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1.
Viruses ; 13(7)2021 06 30.
Artigo em Inglês | MEDLINE | ID: mdl-34209034

RESUMO

Host plasma membrane protein SERINC5 is incorporated into budding retrovirus particles where it blocks subsequent entry into susceptible target cells. Three structurally unrelated proteins encoded by diverse retroviruses, human immunodeficiency virus type 1 (HIV-1) Nef, equine infectious anemia virus (EIAV) S2, and ecotropic murine leukemia virus (MLV) GlycoGag, disrupt SERINC5 antiviral activity by redirecting SERINC5 from the site of virion assembly on the plasma membrane to an internal RAB7+ endosomal compartment. Pseudotyping retroviruses with particular glycoproteins, e.g., vesicular stomatitis virus glycoprotein (VSV G), renders the infectivity of particles resistant to inhibition by virion-associated SERINC5. To better understand viral determinants for SERINC5-sensitivity, the effect of SERINC5 was assessed using HIV-1, MLV, and Mason-Pfizer monkey virus (M-PMV) virion cores, pseudotyped with glycoproteins from Arenavirus, Coronavirus, Filovirus, Rhabdovirus, Paramyxovirus, and Orthomyxovirus genera. SERINC5 restricted virions pseudotyped with glycoproteins from several retroviruses, an orthomyxovirus, a rhabdovirus, a paramyxovirus, and an arenavirus. Infectivity of particles pseudotyped with HIV-1, amphotropic-MLV (A-MLV), or influenza A virus (IAV) glycoproteins, was decreased by SERINC5, whether the core was provided by HIV-1, MLV, or M-PMV. In contrast, particles pseudotyped with glycoproteins from M-PMV, parainfluenza virus 5 (PIV5), or rabies virus (RABV) were sensitive to SERINC5, but only with particular retroviral cores. Resistance to SERINC5 did not correlate with reduced SERINC5 incorporation into particles, route of viral entry, or absolute infectivity of the pseudotyped virions. These findings indicate that some non-retroviruses may be sensitive to SERINC5 and that, in addition to the viral glycoprotein, the retroviral core influences sensitivity to SERINC5.


Assuntos
Interações Hospedeiro-Patógeno , Proteínas de Membrana/genética , Proteínas do Envelope Viral , Vírion/metabolismo , Vírus/metabolismo , Células HEK293 , HIV-1/metabolismo , Humanos , Vírus da Leucemia Murina/metabolismo , Proteínas de Membrana/imunologia , Retroviridae/classificação , Retroviridae/metabolismo , Proteínas do Envelope Viral/genética , Proteínas do Envelope Viral/imunologia , Vírion/genética , Internalização do Vírus , Vírus/química , Vírus/classificação , Vírus/genética
2.
Viruses ; 13(3)2021 03 10.
Artigo em Inglês | MEDLINE | ID: mdl-33801908

RESUMO

An evolutionary arms race has been ongoing between retroviruses and their primate hosts for millions of years. Within the last century, a zoonotic transmission introduced the Human Immunodeficiency Virus (HIV-1), a retrovirus, to the human population that has claimed the lives of millions of individuals and is still infecting over a million people every year. To counteract retroviruses such as this, primates including humans have evolved an innate immune sensor for the retroviral capsid lattice known as TRIM5α. Although the molecular basis for its ability to restrict retroviruses is debated, it is currently accepted that TRIM5α forms higher-order assemblies around the incoming retroviral capsid that are not only disruptive for the virus lifecycle, but also trigger the activation of an antiviral state. More recently, it was discovered that TRIM5α restriction is broader than previously thought because it restricts not only the human retroelement LINE-1, but also the tick-borne flaviviruses, an emergent group of RNA viruses that have vastly different strategies for replication compared to retroviruses. This review focuses on the underlying mechanisms of TRIM5α-mediated restriction of retroelements and flaviviruses and how they differ from the more widely known ability of TRIM5α to restrict retroviruses.


