquitin ligase, binds directly to and promotes Tas for ubiquitination and degradation. And the RBCC domain of Trim28 is required for the ubiquitination and degradation of Tas. Conclusions Collectively, our findings not only identify a host factor Trim28 negatively inhibits PFV replication by acting as transcriptional restriction factor enriched in viral LTR promoter through modulating H3K9me3 mark here, but also reveal that Trim28 mediated ubiquitin proteasome degradation of Tas as a mechanism underlying Trim28 restricts Tas-dependent transcription activity of PFV promoter and PFV replication. These findings provide new insights into the process of PFV latency establishment. Graphical Abstract ![]()
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Affiliation(s)
- Peipei Yuan
- Department of Immunology, School of Basic Medical Sciences, Hubei University of Medicine, Shiyan, 442000, China.,Hubei Province Key Laboratory of Allergy and Immunology, School of Basic Medical Sciences, Wuhan University, No. 185, Donghu Road, Wuchang District, Wuhan, 430071, China.,Hubei Key Laboratory of Embryonic Stem Cell Research, Hubei University of Medicine, Shiyan, 442000, Hubei, China
| | - Jun Yan
- Hubei Province Key Laboratory of Allergy and Immunology, School of Basic Medical Sciences, Wuhan University, No. 185, Donghu Road, Wuchang District, Wuhan, 430071, China.,Hubei Provincial Key Laboratory of Developmentally Originated Disease, School of Basic Medical Sciences, Wuhan University, Wuhan, 430071, China
| | - Shuang Wang
- Hubei Province Key Laboratory of Allergy and Immunology, School of Basic Medical Sciences, Wuhan University, No. 185, Donghu Road, Wuchang District, Wuhan, 430071, China.,Hubei Provincial Key Laboratory of Developmentally Originated Disease, School of Basic Medical Sciences, Wuhan University, Wuhan, 430071, China
| | - Yang Guo
- Department of Immunology, School of Basic Medical Sciences, Hubei University of Medicine, Shiyan, 442000, China.,Hubei Key Laboratory of Embryonic Stem Cell Research, Hubei University of Medicine, Shiyan, 442000, Hubei, China
| | - Xueyan Xi
- Department of Immunology, School of Basic Medical Sciences, Hubei University of Medicine, Shiyan, 442000, China.,Hubei Key Laboratory of Embryonic Stem Cell Research, Hubei University of Medicine, Shiyan, 442000, Hubei, China
| | - Song Han
- Hubei Province Key Laboratory of Allergy and Immunology, School of Basic Medical Sciences, Wuhan University, No. 185, Donghu Road, Wuchang District, Wuhan, 430071, China.,Hubei Provincial Key Laboratory of Developmentally Originated Disease, School of Basic Medical Sciences, Wuhan University, Wuhan, 430071, China
| | - Jun Yin
- Hubei Province Key Laboratory of Allergy and Immunology, School of Basic Medical Sciences, Wuhan University, No. 185, Donghu Road, Wuchang District, Wuhan, 430071, China.,Hubei Provincial Key Laboratory of Developmentally Originated Disease, School of Basic Medical Sciences, Wuhan University, Wuhan, 430071, China
| | - Biwen Peng
- Hubei Province Key Laboratory of Allergy and Immunology, School of Basic Medical Sciences, Wuhan University, No. 185, Donghu Road, Wuchang District, Wuhan, 430071, China.,Hubei Provincial Key Laboratory of Developmentally Originated Disease, School of Basic Medical Sciences, Wuhan University, Wuhan, 430071, China
| | - Xiaohua He
- Hubei Province Key Laboratory of Allergy and Immunology, School of Basic Medical Sciences, Wuhan University, No. 185, Donghu Road, Wuchang District, Wuhan, 430071, China.,Hubei Provincial Key Laboratory of Developmentally Originated Disease, School of Basic Medical Sciences, Wuhan University, Wuhan, 430071, China
| | - Jochen Bodem
- Institut für Virologie und Immunbiologie, Julius-Maximilians-Universität Würzburg, 97078, Würzburg, Germany
| | - Wanhong Liu
- Hubei Province Key Laboratory of Allergy and Immunology, School of Basic Medical Sciences, Wuhan University, No. 185, Donghu Road, Wuchang District, Wuhan, 430071, China. .,Hubei Provincial Key Laboratory of Developmentally Originated Disease, School of Basic Medical Sciences, Wuhan University, Wuhan, 430071, China.
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5
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Host Gene Regulation by Transposable Elements: The New, the Old and the Ugly. Viruses 2020; 12:v12101089. [PMID: 32993145 PMCID: PMC7650545 DOI: 10.3390/v12101089] [Citation(s) in RCA: 15] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/17/2020] [Revised: 09/14/2020] [Accepted: 09/23/2020] [Indexed: 12/12/2022] Open
Abstract
The human genome has been under selective pressure to evolve in response to emerging pathogens and other environmental challenges. Genome evolution includes the acquisition of new genes or new isoforms of genes and changes to gene expression patterns. One source of genome innovation is from transposable elements (TEs), which carry their own promoters, enhancers and open reading frames and can act as ‘controlling elements’ for our own genes. TEs include LINE-1 elements, which can retrotranspose intracellularly and endogenous retroviruses (ERVs) that represent remnants of past retroviral germline infections. Although once pathogens, ERVs also represent an enticing source of incoming genetic material that the host can then repurpose. ERVs and other TEs have coevolved with host genes for millions of years, which has allowed them to become embedded within essential gene expression programmes. Intriguingly, these host genes are often subject to the same epigenetic control mechanisms that evolved to combat the TEs that now regulate them. Here, we illustrate the breadth of host gene regulation through TEs by focusing on examples of young (The New), ancient (The Old), and disease-causing (The Ugly) TE integrants.
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6
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Turelli P, Playfoot C, Grun D, Raclot C, Pontis J, Coudray A, Thorball C, Duc J, Pankevich EV, Deplancke B, Busskamp V, Trono D. Primate-restricted KRAB zinc finger proteins and target retrotransposons control gene expression in human neurons. SCIENCE ADVANCES 2020; 6:eaba3200. [PMID: 32923624 PMCID: PMC7455193 DOI: 10.1126/sciadv.aba3200] [Citation(s) in RCA: 54] [Impact Index Per Article: 10.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 11/22/2019] [Accepted: 07/16/2020] [Indexed: 05/11/2023]
Abstract
In the first days of embryogenesis, transposable element-embedded regulatory sequences (TEeRS) are silenced by Kruppel-associated box (KRAB) zinc finger proteins (KZFPs). Many TEeRS are subsequently co-opted in transcription networks, but how KZFPs influence this process is largely unknown. We identify ZNF417 and ZNF587 as primate-specific KZFPs repressing HERVK (human endogenous retrovirus K) and SVA (SINE-VNTR-Alu) integrants in human embryonic stem cells (ESCs). Expressed in specific regions of the human developing and adult brain, ZNF417/587 keep controlling TEeRS in ESC-derived neurons and brain organoids, secondarily influencing the differentiation and neurotransmission profile of neurons and preventing the induction of neurotoxic retroviral proteins and an interferon-like response. Thus, evolutionarily recent KZFPs and their TE targets partner up to influence human neuronal differentiation and physiology.
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Affiliation(s)
- Priscilla Turelli
- School of Life Sciences, Ecole Polytechnique Fédérale de Lausanne (EPFL), Lausanne, Switzerland
| | - Christopher Playfoot
- School of Life Sciences, Ecole Polytechnique Fédérale de Lausanne (EPFL), Lausanne, Switzerland
| | - Dephine Grun
- School of Life Sciences, Ecole Polytechnique Fédérale de Lausanne (EPFL), Lausanne, Switzerland
| | - Charlène Raclot
- School of Life Sciences, Ecole Polytechnique Fédérale de Lausanne (EPFL), Lausanne, Switzerland
| | - Julien Pontis
- School of Life Sciences, Ecole Polytechnique Fédérale de Lausanne (EPFL), Lausanne, Switzerland
| | - Alexandre Coudray
- School of Life Sciences, Ecole Polytechnique Fédérale de Lausanne (EPFL), Lausanne, Switzerland
| | - Christian Thorball
- School of Life Sciences, Ecole Polytechnique Fédérale de Lausanne (EPFL), Lausanne, Switzerland
| | - Julien Duc
- School of Life Sciences, Ecole Polytechnique Fédérale de Lausanne (EPFL), Lausanne, Switzerland
| | - Eugenia V. Pankevich
- School of Life Sciences, Ecole Polytechnique Fédérale de Lausanne (EPFL), Lausanne, Switzerland
- CeMM Research Center for Molecular Medicine of the Austrian Academy of Sciences, Vienna, Austria
| | - Bart Deplancke
- School of Life Sciences, Ecole Polytechnique Fédérale de Lausanne (EPFL), Lausanne, Switzerland
| | - Volker Busskamp
- Center for Regenerative Therapies, Technische Universität Dresden, Dresden, Germany
- Faculty of Medicine, Department of Ophthalmology, University of Bonn, Bonn, Germany
| | - Didier Trono
- School of Life Sciences, Ecole Polytechnique Fédérale de Lausanne (EPFL), Lausanne, Switzerland
- Corresponding author.
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7
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Wolf G, de Iaco A, Sun MA, Bruno M, Tinkham M, Hoang D, Mitra A, Ralls S, Trono D, Macfarlan TS. KRAB-zinc finger protein gene expansion in response to active retrotransposons in the murine lineage. eLife 2020; 9:56337. [PMID: 32479262 PMCID: PMC7289599 DOI: 10.7554/elife.56337] [Citation(s) in RCA: 80] [Impact Index Per Article: 16.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/24/2020] [Accepted: 05/31/2020] [Indexed: 11/13/2022] Open
Abstract
The Krüppel-associated box zinc finger protein (KRAB-ZFP) family diversified in mammals. The majority of human KRAB-ZFPs bind transposable elements (TEs), however, since most TEs are inactive in humans it is unclear whether KRAB-ZFPs emerged to suppress TEs. We demonstrate that many recently emerged murine KRAB-ZFPs also bind to TEs, including the active ETn, IAP, and L1 families. Using a CRISPR/Cas9-based engineering approach, we genetically deleted five large clusters of KRAB-ZFPs and demonstrate that target TEs are de-repressed, unleashing TE-encoded enhancers. Homozygous knockout mice lacking one of two KRAB-ZFP gene clusters on chromosome 2 and chromosome 4 were nonetheless viable. In pedigrees of chromosome 4 cluster KRAB-ZFP mutants, we identified numerous novel ETn insertions with a modest increase in mutants. Our data strongly support the current model that recent waves of retrotransposon activity drove the expansion of KRAB-ZFP genes in mice and that many KRAB-ZFPs play a redundant role restricting TE activity.