Assuntos
Capsídeo/imunologia , Imunidade Inata , Vírus de RNA/imunologia , Vírus de RNA/metabolismo , Proteínas com Motivo Tripartido/metabolismo , Ubiquitina-Proteína Ligases/metabolismo , Capsídeo/metabolismo , Proteínas de Transporte/genética , Flavivirus/imunologia , Flavivirus/metabolismo , Humanos , Vírus de RNA/classificação , Vírus de RNA/genética , Retroviridae/imunologia , Retroviridae/metabolismo , Infecções por Retroviridae/imunologia , Infecções por Retroviridae/prevenção & controle , Proteínas com Motivo Tripartido/genética , Proteínas com Motivo Tripartido/imunologia , Ubiquitina-Proteína Ligases/genética , Ubiquitina-Proteína Ligases/imunologia
3.
Nat Commun ; 12(1): 2187, 2021 04 12.
Artigo em Inglês | MEDLINE | ID: mdl-33846346

RESUMO

The RNA-sensing pathway contributes to type I interferon (IFN) production induced by DNA damaging agents. However, the potential involvement of RNA sensors in DNA repair is unknown. Here, we found that retinoic acid-inducible gene I (RIG-I), a key cytosolic RNA sensor that recognizes RNA virus and initiates the MAVS-IRF3-type I IFN signaling cascade, is recruited to double-stranded breaks (DSBs) and suppresses non-homologous end joining (NHEJ). Mechanistically, RIG-I interacts with XRCC4, and the RIG-I/XRCC4 interaction impedes the formation of XRCC4/LIG4/XLF complex at DSBs. High expression of RIG-I compromises DNA repair and sensitizes cancer cells to irradiation treatment. In contrast, depletion of RIG-I renders cells resistant to irradiation in vitro and in vivo. In addition, this mechanism suggests a protective role of RIG-I in hindering retrovirus integration into the host genome by suppressing the NHEJ pathway. Reciprocally, XRCC4, while suppressed for its DNA repair function, has a critical role in RIG-I immune signaling through RIG-I interaction. XRCC4 promotes RIG-I signaling by enhancing oligomerization and ubiquitination of RIG-I, thereby suppressing RNA virus replication in host cells. In vivo, silencing XRCC4 in mouse lung promotes influenza virus replication in mice and these mice display faster body weight loss, poorer survival, and a greater degree of lung injury caused by influenza virus infection. This reciprocal regulation of RIG-I and XRCC4 reveals a new function of RIG-I in suppressing DNA repair and virus integration into the host genome, and meanwhile endues XRCC4 with a crucial role in potentiating innate immune response, thereby helping host to prevail in the battle against virus.


Assuntos
Proteína DEAD-box 58/metabolismo , Reparo do DNA , Proteínas de Ligação a DNA/metabolismo , Receptores Imunológicos/metabolismo , Transdução de Sinais/imunologia , Células A549 , Animais , Quebras de DNA de Cadeia Dupla/efeitos da radiação , Reparo do DNA por Junção de Extremidades/efeitos da radiação , Reparo do DNA/efeitos da radiação , Relação Dose-Resposta à Radiação , Genoma Humano , Células HEK293 , Humanos , Camundongos , Radiação Ionizante , Retroviridae/metabolismo , Replicação Viral/efeitos da radiação
4.
Commun Biol ; 4(1): 325, 2021 03 11.
Artigo em Inglês | MEDLINE | ID: mdl-33707565

RESUMO

Congenital cataracts are associated with gene mutations, yet the underlying mechanism remains largely unknown. Here we reported an embryonic chick lens model that closely recapitulates the process of cataract formation. We adopted dominant-negative site mutations that cause congenital cataracts, connexin, Cx50E48K, aquaporin 0, AQP0R33C, αA-crystallin, CRYAA R12C and R54C. The recombinant retroviruses containing these mutants were microinjected into the occlusive lumen of chick lenses at early embryonic development. Cx50E48K expression developed cataracts associated with disorganized nuclei and enlarged extracellular spaces. Expression of AQP0R33C resulted in cortical cataracts, enlarged extracellular spaces and distorted fiber cell organization. αA crystallin mutations distorted lens light transmission and increased crystalline protein aggregation. Together, retroviral expression of congenital mutant genes in embryonic chick lenses closely mimics characteristics of human congenital cataracts. This model will provide an effective, reliable in vivo system to investigate the development and underlying mechanism of cataracts and other genetic diseases.