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Affiliation(s)
- Gernot Wolf
- The Eunice Kennedy Shriver National Institute of Child Health and Human Development, The National Institutes of Health, Bethesda, United States
| | - Alberto de Iaco
- School of Life Sciences, École Polytechnique Fédérale de Lausanne (EPFL), Lausanne, Switzerland
| | - Ming-An Sun
- The Eunice Kennedy Shriver National Institute of Child Health and Human Development, The National Institutes of Health, Bethesda, United States
| | - Melania Bruno
- The Eunice Kennedy Shriver National Institute of Child Health and Human Development, The National Institutes of Health, Bethesda, United States
| | - Matthew Tinkham
- The Eunice Kennedy Shriver National Institute of Child Health and Human Development, The National Institutes of Health, Bethesda, United States
| | - Don Hoang
- The Eunice Kennedy Shriver National Institute of Child Health and Human Development, The National Institutes of Health, Bethesda, United States
| | - Apratim Mitra
- The Eunice Kennedy Shriver National Institute of Child Health and Human Development, The National Institutes of Health, Bethesda, United States
| | - Sherry Ralls
- The Eunice Kennedy Shriver National Institute of Child Health and Human Development, The National Institutes of Health, Bethesda, United States
| | - Didier Trono
- School of Life Sciences, École Polytechnique Fédérale de Lausanne (EPFL), Lausanne, Switzerland
| | - Todd S Macfarlan
- The Eunice Kennedy Shriver National Institute of Child Health and Human Development, The National Institutes of Health, Bethesda, United States
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8
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TRIM5α self-assembly and compartmentalization of the HIV-1 viral capsid. Nat Commun 2020; 11:1307. [PMID: 32161265 PMCID: PMC7066149 DOI: 10.1038/s41467-020-15106-1] [Citation(s) in RCA: 54] [Impact Index Per Article: 10.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/30/2019] [Accepted: 02/19/2020] [Indexed: 12/20/2022] Open
Abstract
The tripartite-motif protein, TRIM5α, is an innate immune sensor that potently restricts retrovirus infection by binding to human immunodeficiency virus capsids. Higher-ordered oligomerization of this protein forms hexagonally patterned structures that wrap around the viral capsid, despite an anomalously low affinity for the capsid protein (CA). Several studies suggest TRIM5α oligomerizes into a lattice with a symmetry and spacing that matches the underlying capsid, to compensate for the weak affinity, yet little is known about how these lattices form. Using a combination of computational simulations and electron cryo-tomography imaging, we reveal the dynamical mechanisms by which these lattices self-assemble. Constrained diffusion allows the lattice to reorganize, whereas defects form on highly curved capsid surfaces to alleviate strain and lattice symmetry mismatches. Statistical analysis localizes the TRIM5α binding interface at or near the CypA binding loop of CA. These simulations elucidate the molecular-scale mechanisms of viral capsid cellular compartmentalization by TRIM5α. Tripartite-motif containing (TRIM) proteins modulate cellular responses to viral infection. Here the authors use molecular dynamics simulations to demonstrate that TRIM5α uses a two-dimensional lattice hopping mechanism to aggregate on the HIV capsid surface and initiate lattice growth.
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9
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Zaikos TD, Terry VH, Sebastian Kettinger NT, Lubow J, Painter MM, Virgilio MC, Neevel A, Taschuk F, Onafuwa-Nuga A, McNamara LA, Riddell J, Bixby D, Markowitz N, Collins KL. Hematopoietic Stem and Progenitor Cells Are a Distinct HIV Reservoir that Contributes to Persistent Viremia in Suppressed Patients. Cell Rep 2019; 25:3759-3773.e9. [PMID: 30590047 DOI: 10.1016/j.celrep.2018.11.104] [Citation(s) in RCA: 34] [Impact Index Per Article: 5.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/26/2018] [Revised: 09/12/2018] [Accepted: 11/29/2018] [Indexed: 01/03/2023] Open
Abstract
Long-lived reservoirs of persistent HIV are a major barrier to a cure. CD4+ hematopoietic stem and progenitor cells (HSPCs) have the capacity for lifelong survival, self-renewal, and the generation of daughter cells. Recent evidence shows that they are also susceptible to HIV infection in vitro and in vivo. Whether HSPCs harbor infectious virus or contribute to plasma virus (PV) is unknown. Here, we provide strong evidence that clusters of identical proviruses from HSPCs and their likely progeny often match residual PV. A higher proportion of these sequences match residual PV than proviral genomes from bone marrow and peripheral blood mononuclear cells that are observed only once. Furthermore, an analysis of near-full-length genomes isolated from HSPCs provides evidence that HSPCs harbor functional HIV proviral genomes that often match residual PV. These results support the conclusion that HIV-infected HSPCs form a distinct and functionally significant reservoir of persistent HIV in infected people.
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Affiliation(s)
- Thomas D Zaikos
- Department of Microbiology and Immunology, University of Michigan, Ann Arbor, MI, USA
| | - Valeri H Terry
- Department of Internal Medicine, University of Michigan, Ann Arbor, MI, USA
| | - Nadia T Sebastian Kettinger
- Program in Cellular and Molecular Biology, University of Michigan, Ann Arbor, MI, USA; Medical Scientist Training Program, University of Michigan, Ann Arbor, MI, USA
| | - Jay Lubow
- Department of Microbiology and Immunology, University of Michigan, Ann Arbor, MI, USA
| | - Mark M Painter
- Graduate Program in Immunology, University of Michigan, Ann Arbor, MI, USA
| | - Maria C Virgilio
- Program in Cellular and Molecular Biology, University of Michigan, Ann Arbor, MI, USA
| | - Andrew Neevel
- Department of Internal Medicine, University of Michigan, Ann Arbor, MI, USA
| | - Frances Taschuk
- Department of Internal Medicine, University of Michigan, Ann Arbor, MI, USA
| | | | - Lucy A McNamara
- Department of Microbiology and Immunology, University of Michigan, Ann Arbor, MI, USA
| | - James Riddell
- Division of Infectious Disease, Department of Internal Medicine, University of Michigan, Ann Arbor, MI, USA
| | - Dale Bixby
- Division of Oncology, Department of Internal Medicine, University of Michigan, Ann Arbor, MI, USA
| | - Norman Markowitz
- Division of Infectious Diseases, Henry Ford Hospital, Detroit, MI, USA
| | - Kathleen L Collins
- Department of Microbiology and Immunology, University of Michigan, Ann Arbor, MI, USA; Department of Internal Medicine, University of Michigan, Ann Arbor, MI, USA; Program in Cellular and Molecular Biology, University of Michigan, Ann Arbor, MI, USA; Medical Scientist Training Program, University of Michigan, Ann Arbor, MI, USA; Division of Infectious Disease, Department of Internal Medicine, University of Michigan, Ann Arbor, MI, USA; Graduate Program in Immunology, University of Michigan, Ann Arbor, MI, USA.
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10
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Wu X, Kwong AC, Rice CM. Antiviral resistance of stem cells. Curr Opin Immunol 2019; 56:50-59. [PMID: 30352329 PMCID: PMC6462420 DOI: 10.1016/j.coi.2018.10.004] [Citation(s) in RCA: 17] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/27/2018] [Revised: 09/27/2018] [Accepted: 10/02/2018] [Indexed: 01/07/2023]
Abstract
Stem cells are important for growth and regeneration given their ability to self-renew and differentiate into mature cells. Resistance to certain viral infections has been established as a phenotype of stem cells, a protection in line with their important physiological function. Antiviral resistance is critical to all cells, but it is differentially regulated between stem cells and differentiated cells. Stem cells utilize antiviral RNA interference, interferon-independent repression of endogenous retroviruses and intrinsic expression of antiviral interferon-stimulated genes. Differentiated cells often rely on the interferon-associated protein-based response to induce a local antiviral state. This review outlines the antiviral resistance mechanisms of stem cells and discusses some ideas as to why stem cells and differentiated cells may have evolved to utilize distinct mechanisms.
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Affiliation(s)
- Xianfang Wu
- Laboratory of Virology and Infectious Disease, The Rockefeller University, New York, NY 10065, United States
| | - Andrew C Kwong
- Laboratory of Virology and Infectious Disease, The Rockefeller University, New York, NY 10065, United States; Department of Biochemistry and Biomedical Sciences, McMaster University, Hamilton, ON, Canada
| | - Charles M Rice
- Laboratory of Virology and Infectious Disease, The Rockefeller University, New York, NY 10065, United States.
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11
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Treger RS, Pope SD, Kong Y, Tokuyama M, Taura M, Iwasaki A. The Lupus Susceptibility Locus Sgp3 Encodes the Suppressor of Endogenous Retrovirus Expression SNERV. Immunity 2019; 50:334-347.e9. [PMID: 30709743 DOI: 10.1016/j.immuni.2018.12.022] [Citation(s) in RCA: 56] [Impact Index Per Article: 9.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/02/2018] [Revised: 10/18/2018] [Accepted: 12/17/2018] [Indexed: 12/24/2022]
Abstract
Elevated endogenous retrovirus (ERV) transcription and anti-ERV antibody reactivity are implicated in lupus pathogenesis. Overproduction of non-ecotropic ERV (NEERV) envelope glycoprotein gp70 and resultant nephritis occur in lupus-prone mice, but whether NEERV mis-expression contributes to lupus etiology is unclear. Here we identified suppressor of NEERV (Snerv) 1 and 2, Krüppel-associated box zinc-finger proteins (KRAB-ZFPs) that repressed NEERV by binding the NEERV long terminal repeat to recruit the transcriptional regulator KAP1. Germline Snerv1/Snerv2 deletion increased activating chromatin modifications, transcription, and gp70 expression from NEERV loci. F1 crosses of lupus-prone New Zealand Black (NZB) and 129 mice to Snerv1/Snerv2-/- mice failed to restore NEERV repression, demonstrating that loss of SNERV underlies the lupus autoantigen gp70 overproduction that promotes nephritis in susceptible mice and that SNERV encodes for Sgp3 (in NZB mice) and Gv-1 loci (in 129 mice). Increased ERV expression in lupus patients inversely correlated with three putative ERV-suppressing KRAB-ZFPs, suggesting that loss of KRAB-ZFP-mediated ERV control may contribute to human lupus pathogenesis.
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Affiliation(s)
- Rebecca S Treger
- Department of Immunobiology, Yale University School of Medicine, New Haven, CT 06520, USA
| | - Scott D Pope
- Department of Immunobiology, Yale University School of Medicine, New Haven, CT 06520, USA
| | - Yong Kong
- Department of Immunobiology, Yale University School of Medicine, New Haven, CT 06520, USA; Department of Molecular Biophysics and Biochemistry, W.M. Keck Foundation Biotechnology Resource Laboratory, Yale University School of Medicine, New Haven, CT 06520, USA
| | - Maria Tokuyama
- Department of Immunobiology, Yale University School of Medicine, New Haven, CT 06520, USA
| | - Manabu Taura
- Department of Immunobiology, Yale University School of Medicine, New Haven, CT 06520, USA
| | - Akiko Iwasaki
- Department of Immunobiology, Yale University School of Medicine, New Haven, CT 06520, USA; Department of Molecular, Cellular and Developmental Biology, Yale University, New Haven, CT 06520, USA; Howard Hughes Medical Institute, Chevy Chase, MD 20815, USA.
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12
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Tie CH, Fernandes L, Conde L, Robbez-Masson L, Sumner RP, Peacock T, Rodriguez-Plata MT, Mickute G, Gifford R, Towers GJ, Herrero J, Rowe HM. KAP1 regulates endogenous retroviruses in adult human cells and contributes to innate immune control. EMBO Rep 2018; 19:e45000. [PMID: 30061100 PMCID: PMC6172469 DOI: 10.15252/embr.201745000] [Citation(s) in RCA: 80] [Impact Index Per Article: 11.4] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/11/2017] [Revised: 06/21/2018] [Accepted: 07/09/2018] [Indexed: 12/19/2022] Open
Abstract
Endogenous retroviruses (ERVs) have accumulated in vertebrate genomes and contribute to the complexity of gene regulation. KAP1 represses ERVs during development by its recruitment to their repetitive sequences through KRAB zinc-finger proteins (KZNFs), but little is known about the regulation of ERVs in adult tissues. We observed that KAP1 repression of HERVK14C was conserved in differentiated human cells and performed KAP1 knockout to obtain an overview of KAP1 function. Our results show that KAP1 represses ERVs (including HERV-T and HERV-S) and ZNF genes, both of which overlap with KAP1 binding sites and H3K9me3 in multiple cell types. Furthermore, this pathway is functionally conserved in adult human peripheral blood mononuclear cells. Cytosine methylation that acts on KAP1 regulated loci is necessary to prevent an interferon response, and KAP1-depletion leads to activation of some interferon-stimulated genes. Finally, loss of KAP1 leads to a decrease in H3K9me3 enrichment at ERVs and ZNF genes and an RNA-sensing response mediated through MAVS signaling. These data indicate that the KAP1-KZNF pathway contributes to genome stability and innate immune control in adult human cells.
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Affiliation(s)
- Christopher Hc Tie
- Division of Infection and Immunity, University College London, London, UK
| | - Liane Fernandes
- Division of Infection and Immunity, University College London, London, UK
| | - Lucia Conde
- Bill Lyons Informatics Centre, UCL Cancer Institute, London, UK
| | | | - Rebecca P Sumner
- Division of Infection and Immunity, University College London, London, UK
| | - Tom Peacock
- Division of Infection and Immunity, University College London, London, UK
| | | | - Greta Mickute
- Division of Infection and Immunity, University College London, London, UK
| | - Robert Gifford
- MRC-University of Glasgow Centre for Virus Research, Glasgow, UK
| | - Greg J Towers
- Division of Infection and Immunity, University College London, London, UK
| | - Javier Herrero
- Bill Lyons Informatics Centre, UCL Cancer Institute, London, UK
| | - Helen M Rowe
- Division of Infection and Immunity, University College London, London, UK
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13
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Krischuns T, Günl F, Henschel L, Binder M, Willemsen J, Schloer S, Rescher U, Gerlt V, Zimmer G, Nordhoff C, Ludwig S, Brunotte L. Phosphorylation of TRIM28 Enhances the Expression of IFN-β and Proinflammatory Cytokines During HPAIV Infection of Human Lung Epithelial Cells. Front Immunol 2018; 9:2229. [PMID: 30323812 PMCID: PMC6172303 DOI: 10.3389/fimmu.2018.02229] [Citation(s) in RCA: 64] [Impact Index Per Article: 9.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/31/2018] [Accepted: 09/07/2018] [Indexed: 01/28/2023] Open
Abstract
Human infection with highly pathogenic avian influenza viruses (HPAIV) is often associated with severe tissue damage due to hyperinduction of interferons and proinflammatory cytokines. The reasons for this excessive cytokine expression are still incompletely understood, which has hampered the development of efficient immunomodulatory treatment options. The host protein TRIM28 associates to the promoter regions of over 13,000 genes and is recognized as a genomic corepressor and negative immune regulator. TRIM28 corepressor activity is regulated by post-translational modifications, specifically phosphorylation of S473, which modulates binding of TRIM28 to the heterochromatin-binding protein HP1. Here, we identified TRIM28 as a key immune regulator leading to increased IFN-β and proinflammatory cytokine levels during infection with HPAIV. Using influenza A virus strains of the subtype H1N1 as well as HPAIV of subtypes H7N7, H7N9, and H5N1, we could demonstrate that strain-specific phosphorylation of TRIM28 S473 is induced by a signaling cascade constituted of PKR, p38 MAPK, and MSK1 in response to RIG-I independent sensing of viral RNA. Furthermore, using chemical inhibitors as well as knockout cell lines, our results suggest that phosphorylation of S473 facilitates a functional switch leading to increased levels of IFN-β, IL-6, and IL-8. In summary, we have identified TRIM28 as a critical factor controlling excessive expression of type I IFNs as well as proinflammatory cytokines during infection with H5N1, H7N7, and H7N9 HPAIV. In addition, our data indicate a novel mechanism of PKR-mediated IFN-β expression, which could lay the ground for novel treatment options aiming at rebalancing dysregulated immune responses during severe HPAIV infection.
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Affiliation(s)
- Tim Krischuns
- Institute of Virology Muenster, Westfaelische Wilhelms-University Muenster, Muenster, Germany
- Cluster of Excellence “Cells in Motion”, Westfaelische Wilhelms-University Muenster, Muenster, Germany
| | - Franziska Günl
- Institute of Virology Muenster, Westfaelische Wilhelms-University Muenster, Muenster, Germany
- Cluster of Excellence “Cells in Motion”, Westfaelische Wilhelms-University Muenster, Muenster, Germany
| | - Lea Henschel
- Institute of Virology Muenster, Westfaelische Wilhelms-University Muenster, Muenster, Germany
- Cluster of Excellence “Cells in Motion”, Westfaelische Wilhelms-University Muenster, Muenster, Germany
| | - Marco Binder
- Research Group “Dynamics of Early Viral Infection and the Innate Antiviral Response”, Division Virus-Associated Carcinogenesis (F170), German Cancer Research Center (DKFZ), Heidelberg, Germany
| | - Joschka Willemsen
- Research Group “Dynamics of Early Viral Infection and the Innate Antiviral Response”, Division Virus-Associated Carcinogenesis (F170), German Cancer Research Center (DKFZ), Heidelberg, Germany
| | - Sebastian Schloer
- Cluster of Excellence “Cells in Motion”, Westfaelische Wilhelms-University Muenster, Muenster, Germany
- Center for Molecular Biology of Inflammation, Institute of Medical Biochemistry, Westfaelische Wilhelms-University Muenster, Muenster, Germany
| | - Ursula Rescher
- Cluster of Excellence “Cells in Motion”, Westfaelische Wilhelms-University Muenster, Muenster, Germany
- Center for Molecular Biology of Inflammation, Institute of Medical Biochemistry, Westfaelische Wilhelms-University Muenster, Muenster, Germany
| | - Vanessa Gerlt
- Institute of Virology Muenster, Westfaelische Wilhelms-University Muenster, Muenster, Germany
- Cluster of Excellence “Cells in Motion”, Westfaelische Wilhelms-University Muenster, Muenster, Germany
| | - Gert Zimmer
- Institute of Virology and Immunology (IVI), Bern, Switzerland
- Department of Infectious Diseases and Pathobiology (DIP), Vetsuisse Faculty, University of Bern, Bern, Switzerland
| | - Carolin Nordhoff
- Institute of Virology Muenster, Westfaelische Wilhelms-University Muenster, Muenster, Germany
- Cluster of Excellence “Cells in Motion”, Westfaelische Wilhelms-University Muenster, Muenster, Germany
| | - Stephan Ludwig
- Institute of Virology Muenster, Westfaelische Wilhelms-University Muenster, Muenster, Germany
- Cluster of Excellence “Cells in Motion”, Westfaelische Wilhelms-University Muenster, Muenster, Germany
| | - Linda Brunotte
- Institute of Virology Muenster, Westfaelische Wilhelms-University Muenster, Muenster, Germany
- Cluster of Excellence “Cells in Motion”, Westfaelische Wilhelms-University Muenster, Muenster, Germany
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14
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Yaparla A, Popovic M, Grayfer L. Differentiation-dependent antiviral capacities of amphibian ( Xenopus laevis) macrophages. J Biol Chem 2017; 293:1736-1744. [PMID: 29259133 DOI: 10.1074/jbc.m117.794065] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/30/2017] [Revised: 12/12/2017] [Indexed: 12/30/2022] Open
Abstract
Infections by ranaviruses such as Frog virus 3 (Fv3), are significantly contributing to worldwide amphibian population declines. Notably, amphibian macrophages (Mφs) are important to both the Fv3 infection strategies and the immune defense against this pathogen. However, the mechanisms underlying amphibian Mφ Fv3 susceptibility and resistance remain unknown. Mφ differentiation is mediated by signaling through the colony-stimulating factor-1 receptor (CSF-1R) which is now known to be bound not only by CSF-1, but also by the unrelated interleukin-34 (IL-34) cytokine. Pertinently, amphibian (Xenopus laevis) Mφs differentiated by CSF-1 and IL-34 are highly susceptible and resistant to Fv3, respectively. Accordingly, in the present work, we elucidate the facets of this Mφ Fv3 susceptibility and resistance. Because cellular resistance to viral replication is marked by expression of antiviral restriction factors, it was intuitive to find that IL-34-Mφs possess significantly greater mRNA levels of select restriction factor genes than CSF-1-Mφs. Xenopodinae amphibians have highly expanded repertoires of antiviral interferon (IFN) cytokine gene families, and our results indicated that in comparison with the X. laevis CSF-1-Mφs, the IL-34-Mφs express substantially greater transcripts of representative IFN genes, belonging to distinct gene family clades, as well as their cognate receptor genes. Finally, we demonstrate that IL-34-Mφ-conditioned supernatants confer IFN-mediated anti-Fv3 protection to the virally susceptible X. laevis kidney (A6) cell line. Together, this work underlines the differentiation pathways leading to Fv3-susceptible and -resistant amphibian Mφ populations and defines the molecular mechanisms responsible for these differences.
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Affiliation(s)
- Amulya Yaparla
- From the Department of Biological Sciences, George Washington University, Washington, D. C. 20052-0066
| | - Milan Popovic
- From the Department of Biological Sciences, George Washington University, Washington, D. C. 20052-0066
| | - Leon Grayfer
- From the Department of Biological Sciences, George Washington University, Washington, D. C. 20052-0066
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15
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LTR-Retrotransposon Control by tRNA-Derived Small RNAs. Cell 2017; 170:61-71.e11. [PMID: 28666125 DOI: 10.1016/j.cell.2017.06.013] [Citation(s) in RCA: 286] [Impact Index Per Article: 35.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/04/2016] [Revised: 04/13/2017] [Accepted: 05/09/2017] [Indexed: 12/31/2022]
Abstract
Transposon reactivation is an inherent danger in cells that lose epigenetic silencing during developmental reprogramming. In the mouse, long terminal repeat (LTR)-retrotransposons, or endogenous retroviruses (ERV), account for most novel insertions and are expressed in the absence of histone H3 lysine 9 trimethylation in preimplantation stem cells. We found abundant 18 nt tRNA-derived small RNA (tRF) in these cells and ubiquitously expressed 22 nt tRFs that include the 3' terminal CCA of mature tRNAs and target the tRNA primer binding site (PBS) essential for ERV reverse transcription. We show that the two most active ERV families, IAP and MusD/ETn, are major targets and are strongly inhibited by tRFs in retrotransposition assays. 22 nt tRFs post-transcriptionally silence coding-competent ERVs, while 18 nt tRFs specifically interfere with reverse transcription and retrotransposon mobility. The PBS offers a unique target to specifically inhibit LTR-retrotransposons, and tRF-targeting is a potentially highly conserved mechanism of small RNA-mediated transposon control.
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16
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Evsikov AV, Marín de Evsikova C. Friend or Foe: Epigenetic Regulation of Retrotransposons in Mammalian Oogenesis and Early Development. THE YALE JOURNAL OF BIOLOGY AND MEDICINE 2016; 89:487-497. [PMID: 28018140 PMCID: PMC5168827] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Indexed: 11/25/2022]
Abstract
Epigenetics is the study of phenotypic variation arising from developmental and environmental factors regulating gene transcription at molecular, cellular, and physiological levels. A naturally occurring biological process driven by epigenetics is the egg-to-embryo developmental transition when two fully differentiated adult cells - egg and sperm - revert to an early stem cell type with totipotency but subsequently differentiates into pluripotent embryonic stem cells that give rise to any cell type. Transposable elements (TEs) are active in mammalian oocytes and early embryos, and this activity, albeit counterintuitive because TEs can lead to genomic instability in somatic cells, correlates to successful development. TEs bridge genetic and epigenetic landscapes because TEs are genetic elements whose silencing and de-repression are regulated by epigenetic mechanisms that are sensitive to environmental factors. Ultimately, transposition events can change size, content, and function of mammalian genomes. Thus, TEs act beyond mutagenic agents reshuffling the genomes, and epigenetic regulation of TEs may act as a proximate mechanism by which evolutionary forces increase a species' hidden reserve of epigenetic and phenotypic variability facilitating the adaptation of genomes to their environment.
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Affiliation(s)
- Alexei V. Evsikov
- To whom all correspondence should be addressed: Caralina Marín de Evsikova, Alexei V. Evsikov, Department of Molecular Medicine, Morsani College of Medicine, University of South Florida, 12901 Bruce B Downs Blvd., MDC07, Tampa, FL 33612, CMdE: ; (813) 974 2248; AVE: ; (813) 974 6922, Fax: 813-974-7357
| | - Caralina Marín de Evsikova
- To whom all correspondence should be addressed: Caralina Marín de Evsikova, Alexei V. Evsikov, Department of Molecular Medicine, Morsani College of Medicine, University of South Florida, 12901 Bruce B Downs Blvd., MDC07, Tampa, FL 33612, CMdE: ; (813) 974 2248; AVE: ; (813) 974 6922, Fax: 813-974-7357
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17
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Transcriptional Silencing of Moloney Murine Leukemia Virus in Human Embryonic Carcinoma Cells. J Virol 2016; 91:JVI.02075-16. [PMID: 27795446 DOI: 10.1128/jvi.02075-16] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/21/2016] [Accepted: 10/21/2016] [Indexed: 11/20/2022] Open
Abstract
Embryonic carcinoma (EC) cells are malignant counterparts of embryonic stem (ES) cells and serve as useful models for investigating cellular differentiation and human embryogenesis. Though the susceptibility of murine EC cells to retroviral infection has been extensively analyzed, few studies of retrovirus infection of human EC cells have been performed. We tested the susceptibility of human EC cells to transduction by retroviral vectors derived from three different retroviral genera. We show that human EC cells efficiently express reporter genes delivered by vectors based on human immunodeficiency virus type 1 (HIV-1) and Mason-Pfizer monkey virus (M-PMV) but not Moloney murine leukemia virus (MLV). In human EC cells, MLV integration occurs normally, but no viral gene expression is observed. The block to MLV expression of MLV genomes is relieved upon cellular differentiation. The lack of gene expression is correlated with transcriptional silencing of the MLV promoter through the deposition of repressive histone marks as well as DNA methylation. Moreover, depletion of SETDB1, a histone methyltransferase, resulted in a loss of transcriptional silencing and upregulation of MLV gene expression. Finally, we provide evidence showing that the lack of MLV gene expression may be attributed in part to the lack of MLV enhancer function in human EC cells. IMPORTANCE Human embryonic carcinoma (EC) cells are shown to restrict the expression of murine leukemia virus genomes but not retroviral genomes of the lentiviral or betaretroviral families. The block occurs at the level of transcription and is accompanied by the deposition of repressive histone marks and methylation of the integrated proviral DNA. The host machinery required for silencing in human EC cells is distinct from that in murine EC cell lines: the histone methyltransferase SETDB1 is required, but the widely utilized corepressor TRIM28/Kap1 is not. A transcriptional enhancer element from the Mason-Pfizer monkey virus can override the silencing and promote transcription of chimeric proviral DNAs. The findings reveal novel features of human EC gene regulation not present in their murine counterparts.
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18
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Ecco G, Cassano M, Kauzlaric A, Duc J, Coluccio A, Offner S, Imbeault M, Rowe HM, Turelli P, Trono D. Transposable Elements and Their KRAB-ZFP Controllers Regulate Gene Expression in Adult Tissues. Dev Cell 2016; 36:611-23. [PMID: 27003935 DOI: 10.1016/j.devcel.2016.02.024] [Citation(s) in RCA: 168] [Impact Index Per Article: 18.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/08/2015] [Revised: 02/10/2016] [Accepted: 02/24/2016] [Indexed: 12/21/2022]
Abstract
KRAB-containing zinc finger proteins (KRAB-ZFPs) are early embryonic controllers of transposable elements (TEs), which they repress with their cofactor KAP1 through histone and DNA methylation, a process thought to result in irreversible silencing. Using a target-centered functional screen, we matched murine TEs with their cognate KRAB-ZFP. We found the paralogs ZFP932 and Gm15446 to bind overlapping but distinguishable subsets of ERVK (endogenous retrovirus K), repress these elements in embryonic stem cells, and regulate secondarily the expression of neighboring genes. Most importantly, we uncovered that these KRAB-ZFPs and KAP1 control TEs in adult tissues, in cell culture and in vivo, where they partner up to modulate cellular genes. Therefore, TEs and KRAB-ZFPs establish transcriptional networks that likely regulate not only development but also many physiological events. Given the high degree of species specificity of TEs and KRAB-ZFPs, these results have important implications for understanding the biology of higher vertebrates, including humans.
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Affiliation(s)
- Gabriela Ecco
- School of Life Sciences, École Polytechnique Fédérale de Lausanne (EPFL), Station 19, 1015 Lausanne, Switzerland
| | - Marco Cassano
- School of Life Sciences, École Polytechnique Fédérale de Lausanne (EPFL), Station 19, 1015 Lausanne, Switzerland
| | - Annamaria Kauzlaric
- School of Life Sciences, École Polytechnique Fédérale de Lausanne (EPFL), Station 19, 1015 Lausanne, Switzerland
| | - Julien Duc
- School of Life Sciences, École Polytechnique Fédérale de Lausanne (EPFL), Station 19, 1015 Lausanne, Switzerland
| | - Andrea Coluccio
- School of Life Sciences, École Polytechnique Fédérale de Lausanne (EPFL), Station 19, 1015 Lausanne, Switzerland
| | - Sandra Offner
- School of Life Sciences, École Polytechnique Fédérale de Lausanne (EPFL), Station 19, 1015 Lausanne, Switzerland
| | - Michaël Imbeault
- School of Life Sciences, École Polytechnique Fédérale de Lausanne (EPFL), Station 19, 1015 Lausanne, Switzerland
| | - Helen M Rowe
- School of Life Sciences, École Polytechnique Fédérale de Lausanne (EPFL), Station 19, 1015 Lausanne, Switzerland
| | - Priscilla Turelli
- School of Life Sciences, École Polytechnique Fédérale de Lausanne (EPFL), Station 19, 1015 Lausanne, Switzerland
| | - Didier Trono
- School of Life Sciences, École Polytechnique Fédérale de Lausanne (EPFL), Station 19, 1015 Lausanne, Switzerland.
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19
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Griffin DO, Goff SP. Restriction of HIV-1-based lentiviral vectors in adult primary marrow-derived and peripheral mobilized human CD34+ hematopoietic stem and progenitor cells occurs prior to viral DNA integration. Retrovirology 2016; 13:14. [PMID: 26945863 PMCID: PMC4779582 DOI: 10.1186/s12977-016-0246-0] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/16/2015] [Accepted: 02/18/2016] [Indexed: 01/10/2023] Open
Abstract
Background Gene therapy is currently being attempted using a number of delivery vehicles including lentiviral-based vectors. The delivery and insertion of a gene using lentiviral-based vectors involves multiple discrete steps, including reverse transcription of viral RNA into DNA, nuclear entry, integration of viral DNA into the host genome and expression of integrated genes. Transduction of murine stem cells by the murine leukemia viruses is inefficient because the expression of the integrated DNA is profoundly blocked. Transduction of human stem cells by lentivirus vectors is also inefficient, but the cause and specific part of the retroviral lifecycle where this block occurs is unknown. Results Here we demonstrate that the dominant point of restriction of an HIV-1-based lentiviral vector in adult human hematopoietic stem and progenitor cells (HSPCs) from bone marrow and also those obtained following peripheral mobilization is prior to viral DNA integration. We specifically show that restriction of HSPCs to an HIV-1-based lentiviral vector is prior to formation of nuclear DNA forms. Conclusions Murine restriction of MLV and human cellular restriction of HIV-1 are fundamentally different. While murine restriction of MLV occurs post integration, human restriction of HIV-1 occurs before integration.
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Affiliation(s)
- Daniel O Griffin
- Department of Biochemistry and Molecular Biophysics, Columbia University Medical Center, HHSC 1310c, 701 West 168th Street, New York, NY, 10032, USA. .,Division of Infectious Diseases, Department of Medicine, Columbia University Medical Center, New York, NY, 10032, USA.
| | - Stephen P Goff
- Department of Biochemistry and Molecular Biophysics, Columbia University Medical Center, HHSC 1310c, 701 West 168th Street, New York, NY, 10032, USA. .,Howard Hughes Medical Institute, Columbia University Medical Center, New York, NY, 10032, USA. .,Department of Microbiology and Immunology, Columbia University Medical Center, New York, NY, 10032, USA.
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20
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Christensen T. Human endogenous retroviruses in neurologic disease. APMIS 2016; 124:116-26. [DOI: 10.1111/apm.12486] [Citation(s) in RCA: 39] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/26/2015] [Accepted: 10/26/2015] [Indexed: 12/13/2022]
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21
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HIV-1 Is Restricted prior to Integration of Viral DNA in Primary Cord-Derived Human CD34+ Cells. J Virol 2015; 89:8096-100. [PMID: 25995256 DOI: 10.1128/jvi.01044-15] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/18/2023] Open
Abstract
Certain cells have the ability to block retroviral infection at specific stages of the viral cycle by the activities of well-characterized factors and transcriptional silencing machinery. Infection of murine stem cells (MSCs) by the murine leukemia viruses (MLVs) is profoundly blocked postintegration by transcriptional silencing. Here, we show that a dominant point of restriction of HIV-1 in human CD34+ cells is prior to integration of viral DNA and that HIV-1 restriction by human CD34+ cells is fundamentally different from MLV restriction by mouse cells.
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22
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Yang BX, El Farran CA, Guo HC, Yu T, Fang HT, Wang HF, Schlesinger S, Seah YFS, Goh GYL, Neo SP, Li Y, Lorincz MC, Tergaonkar V, Lim TM, Chen L, Gunaratne J, Collins JJ, Goff SP, Daley GQ, Li H, Bard FA, Loh YH. Systematic identification of factors for provirus silencing in embryonic stem cells. Cell 2015; 163:230-45. [PMID: 26365490 DOI: 10.1016/j.cell.2015.08.037] [Citation(s) in RCA: 153] [Impact Index Per Article: 15.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/26/2014] [Revised: 07/07/2015] [Accepted: 08/13/2015] [Indexed: 12/25/2022]
Abstract
Embryonic stem cells (ESCs) repress the expression of exogenous proviruses and endogenous retroviruses (ERVs). Here, we systematically dissected the cellular factors involved in provirus repression in embryonic carcinomas (ECs) and ESCs by a genome-wide siRNA screen. Histone chaperones (Chaf1a/b), sumoylation factors (Sumo2/Ube2i/Sae1/Uba2/Senp6), and chromatin modifiers (Trim28/Eset/Atf7ip) are key determinants that establish provirus silencing. RNA-seq analysis uncovered the roles of Chaf1a/b and sumoylation modifiers in the repression of ERVs. ChIP-seq analysis demonstrates direct recruitment of Chaf1a and Sumo2 to ERVs. Chaf1a reinforces transcriptional repression via its interaction with members of the NuRD complex (Kdm1a, Hdac1/2) and Eset, while Sumo2 orchestrates the provirus repressive function of the canonical Zfp809/Trim28/Eset machinery by sumoylation of Trim28. Our study reports a genome-wide atlas of functional nodes that mediate proviral silencing in ESCs and illuminates the comprehensive, interconnected, and multi-layered genetic and epigenetic mechanisms by which ESCs repress retroviruses within the genome.
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Affiliation(s)
- Bin Xia Yang
- Epigenetics and Cell Fates Laboratory, A(∗)STAR Institute of Molecular and Cell Biology, 61 Biopolis Drive Proteos, Singapore 138673, Singapore
| | - Chadi A El Farran
- Epigenetics and Cell Fates Laboratory, A(∗)STAR Institute of Molecular and Cell Biology, 61 Biopolis Drive Proteos, Singapore 138673, Singapore; Department of Biological Sciences, National University of Singapore, Singapore 117543, Singapore
| | - Hong Chao Guo
- College of Life Sciences, Nankai University, Tianjin 300071, China
| | - Tao Yu
- Epigenetics and Cell Fates Laboratory, A(∗)STAR Institute of Molecular and Cell Biology, 61 Biopolis Drive Proteos, Singapore 138673, Singapore; Department of Biological Sciences, National University of Singapore, Singapore 117543, Singapore
| | - Hai Tong Fang
- Epigenetics and Cell Fates Laboratory, A(∗)STAR Institute of Molecular and Cell Biology, 61 Biopolis Drive Proteos, Singapore 138673, Singapore
| | - Hao Fei Wang
- Epigenetics and Cell Fates Laboratory, A(∗)STAR Institute of Molecular and Cell Biology, 61 Biopolis Drive Proteos, Singapore 138673, Singapore; Department of Biological Sciences, National University of Singapore, Singapore 117543, Singapore
| | - Sharon Schlesinger
- Department of Biochemistry and Molecular Biophysics, Columbia University, New York, NY 10032, USA; Department of Microbiology and Immunology, Columbia University, New York, NY 10032, USA
| | - Yu Fen Samantha Seah
- Epigenetics and Cell Fates Laboratory, A(∗)STAR Institute of Molecular and Cell Biology, 61 Biopolis Drive Proteos, Singapore 138673, Singapore
| | - Germaine Yen Lin Goh
- Membrane Traffic Laboratory, A(∗)STAR Institute of Molecular and Cell Biology, 61 Biopolis Drive Proteos, Singapore 138673, Singapore
| | - Suat Peng Neo
- Quantitative Proteomics Group, A(∗)STAR Institute of Molecular and Cell Biology, 61 Biopolis Drive Proteos, Singapore 138673, Singapore
| | - Yinghui Li
- Division of Cancer Genetics and Therapeutics, Laboratory of NF-κB Signaling, A(∗)STAR Institute of Molecular and Cell Biology, 61 Biopolis Drive Proteos, Singapore 138673, Singapore
| | - Matthew C Lorincz
- Department of Medical Genetics, Life Sciences Institute, University of British Columbia, Vancouver, BC V6T 1Z3, Canada
| | - Vinay Tergaonkar
- Division of Cancer Genetics and Therapeutics, Laboratory of NF-κB Signaling, A(∗)STAR Institute of Molecular and Cell Biology, 61 Biopolis Drive Proteos, Singapore 138673, Singapore; Department of Biochemistry, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 119077, Singapore
| | - Tit-Meng Lim
- Department of Biological Sciences, National University of Singapore, Singapore 117543, Singapore
| | - Lingyi Chen
- College of Life Sciences, Nankai University, Tianjin 300071, China
| | - Jayantha Gunaratne
- Quantitative Proteomics Group, A(∗)STAR Institute of Molecular and Cell Biology, 61 Biopolis Drive Proteos, Singapore 138673, Singapore; Department of Anatomy, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 117597, Singapore
| | - James J Collins
- Department of Biological Engineering, Synthetic Biology Center, Institute for Medical Engineering and Science, Massachusetts Institute of Technology, Cambridge, MA 02139, USA; Broad Institute of MIT and Harvard, Cambridge, MA 02139, USA; Wyss Institute for Biologically Inspired Engineering, Harvard University, Boston, MA 02115, USA; Howard Hughes Medical Institute, Boston, MA 02115, USA
| | - Stephen P Goff
- Department of Biochemistry and Molecular Biophysics, Columbia University, New York, NY 10032, USA; Department of Microbiology and Immunology, Columbia University, New York, NY 10032, USA; Howard Hughes Medical Institute, New York, NY 10032, USA
| | - George Q Daley
- Howard Hughes Medical Institute, Boston, MA 02115, USA; Stem Cell Transplantation Program, Division of Pediatric Hematology/Oncology, Boston Children's Hospital and Dana-Farber Cancer Institute, Boston, MA 02115, USA; Department of Biological Chemistry and Molecular Pharmacology, Harvard Medical School, Boston, MA 02115, USA; Harvard Stem Cell Institute, Boston, MA 02115, USA; Manton Center for Orphan Disease Research, Boston, MA 02115, USA
| | - Hu Li
- Center for Individualized Medicine, Department of Molecular Pharmacology & Experimental Therapeutics, Mayo Clinic, Rochester, MN 55905, USA
| | - Frederic A Bard
- Membrane Traffic Laboratory, A(∗)STAR Institute of Molecular and Cell Biology, 61 Biopolis Drive Proteos, Singapore 138673, Singapore; Department of Biochemistry, Yong Loo Lin School of Medicine, National University of Singapore, Singapore 119077, Singapore
| | - Yuin-Han Loh
- Epigenetics and Cell Fates Laboratory, A(∗)STAR Institute of Molecular and Cell Biology, 61 Biopolis Drive Proteos, Singapore 138673, Singapore; Department of Biological Sciences, National University of Singapore, Singapore 117543, Singapore.
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Cheng CT, Kuo CY, Ann DK. KAPtain in charge of multiple missions: Emerging roles of KAP1. World J Biol Chem 2014; 5:308-320. [PMID: 25225599 PMCID: PMC4160525 DOI: 10.4331/wjbc.v5.i3.308] [Citation(s) in RCA: 75] [Impact Index Per Article: 6.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/28/2013] [Revised: 03/21/2014] [Accepted: 06/20/2014] [Indexed: 02/05/2023] Open
Abstract
KAP1/TRIM28/TIF1β was identified nearly twenty years ago as a universal transcriptional co-repressor because it interacts with a large KRAB-containing zinc finger protein (KRAB-ZFP) transcription factor family. Many studies demonstrate that KAP1 affects gene expression by regulating the transcription of KRAB-ZFP-specific loci, trans-repressing as a transcriptional co-repressor or epigenetically modulating chromatin structure. Emerging evidence suggests that KAP1 also functions independent of gene regulation by serving as a SUMO/ubiquitin E3 ligase or signaling scaffold protein to mediate signal transduction. KAP1 is subjected to multiple post-translational modifications (PTMs), including serine/tyrosine phosphorylation, SUMOylation, and acetylation, which coordinately regulate KAP1 function and its protein abundance. KAP1 is involved in multiple aspects of cellular activities, including DNA damage response, virus replication, cytokine production and stem cell pluripotency. Moreover, knockout of KAP1 results in embryonic lethality, indicating that KAP1 is crucial for embryonic development and possibly impacts a wide-range of (patho)physiological manifestations. Indeed, studies from conditional knockout mouse models reveal that KAP1-deficiency significantly impairs vital physiological processes, such as immune maturation, stress vulnerability, hepatic metabolism, gamete development and erythropoiesis. In this review, we summarize and evaluate current literatures involving the biochemical and physiological functions of KAP1. In addition, increasing studies on the clinical relevance of KAP1 in cancer will also be discussed.
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Turelli P, Castro-Diaz N, Marzetta F, Kapopoulou A, Raclot C, Duc J, Tieng V, Quenneville S, Trono D. Interplay of TRIM28 and DNA methylation in controlling human endogenous retroelements. Genome Res 2014; 24:1260-70. [PMID: 24879559 PMCID: PMC4120080 DOI: 10.1101/gr.172833.114] [Citation(s) in RCA: 147] [Impact Index Per Article: 13.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/15/2022]
Abstract
Reverse transcription-derived sequences account for at least half of the human genome. Although these retroelements are formidable motors of evolution, they can occasionally cause disease, and accordingly are inactivated during early embryogenesis through epigenetic mechanisms. In the mouse, at least for endogenous retroviruses, important mediators of this process are the tetrapod-specific KRAB-containing zinc finger proteins (KRAB-ZFPs) and their cofactor TRIM28. The present study demonstrates that KRAB/TRIM28-mediated regulation is responsible for controlling a very broad range of human-specific endogenous retroelements (EREs) in human embryonic stem (ES) cells and that it exerts, as a consequence, a marked effect on the transcriptional dynamics of these cells. It further reveals reciprocal dependence between TRIM28 recruitment at specific families of EREs and DNA methylation. It finally points to the importance of persistent TRIM28-mediated control of ERE transcriptional impact beyond their presumed inactivation by DNA methylation.
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Affiliation(s)
- Priscilla Turelli
- School of Life Sciences and Frontiers in Genetics Program, Ecole Polytechnique Fédérale de Lausanne (EPFL), 1015 Lausanne, Switzerland
| | - Nathaly Castro-Diaz
- School of Life Sciences and Frontiers in Genetics Program, Ecole Polytechnique Fédérale de Lausanne (EPFL), 1015 Lausanne, Switzerland
| | - Flavia Marzetta
- School of Life Sciences and Frontiers in Genetics Program, Ecole Polytechnique Fédérale de Lausanne (EPFL), 1015 Lausanne, Switzerland
| | - Adamandia Kapopoulou
- School of Life Sciences and Frontiers in Genetics Program, Ecole Polytechnique Fédérale de Lausanne (EPFL), 1015 Lausanne, Switzerland
| | - Charlène Raclot
- School of Life Sciences and Frontiers in Genetics Program, Ecole Polytechnique Fédérale de Lausanne (EPFL), 1015 Lausanne, Switzerland
| | - Julien Duc
- School of Life Sciences and Frontiers in Genetics Program, Ecole Polytechnique Fédérale de Lausanne (EPFL), 1015 Lausanne, Switzerland
| | - Vannary Tieng
- Department of Pathology, Faculty of Medicine, University of Geneva, 1211 Geneva 4, Switzerland
| | - Simon Quenneville
- School of Life Sciences and Frontiers in Genetics Program, Ecole Polytechnique Fédérale de Lausanne (EPFL), 1015 Lausanne, Switzerland
| | - Didier Trono
- School of Life Sciences and Frontiers in Genetics Program, Ecole Polytechnique Fédérale de Lausanne (EPFL), 1015 Lausanne, Switzerland;
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25
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Dewannieux M, Heidmann T. Endogenous retroviruses: acquisition, amplification and taming of genome invaders. Curr Opin Virol 2013; 3:646-56. [DOI: 10.1016/j.coviro.2013.08.005] [Citation(s) in RCA: 92] [Impact Index Per Article: 7.7] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/10/2013] [Revised: 08/13/2013] [Accepted: 08/14/2013] [Indexed: 12/12/2022]
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EBP1, a novel host factor involved in primer binding site-dependent restriction of moloney murine leukemia virus in embryonic cells. J Virol 2013; 88:1825-9. [PMID: 24227866 DOI: 10.1128/jvi.02578-13] [Citation(s) in RCA: 18] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
Mouse embryonic cells are unable to support the replication of Moloney murine leukemia virus (MLV). The integrated viral DNA is transcriptionally silenced, largely due to binding of host transcriptional repressors to the primer binding site (PBS) of the provirus. We have previously shown that a PBS DNA-binding repressor complex contains ZFP809 and TRIM28. Here, we identified ErbB3-binding protein 1 (EBP1) to be a novel component of the ZFP809-TRIM28 silencing complex and show that EBP1 depletion reduces PBS-mediated retroviral silencing.
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Wolf G, Nielsen AL, Mikkelsen JG, Pedersen FS. Epigenetic marking and repression of porcine endogenous retroviruses. J Gen Virol 2013; 94:960-970. [PMID: 23324470 DOI: 10.1099/vir.0.049288-0] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/28/2022] Open
Abstract
Endogenous retroviruses (ERVs) are remnants of retroviral germ line infections and have been identified in all mammals investigated so far. Although the majority of ERVs are degenerated, some mammalian species, such as mice and pigs, carry replication-competent ERVs capable of forming infectious viral particles. In mice, ERVs are silenced by DNA methylation and histone modifications and some exogenous retroviruses were shown to be transcriptionally repressed after integration by a primer-binding site (PBS) targeting mechanism. However, epigenetic repression of porcine ERVs (PERVs) has remained largely unexplored so far. In this study, we screened the pig genome for PERVs using LTRharvest, a tool for de novo detection of ERVs, and investigated various aspects of epigenetic repression of three unrelated PERV families. We found that these PERV families are differentially up- or downregulated upon chemical inhibition of DNA methylation and histone deacetylation in cultured porcine cells. Furthermore, chromatin immunoprecipitation analysis revealed repressive histone methylation marks at PERV loci in primary porcine embryonic germ cells and immortalized embryonic kidney cells. PERV elements belonging to the PERV-γ1 family, which is the only known PERV family that has remained active up to the present, were marked by significantly higher levels of histone methylations than PERV-γ2 and PERV-β3 proviruses. Finally, we tested three PERV-associated PBS sequences for repression activity in murine and porcine cells using retroviral transduction experiments and showed that none of these PBS sequences induced immediate transcriptional silencing in the tested primary porcine cells.
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Affiliation(s)
- Gernot Wolf
- Department of Molecular Biology and Genetics, University of Aarhus, DK-8000 Aarhus C, Denmark
| | | | | | - Finn Skou Pedersen
- Department of Molecular Biology and Genetics, University of Aarhus, DK-8000 Aarhus C, Denmark
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Global and stage specific patterns of Krüppel-associated-box zinc finger protein gene expression in murine early embryonic cells. PLoS One 2013; 8:e56721. [PMID: 23451074 PMCID: PMC3579818 DOI: 10.1371/journal.pone.0056721] [Citation(s) in RCA: 40] [Impact Index Per Article: 3.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/25/2012] [Accepted: 01/14/2013] [Indexed: 01/24/2023] Open
Abstract
Highly coordinated transcription networks orchestrate the self-renewal of pluripotent stem cell and the earliest steps of mammalian development. KRAB-containing zinc finger proteins represent the largest group of transcription factors encoded by the genomes of higher vertebrates including mice and humans. Together with their putatively universal cofactor KAP1, they have been implicated in events as diverse as the silencing of endogenous retroelements, the maintenance of imprinting and the pluripotent self-renewal of embryonic stem cells, although the genomic targets and specific functions of individual members of this gene family remain largely undefined. Here, we first generated a list of Ensembl-annotated KRAB-containing genes encoding the mouse and human genomes. We then defined the transcription levels of these genes in murine early embryonic cells. We found that the majority of KRAB-ZFP genes are expressed in mouse pluripotent stem cells and other early progenitors. However, we also identified distinctively cell- or stage-specific patterns of expression, some of which are pluripotency-restricted. Finally, we determined that individual KRAB-ZFP genes exhibit highly distinctive modes of expression, even when grouped in genomic clusters, and that these cannot be correlated with the presence of prototypic repressive or activating chromatin marks. These results pave the way to delineating the role of specific KRAB-ZFPs in early embryogenesis.
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Fletcher AJ, Towers GJ. Inhibition of retroviral replication by members of the TRIM protein family. Curr Top Microbiol Immunol 2013; 371:29-66. [PMID: 23686231 DOI: 10.1007/978-3-642-37765-5_2] [Citation(s) in RCA: 18] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
The TRIM protein family is emerging as a central component of mammalian antiviral innate immunity. Beginning with the identification of TRIM5α as a mammalian post-entry restriction factor against retroviruses, to the repeated observation that many TRIMs ubiquitinate and regulate signaling pathways, the past decade has witnessed an intense research effort to understand how TRIM proteins influence immunity. The list of viral families targeted directly or indirectly by TRIM proteins has grown to include adenoviruses, hepadnaviruses, picornaviruses, flaviviruses, orthomyxoviruses, paramyxoviruses, herpesviruses, rhabdoviruses and arenaviruses. We have come to appreciate how, through intense bouts of positive selection, some TRIM genes have been honed into species-specific restriction factors. Similarly, in the case of TRIMCyp, we are beginning to understand how viruses too have mutated to evade restriction, suggesting that TRIM and viruses have coevolved for millions of years of primate evolution. Recently, TRIM5α returned to the limelight when it was shown to trigger the expression of antiviral genes upon recognition of an incoming virus, a paradigm shift that demonstrated that restriction factors make excellent pathogen sensors. However, it remains unclear how many of ~100 human TRIM genes are antiviral, despite the expression of many of these genes being upregulated by interferon and upon viral infection. TRIM proteins do not conform to one type of antiviral mechanism, reflecting the diversity of viruses they target. Moreover, the cofactors of restriction remain largely enigmatic. The control of retroviral replication remains an important medical subject and provides a useful backdrop for reviewing how TRIM proteins act to repress viral replication.
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Affiliation(s)
- Adam J Fletcher
- MRC Centre for Medical Molecular Virology, University College, London, UK.
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Abstract
Most proteins of the TRIM family (also known as RBCC family) are ubiquitin ligases that share a peculiar protein structure, characterized by including an N-terminal RING finger domain closely followed by one or two B-boxes. Additional protein domains found at their C termini have been used to classify TRIM proteins into classes. TRIMs are involved in multiple cellular processes and many of them are essential components of the innate immunity system of animal species. In humans, it has been shown that mutations in several TRIM-encoding genes lead to diverse genetic diseases and contribute to several types of cancer. They had been hitherto detected only in animals. In this work, by comprehensively analyzing the available diversity of TRIM and TRIM-like protein sequences and evaluating their evolutionary patterns, an improved classification of the TRIM family is obtained. Members of one of the TRIM subfamilies defined, called Subfamily A, turn to be present not only in animals, but also in many other eukaryotes, such as fungi, apusozoans, alveolates, excavates and plants. The rest of subfamilies are animal-specific and several of them originated only recently. Subfamily A proteins are characterized by containing a MATH domain, suggesting a potential evolutionary connection between TRIM proteins and a different type of ubiquitin ligases, known as TRAFs, which contain quite similar MATH domains. These results indicate that the TRIM family emerged much earlier than so far thought and contribute to our understanding of its origin and diversification. The structural and evolutionary links with the TRAF family of ubiquitin ligases can be experimentally explored to determine whether functional connections also exist.
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Affiliation(s)
- Ignacio Marín
- Instituto de Biomedicina de Valencia (IBV-CSIC), Consejo Superior de Investigaciones Científicas, Valencia, Spain.
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Abstract
Studies of retroviruses have been instrumental in revealing the existence of an array of antiviral proteins, or restriction factors, and the mechanisms by which they function. Some restriction factors appear to specifically inhibit retrovirus replication, while others have a broader antiviral action. Here, we briefly review current understanding of the mechanisms by which several such proteins exert antiviral activity. We also discuss how retroviruses have evolved to evade or antagonize antiviral proteins, including through the action of viral accessory proteins. Restriction factors, their viral targets and antagonists have exerted evolutionary pressure on each other, resulting in specialization and barriers to cross-species transmission. Potentially, this recently revealed intrinsic system of antiviral immunity might be mobilized for therapeutic benefit.
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Affiliation(s)
- Theodora Hatziioannou
- Aaron Diamond AIDS Research Center, The Rockefeller University, New York, NY 10016, United States
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32
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Sharma P, Azebi S, England P, Christensen T, Møller-Larsen A, Petersen T, Batsché E, Muchardt C. Citrullination of histone H3 interferes with HP1-mediated transcriptional repression. PLoS Genet 2012; 8:e1002934. [PMID: 23028349 PMCID: PMC3441713 DOI: 10.1371/journal.pgen.1002934] [Citation(s) in RCA: 72] [Impact Index Per Article: 5.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/22/2012] [Accepted: 07/16/2012] [Indexed: 11/18/2022] Open
Abstract
Multiple Sclerosis (MS) is an autoimmune disease associated with abnormal expression of a subset of cytokines, resulting in inappropriate T-lymphocyte activation and uncontrolled immune response. A key issue in the field is the need to understand why these cytokines are transcriptionally activated in the patients. Here, we have examined several transcription units subject to pathological reactivation in MS, including the TNFα and IL8 cytokine genes and also several Human Endogenous RetroViruses (HERVs). We find that both the immune genes and the HERVs require the heterochromatin protein HP1α for their transcriptional repression. We further show that the Peptidylarginine Deiminase 4 (PADI4), an enzyme with a suspected role in MS, weakens the binding of HP1α to tri-methylated histone H3 lysine 9 by citrullinating histone H3 arginine 8. The resulting de-repression of both cytokines and HERVs can be reversed with the PADI-inhibitor Cl-amidine. Finally, we show that in peripheral blood mononuclear cells (PBMCs) from MS patients, the promoters of TNFα, and several HERVs share a deficit in HP1α recruitment and an augmented accumulation of histone H3 with a double citrulline 8 tri-methyl lysine 9 modifications. Thus, our study provides compelling evidence that HP1α and PADI4 are regulators of both immune genes and HERVs, and that multiple events of transcriptional reactivation in MS patients can be explained by the deficiency of a single mechanism of gene silencing.
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Affiliation(s)
- Priyanka Sharma
- Institut Pasteur, Département de Biologie du Développement, CNRS URA2578, Unité de Régulation Epigénétique, Paris, France
| | - Saliha Azebi
- Institut Pasteur, Département de Biologie du Développement, CNRS URA2578, Unité de Régulation Epigénétique, Paris, France
| | - Patrick England
- Institut Pasteur, Département de Biologie Structurale et Chimie, CNRS UMR3528, Plate-Forme de Biophysique des Macromolécules et de Leurs Interactions, Paris, France
| | | | | | - Thor Petersen
- Department of Neurology, Aarhus University Hospital, Aarhus, Denmark
| | - Eric Batsché
- Institut Pasteur, Département de Biologie du Développement, CNRS URA2578, Unité de Régulation Epigénétique, Paris, France
| | - Christian Muchardt
- Institut Pasteur, Département de Biologie du Développement, CNRS URA2578, Unité de Régulation Epigénétique, Paris, France
- * E-mail:
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Suzuki Y, Chew ML, Suzuki Y. Role of host-encoded proteins in restriction of retroviral integration. Front Microbiol 2012; 3:227. [PMID: 22737148 PMCID: PMC3381236 DOI: 10.3389/fmicb.2012.00227] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/13/2012] [Accepted: 06/02/2012] [Indexed: 11/13/2022] Open
Abstract
In retroviral infections, a copy of the viral DNA is first synthesized from genomic RNA by reverse transcription and subsequently integrated into host chromatin. This integration step, executed by the viral enzyme integrase (IN), is one of the hallmarks of retroviral infection. Although an obligate role for IN in retroviral integration has been clearly defined by numerous biochemical analysis of its recombinant protein and genetic analysis of the viral IN gene, several host cellular proteins have also been implicated as key factors involved in the integration step during viral replication. Although studies on integration cofactors have mostly emphasized factors that aid the integration process either through direct or indirect association with IN, it has become apparent that host cells may also harbor proteins that act as inhibitors of retroviral integration. Intriguingly, some of these inhibitory proteins appear to hamper the integration process via posttranslational modifications of the components of the preintegration complex including IN. A better understanding of the molecular mechanisms leading to the inhibition of integration will provide us with clues for the development of new strategies for treating retroviral infections. In this review, we draw attention to recent insights regarding potential host cellular factors that restrict integration, and illustrate how these inhibitory effects are achieved.
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Affiliation(s)
- Yasutsugu Suzuki
- Department of Microbiology, Yong Loo Lin School of Medicine, National University of Singapore, Singapore
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Armstrong L, Lako M, Buckley N, Lappin TRJ, Murphy MJ, Nolta JA, Pittenger M, Stojkovic M. Editorial: Our top 10 developments in stem cell biology over the last 30 years. Stem Cells 2012; 30:2-9. [PMID: 22162299 DOI: 10.1002/stem.1007] [Citation(s) in RCA: 25] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/15/2022]
Abstract
To celebrate 30 years of peer-reviewed publication of cutting edge stem cell research in Stem Cells, the first journal devoted to this promising field, we pause to review how far we have come in the three-decade lifetime of the Journal. To do this, we will present our views of the 10 most significant developments that have advanced stem cell biology where it is today. With the increasing rate of new data, it is natural that the bulk of these developments would have occurred in recent years, but we must not think that stem cell biology is a young science. The idea of a stem cell has actually been around for quite a long time having appeared in the scientific literature as early as 1868 with Haeckels' concept of a stamzelle as an uncommitted or undifferentiated cell responsible for producing many types of new cells to repair the body [Naturliche Schopfungsgeschichte, 1868; Berlin: Georg Reimer] but it took many years to obtain hard evidence in support of this theory. Not until the work of James Till and Ernest McCulloch in the 1960s did we have proof of the existence of stem cells and until the derivation of embryonal carcinoma cells in the 1960s-1970s and the first embryonic stem cell in 1981, such adult or tissue-specific stem cells were the only known class. The first issue of Stem Cells was published in 1981; no small wonder that most of its papers were devoted to hematopoietic progenitors. More recently, induced pluripotent stem cells (iPSCs) have been developed, and this is proving to be a fertile area of investigation as shown by the volume of publications appearing not only in Stem Cells but also in other journals over the last 5 years. The reader will note that many of the articles in this special issue are concerned with iPSC; however, this reflects the current surge of interest in the topic rather than any deliberate attempt to ignore other areas of stem cell investigation.
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Shibata M, Blauvelt KE, Liem KF, García-García MJ. TRIM28 is required by the mouse KRAB domain protein ZFP568 to control convergent extension and morphogenesis of extra-embryonic tissues. Development 2012; 138:5333-43. [PMID: 22110054 DOI: 10.1242/dev.072546] [Citation(s) in RCA: 24] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/08/2023]
Abstract
TRIM28 is a transcriptional regulator that is essential for embryonic development and is implicated in a variety of human diseases. The roles of TRIM28 in distinct biological processes are thought to depend on its interaction with factors that determine its DNA target specificity. However, functional evidence linking TRIM28 to specific co-factors is scarce. chatwo, a hypomorphic allele of Trim28, causes embryonic lethality and defects in convergent extension and morphogenesis of extra-embryonic tissues. These phenotypes are remarkably similar to those of mutants in the Krüppel-associated box (KRAB) zinc finger protein ZFP568, providing strong genetic evidence that ZFP568 and TRIM28 control morphogenesis through a common molecular mechanism. We determined that chatwo mutations decrease TRIM28 protein stability and repressive activity, disrupting both ZFP568-dependent and ZFP568-independent roles of TRIM28. These results, together with the analysis of embryos bearing a conditional inactivation of Trim28 in embryonic-derived tissues, revealed that TRIM28 is differentially required by ZFP568 and other factors during the early stages of mouse embryogenesis. In addition to uncovering novel roles of TRIM28 in convergent extension and morphogenesis of extra-embryonic tissues, our characterization of chatwo mutants demonstrates that KRAB domain proteins are essential to determine some of the biological functions of TRIM28.
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Affiliation(s)
- Maho Shibata
- Department of Molecular Biology and Genetics, Cornell University, 526 Campus Road, Ithaca, NY 14853, USA
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36
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Allouch A, Di Primio C, Alpi E, Lusic M, Arosio D, Giacca M, Cereseto A. The TRIM family protein KAP1 inhibits HIV-1 integration. Cell Host Microbe 2011; 9:484-95. [PMID: 21669397 DOI: 10.1016/j.chom.2011.05.004] [Citation(s) in RCA: 100] [Impact Index Per Article: 7.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/03/2010] [Revised: 03/30/2011] [Accepted: 05/13/2011] [Indexed: 01/05/2023]
Abstract
The integration of viral cDNA into the host genome is a critical step in the life cycle of HIV-1. This step is catalyzed by integrase (IN), a viral enzyme that is positively regulated by acetylation via the cellular histone acetyl transferase (HAT) p300. To investigate the relevance of IN acetylation, we searched for cellular proteins that selectively bind acetylated IN and identified KAP1, a protein belonging to the TRIM family of antiviral proteins. KAP1 binds acetylated IN and induces its deacetylation through the formation of a protein complex which includes the deacetylase HDAC1. Modulation of intracellular KAP1 levels in different cell types including T cells, the primary HIV-1 target, revealed that KAP1 curtails viral infectivity by selectively affecting HIV-1 integration. This study identifies KAP1 as a cellular factor restricting HIV-1 infection and underscores the relevance of IN acetylation as a crucial step in the viral infectious cycle.
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Affiliation(s)
- Awatef Allouch
- Molecular Biology Laboratory, Scuola Normale Superiore, Piazza dei Cavalieri 7, Pisa 56126, Italy
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The novel Nrf2-interacting factor KAP1 regulates susceptibility to oxidative stress by promoting the Nrf2-mediated cytoprotective response. Biochem J 2011; 436:387-97. [PMID: 21382013 DOI: 10.1042/bj20101748] [Citation(s) in RCA: 23] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/07/2023]
Abstract
The transcription factor Nrf2 (nuclear factor-erythroid 2-related factor 2) co-ordinately regulates ARE (antioxidant-response element)-mediated induction of cytoprotective genes in response to electrophiles and oxidative stress; however, the molecular mechanism controlling Nrf2-dependent gene expression is not fully understood. To identify factors that regulate Nrf2-dependent transcription, we searched for proteins that interact with the Nrf2-NT (N-terminal Nrf2 transactivation domain) by affinity purification from HeLa nuclear extracts. In the present study, we identified KAP1 [KRAB (Krüppel-associated box)-associated protein 1] as a novel Nrf2-NT-interacting protein. Pull-down analysis confirmed the interaction between KAP1 and Nrf2 in cultured cells and demonstrated that the N-terminal region of KAP1 binds to Nrf2-NT in vitro. Reporter assays showed that KAP1 facilitates Nrf2 transactivation activity in a dose-dependent manner. Furthermore, the induction of the Nrf2-dependent expression of HO-1 (haem oxygenase-1) and NQO1 [NAD(P)H quinone oxidoreductase 1] by DEM (diethyl maleate) was attenuated by KAP1 knockdown in NIH 3T3 fibroblasts. This finding established that KAP1 acts as a positive regulator of Nrf2. Although Nrf2 nuclear accumulation was unaffected by KAP1 knockdown, the ability of Nrf2 to bind to the regulatory region of HO-1 and NQO1 was reduced. Moreover, KAP1 knockdown enhanced the sensitivity of NIH 3T3 cells to tert-butylhydroquinone, H2O2 and diamide. These results support our contention that KAP1 participates in the oxidative stress response by maximizing Nrf2-dependent transcription.
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Yu S, Gao B, Duan Z, Xu W, Xiong S. Identification of tripartite motif-containing 22 (TRIM22) as a novel NF-κB activator. Biochem Biophys Res Commun 2011; 410:247-51. [PMID: 21651891 DOI: 10.1016/j.bbrc.2011.05.124] [Citation(s) in RCA: 48] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/15/2011] [Accepted: 05/21/2011] [Indexed: 01/01/2023]
Abstract
Increasing evidence suggests that TRIM family proteins may play important roles in the regulation of innate immune signaling pathways. Here we report TRIM22 is involved in the activation of NF-κB. It was found that overexpression of TRIM22 could dose-dependently activate NF-κB as demonstrated by reporter gene assay and electrophoretic mobility shift assay, but had no effect on the activity of other transcription factors, including NF-AT, AP-1, C/EBP and IRFs. Further study showed that both the N-terminal RING domain and C-terminal SPRY domain were crucial for TRIM22-mediated NF-κB activation. Moreover, our results revealed that TRIM22 overexpression could significantly induce the secretion of pro-inflammatory cytokines by human macrophage cell line U937 in an NF-κB-dependent manner. These data suggested that TRIM22 was a positive regulator of NF-κB-mediated transcription.
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Affiliation(s)
- Shanshan Yu
- Institute for Immunobiology, Department of Immunology, Shanghai Medical College of Fudan University, Shanghai 200032, PR China
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39
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Rowe HM, Trono D. Dynamic control of endogenous retroviruses during development. Virology 2011; 411:273-87. [PMID: 21251689 DOI: 10.1016/j.virol.2010.12.007] [Citation(s) in RCA: 214] [Impact Index Per Article: 15.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/02/2010] [Accepted: 12/06/2010] [Indexed: 02/07/2023]
Abstract
Close to half of the human genome encompasses mobile genetic elements, most of which are retrotransposons. These genetic invaders are formidable evolutionary forces that have shaped the architecture of the genomes of higher organisms, with some conserving the ability to induce new integrants within their hosts' genome. Expectedly, the control of endogenous retroviruses is tight and multi-pronged. It is most crucially established in the germ line and during the first steps of embryogenesis, primarily through transcriptional mechanisms that have likely evolved under their very pressure, but are now engaged in controlling gene expression at large, notably during early development.
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Affiliation(s)
- Helen M Rowe
- National Program, Ecole Polytechnique Fédérale de Lausanne, 1015 Lausanne, Switzerland
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Kamata M, Liu S, Liang M, Nagaoka Y, Chen ISY. Generation of human induced pluripotent stem cells bearing an anti-HIV transgene by a lentiviral vector carrying an internal murine leukemia virus promoter. Hum Gene Ther 2010; 21:1555-67. [PMID: 20524893 DOI: 10.1089/hum.2010.050] [Citation(s) in RCA: 21] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/13/2022] Open
Abstract
The recent development of induced pluripotent stem cells (iPSCs) by ectopic expression of defined reprogramming factors offers enormous therapeutic opportunity. To deliver these factors, murine leukemia virus (MLV)-based vectors have been broadly used in the setting of hematopoietic stem cell transplantation. However, MLV vectors have been implicated in malignancy induced by insertional mutagenesis, whereas lentiviral vectors have not. Furthermore, the infectivity of MLV vectors is limited to dividing cells, whereas lentiviral vectors can also transduce nondividing cells. One important characteristic of MLV vectors is a self-silencing property of the promoter element in pluripotent stem cells, allowing temporal transgene expression in a nonpluripotent state before iPSC derivation. Here we test iPSC generation using a novel chimeric vector carrying a mutant MLV promoter internal to a lentiviral vector backbone, thereby containing the useful properties of both types of vectors. Transgene expression of this chimeric vector was highly efficient compared with that of MLV vectors and was silenced specifically in human embryonic stem cells. Human fetal fibroblasts transduced with the vector encoding each factor were efficiently reprogrammed into a pluripotent state, and these iPSCs had potential to differentiate into a variety of cell types. To explore the possibility of iPSCs for gene therapy, we established iPSC clones expressing a short hairpin RNA (shRNA) targeting chemokine receptor 5 (CCR5), the main coreceptor for HIV-1. Using a reporter construct for CCR5 expression, we confirmed that CCR5 shRNA was expressed and specifically knocked down the reporter expression in iPSCs. These data indicate that our chimeric lentiviral vector is a valuable tool for generation of iPSCs and the combination with vectors encoding transgenes allows for rapid establishment of desired genetically engineered iPSC lines.
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Affiliation(s)
- Masakazu Kamata
- Department of Microbiology, Immunology and Molecular Genetics, University of California at Los Angeles, David Geffen School of Medicine, Los Angeles, CA 90095, USA
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Abstract
The dynamics of embryonic stem cell pluripotency is orchestrated by an interplay of transcriptional and epigenetic regulation in a systematic and modular manner. While the ES cell stage is marked by multiple loci with bivalent chromatin marks that prepare genes for imminent activation on differentiation, this open chromatin conformation is tempered by repressive machinery that prevent premature expression of key developmental genes. This review serves to highlight key ES transcription factors and their known links to the epigenetic machinery via known protein complexes.
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Affiliation(s)
- Clara Y Cheong
- Stem Cell and Developmental Biology, Genome Institute of Singapore, Singapore, Singapore
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Gao B, Wang Y, Xu W, Duan Z, Xiong S. A 5′ Extended IFN-Stimulating Response Element Is Crucial for IFN-γ–Induced Tripartite Motif 22 Expression via Interaction with IFN Regulatory Factor-1. THE JOURNAL OF IMMUNOLOGY 2010; 185:2314-23. [DOI: 10.4049/jimmunol.1001053] [Citation(s) in RCA: 15] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/31/2022]
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Matsui T, Leung D, Miyashita H, Maksakova IA, Miyachi H, Kimura H, Tachibana M, Lorincz MC, Shinkai Y. Proviral silencing in embryonic stem cells requires the histone methyltransferase ESET. Nature 2010; 464:927-31. [PMID: 20164836 DOI: 10.1038/nature08858] [Citation(s) in RCA: 597] [Impact Index Per Article: 39.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/10/2009] [Accepted: 01/16/2010] [Indexed: 12/13/2022]
Abstract
Endogenous retroviruses (ERVs), retrovirus-like elements with long terminal repeats, are widely dispersed in the euchromatic compartment in mammalian cells, comprising approximately 10% of the mouse genome. These parasitic elements are responsible for >10% of spontaneous mutations. Whereas DNA methylation has an important role in proviral silencing in somatic and germ-lineage cells, an additional DNA-methylation-independent pathway also functions in embryonal carcinoma and embryonic stem (ES) cells to inhibit transcription of the exogenous gammaretrovirus murine leukaemia virus (MLV). Notably, a recent genome-wide study revealed that ERVs are also marked by histone H3 lysine 9 trimethylation (H3K9me3) and H4K20me3 in ES cells but not in mouse embryonic fibroblasts. However, the role that these marks have in proviral silencing remains unexplored. Here we show that the H3K9 methyltransferase ESET (also called SETDB1 or KMT1E) and the Krüppel-associated box (KRAB)-associated protein 1 (KAP1, also called TRIM28) are required for H3K9me3 and silencing of endogenous and introduced retroviruses specifically in mouse ES cells. Furthermore, whereas ESET enzymatic activity is crucial for HP1 binding and efficient proviral silencing, the H4K20 methyltransferases Suv420h1 and Suv420h2 are dispensable for silencing. Notably, in DNA methyltransferase triple knockout (Dnmt1(-/-)Dnmt3a(-/-)Dnmt3b(-/-)) mouse ES cells, ESET and KAP1 binding and ESET-mediated H3K9me3 are maintained and ERVs are minimally derepressed. We propose that a DNA-methylation-independent pathway involving KAP1 and ESET/ESET-mediated H3K9me3 is required for proviral silencing during the period early in embryogenesis when DNA methylation is dynamically reprogrammed.
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Affiliation(s)
- Toshiyuki Matsui
- Experimental Research Center for Infectious Diseases, Institute for Virus Research, Kyoto University, 53 Shogoin, Kawara-cho, Sakyo-ku, Kyoto 606-8507, Japan
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Hwang CY, Holl J, Rajan D, Lee Y, Kim S, Um M, Kwon KS, Song B. Hsp70 interacts with the retroviral restriction factor TRIM5alpha and assists the folding of TRIM5alpha. J Biol Chem 2010; 285:7827-37. [PMID: 20053985 DOI: 10.1074/jbc.m109.040618] [Citation(s) in RCA: 11] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/24/2023] Open
Abstract
Tripartite motif (TRIM) protein TRIM5alpha has been shown to restrict human immunodeficiency virus, type 1 infection in Old World monkey cells at the early post-entry step by poorly understood mechanisms. Currently, the physiological function of TRIM5alpha is not known. In this study, we showed that transiently overexpressed TRIM5alpha causes a morphological change in HEK293T cells. A proteomics analysis of the protein complexes that were pulled down with hemagglutinin-tagged TRIM5alpha suggested that the heat shock protein 70 (Hsp70) may serve as a TRIM5alpha-binding partner. The interaction between Hsp70 and TRIM5alpha was confirmed by co-localization and co-immunoprecipitation assays. Co-expression of Hsp70 reversed the TRIM5alpha-induced morphological change in HEK293T cells. Another heat shock protein Hsc70 also bound to TRIM5alpha, but unlike Hsp70, Hsc70 was not able to reverse the TRIM5alpha-induced morphological change, suggesting that Hsp70 specifically reverses the morphological change caused by TRIM5alpha. Studies using a series of TRIM5alpha deletion mutants demonstrate that, although the PRYSPRY domain is critical for binding to Hsp70, the entire TRIM5alpha structure is necessary to induce the morphological change of cells. When the ATPase domain of Hsp70 was mutated, the mutated Hsp70 could not counteract the morphological change induced by TRIM5alpha, indicating that the catalytic activity of Hsp70 protein is important for this function. Co-expression of Hsp70 elevated the levels of TRIM5alpha in the detergent-soluble fraction with a concomitant decrease in the detergent-insoluble fraction. Together these results suggest that Hsp70 plays critical roles in the cellular management against the TRIM5alpha-induced cellular insults.
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Affiliation(s)
- Chae Young Hwang
- Department of Pediatrics, Emory University School of Medicine, Atlanta, Georgia 30322, USA
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Restriction factors of retroviral replication: the example of Tripartite Motif (TRIM) protein 5 alpha and 22. Amino Acids 2009; 39:1-9. [PMID: 19943174 DOI: 10.1007/s00726-009-0393-x] [Citation(s) in RCA: 14] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/28/2009] [Accepted: 11/11/2009] [Indexed: 12/26/2022]
Abstract
Viral tropism, replication, and pathogenesis are determined by multiple interactions between the pathogen and the host. In the case of retroviruses, and in particular, the human immunodeficiency virus, the specific interaction of the envelope protein with the host receptors and co-receptors is essential to gain entry in the cells. After entry, the success of retroviruses to complete their life cycle depends on a complex interplay between the virus and host proteins. Indeed, the cell environment is endowed with a number of factors that actively block distinct stage(s) in the microbial life cycle. Among these restriction factors, Tripartite Motif-5 alpha (TRIM5 alpha) has been extensively studied; however, other TRIM family members have been demonstrated to be anti-retroviral effector proteins. This article reviews, in particular, the current knowledge on the anti-retroviral effects of TRIM5 alpha and TRIM22.
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Gao B, Duan Z, Xu W, Xiong S. Tripartite motif-containing 22 inhibits the activity of hepatitis B virus core promoter, which is dependent on nuclear-located RING domain. Hepatology 2009; 50:424-33. [PMID: 19585648 DOI: 10.1002/hep.23011] [Citation(s) in RCA: 172] [Impact Index Per Article: 10.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 12/12/2022]
Abstract
UNLABELLED Members of the tripartite motif (TRIM) family are a part of the innate immune system to counter intracellular pathogens. TRIM22 has been reported to possess antiretroviral activity. Here we report that TRIM22 is involved in antiviral immunity against hepatitis B virus (HBV). Our results showed that TRIM22, being a strongly induced gene by interferons in human hepatoma HepG2 cells, could inhibit HBV gene expression and replication in a cell culture system as well as in a mouse model system. Importantly, it was found that TRIM22 could inhibit the activity of HBV core promoter (CP) in a dose-dependent manner. However, TRIM22 lacking the C terminal SPRY domain lost this activity. Further study showed that the SPRY domain deletion mutant was localized exclusively to the cytoplasm of HepG2 cells. In contrast, the wild-type TRIM22 was localized to the nucleus, as expected for a transcriptional suppressor. Interestingly, although RING domain mutants of TRIM22 were localized to the nucleus, they could not inhibit HBV CP activity, indicating that TRIM22-mediated anti-HBV activity was dependent on the nuclear-located RING domain. CONCLUSION These findings suggest that TRIM22, which exhibits anti-HBV activity by acting as a transcriptional suppressor, may play an important role in the clearance of HBV.
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Affiliation(s)
- Bo Gao
- Department of Immunology, Institute for Immunobiology, Shanghai Medical College of Fudan University, People's Republic of China
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Abstract
Embryonic stem cells (ESCs) and other primitive stem cells of mice have been known for more than 30 years to potently block retrovirus replication. Infection of ESCs by the murine leukaemia viruses (MLVs) results in the normal establishment of integrated proviral DNA, but this DNA is then transcriptionally silenced, preventing further viral spread. The repression is largely mediated by trans-acting factors that recognize a conserved sequence element termed the primer binding site, an 18-base pair sequence complementary to the 3' end of a cellular transfer RNA. A specific tRNA is annealed to the primer binding site sequence of the viral genomic RNA, and is used to prime DNA synthesis. This same sequence in the context of the integrated proviral DNA is targeted for silencing in ESCs. We have recently shown that a large protein complex binding to the primer binding site in ESCs contains TRIM28 (refs 8, 9), a well-characterized transcriptional co-repressor. An important question remains as to the identity of the factor that directly recognizes integrated retroviral DNAs and recruits TRIM28 to mediate their specific silencing. Here we identify the zinc finger protein ZFP809 as the recognition molecule that bridges the integrated proviral DNA and TRIM28. We show that expression of ZFP809 is sufficient to render even differentiated cells highly resistant to MLV infection. Furthermore, we demonstrate that ZFP809 is able to potently block transcription from DNA constructs of human T-cell lymphotropic virus-1 (HTLV-1), which use the same primer tRNA. These results identify ZFP809 as a DNA-binding factor that specifically recognizes a large subset of mammalian retroviruses and retroelements, targeting them for transcriptional silencing. We propose that ZFP809 evolved as a stem-cell-specific retroviral restriction factor, and therefore constitutes a new component of the intrinsic immune system of stem cells.
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Affiliation(s)
- Daniel Wolf
- Howard Hughes Medical Institute, Department of Biochemistry and Molecular Biophysics, Columbia University, College of Physicians and Surgeons, New York, NY 10032 USA
| | - Stephen P. Goff
- Howard Hughes Medical Institute, Department of Biochemistry and Molecular Biophysics, Columbia University, College of Physicians and Surgeons, New York, NY 10032 USA
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48
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Preferential epigenetic suppression of the autonomous MusD over the nonautonomous ETn mouse retrotransposons. Mol Cell Biol 2009; 29:2456-68. [PMID: 19273603 DOI: 10.1128/mcb.01383-08] [Citation(s) in RCA: 12] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
Nonautonomous retrotransposon subfamilies are often amplified in preference to their coding-competent relatives. However, the mechanisms responsible for such replicative success are poorly understood. Here, we demonstrate that the autonomous MusD long terminal repeat (LTR) retrotransposons are subject to greater epigenetic silencing than their nonautonomous cousins, the early transposons (ETns), which are expressed at a 170-fold-higher level than MusD in mouse embryonic stem (ES) cells. We show that, in ES cells, 5' LTRs and the downstream region of MusD elements are more heavily methylated and are associated with less-activating and more-repressive histone modifications than the highly similar ETnII sequences. The internal region of MusD likely contributes to their silencing, as transgenes with MusD, compared to those with ETnII sequences, show reduced reporter gene expression and a higher level of repressive histone marks. Genomic distribution patterns of MusD and ETn elements are consistent with stronger selection against MusD elements within introns, suggesting that MusD-associated silencing marks can negatively impact genes. We propose a model in which nonautonomous retrotransposons may gain transcriptional and retrotranspositional advantages over their coding-competent counterparts by elimination of the CpG-rich retroviral sequence targeting the autonomous subfamilies for silencing.
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49
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Abstract
Up to 10% of the mouse genome is comprised of endogenous retrovirus (ERV) sequences, and most represent the remains of ancient germ line infections. Our knowledge of the three distinct classes of ERVs is inversely correlated with their copy number, and their characterization has benefited from the availability of divergent wild mouse species and subspecies, and from ongoing analysis of the Mus genome sequence. In contrast to human ERVs, which are nearly all extinct, active mouse ERVs can still be found in all three ERV classes. The distribution and diversity of ERVs has been shaped by host-virus interactions over the course of evolution, but ERVs have also been pivotal in shaping the mouse genome by altering host genes through insertional mutagenesis, by adding novel regulatory and coding sequences, and by their co-option by host cells as retroviral resistance genes. We review mechanisms by which an adaptive coexistence has evolved. (Part of a multi-author review).
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Affiliation(s)
- C. Stocking
- Heinrich-Pette-Institute, Martinistrasse 52, 20251 Hamburg, Germany
| | - C. A. Kozak
- Laboratory of Molecular Microbiology, National Institute of Allergy and Infectious Diseases, National Institutes of Health, 4 Center Drive MSC 0460, Bethesda, Maryland, 20892-0460 USA
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