Assuntos
Aquaporinas/genética , Catarata/congênito , Conexinas/genética , Cristalinas/genética , Proteínas do Olho/genética , Cristalino/anormalidades , Mutação , Animais , Aquaporinas/metabolismo , Catarata/metabolismo , Catarata/patologia , Embrião de Galinha , Conexinas/metabolismo , Cristalinas/metabolismo , Modelos Animais de Doenças , Proteínas do Olho/metabolismo , Técnicas de Transferência de Genes , Predisposição Genética para Doença , Vetores Genéticos , Cristalino/metabolismo , Microinjeções , Fenótipo , Retroviridae/genética , Retroviridae/metabolismo
5.
Viruses ; 13(2)2021 02 12.
Artigo em Inglês | MEDLINE | ID: mdl-33673243

RESUMO

In normal cells APOBEC3 (A3A-A3H) enzymes as part of the innate immune system deaminate cytosine to uracil on single-stranded DNA (ssDNA) to scramble DNA in order to give protection against a range of exogenous retroviruses, DNA-based parasites, and endogenous retroelements. However, some viruses and cancer cells use these enzymes, especially A3A and A3B, to escape the adaptive immune response and thereby lead to the evolution of drug resistance. We have synthesized first-in-class inhibitors featuring modified ssDNA. We present models based on small-angle X-ray scattering (SAXS) data that (1) confirm that the mode of binding of inhibitor to an active A3B C-terminal domain construct in the solution state is the same as the mode of binding substrate to inactive mutants of A3A and A3B revealed in X-ray crystal structures and (2) give insight into the disulfide-linked inactive dimer formed under the oxidizing conditions of purification.


Assuntos
Citidina Desaminase/química , Citidina Desaminase/metabolismo , DNA de Cadeia Simples/química , Antígenos de Histocompatibilidade Menor/química , Antígenos de Histocompatibilidade Menor/metabolismo , RNA Viral/química , Infecções por Retroviridae/enzimologia , Retroviridae/genética , Domínio Catalítico , Citidina Desaminase/genética , DNA de Cadeia Simples/metabolismo , Dimerização , Humanos , Antígenos de Histocompatibilidade Menor/genética , Mutação , RNA Viral/genética , RNA Viral/metabolismo , Retroviridae/metabolismo , Infecções por Retroviridae/genética , Infecções por Retroviridae/virologia , Espalhamento a Baixo Ângulo
6.
Arch Virol ; 166(3): 733-753, 2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-33502593

RESUMO

The chronic dysfunction of neuronal cells, both central and peripheral, a characteristic of neurological disorders, may be caused by irreversible damage and cell death. In 2016, more than 276 million cases of neurological disorders were reported worldwide. Moreover, neurological disorders are the second leading cause of death. Generally, the etiology of neurological diseases is not fully understood. Recent studies have related the onset of neurological disorders to viral infections, which may cause neurological symptoms or lead to immune responses that trigger these pathological signs. Currently, this relationship is mostly based on epidemiological data on infections and seroprevalence of patients who present with neurological disorders. The number of studies aiming to elucidate the mechanism of action by which viral infections may directly or indirectly contribute to the development of neurological disorders has been increasing over the years but these studies are still scarce. Comprehending the pathogenesis of these diseases and exploring novel theories may favor the development of new strategies for diagnosis and therapy in the future. Therefore, the objective of the present study was to review the main pieces of evidence for the relationship between viral infection and neurological disorders such as Alzheimer's disease, Parkinson's disease, Guillain-Barré syndrome, multiple sclerosis, and epilepsy. Viruses belonging to the families Herpesviridae, Orthomyxoviridae, Flaviviridae, and Retroviridae have been reported to be involved in one or more of these conditions. Also, neurological symptoms and the future impact of infection with SARS-CoV-2, a member of the family Coronaviridae that is responsible for the COVID-19 pandemic that started in late 2019, are reported and discussed.


Assuntos
COVID-19/patologia , Doenças do Sistema Nervoso/virologia , Tropismo Viral/fisiologia , Doença de Alzheimer/virologia , COVID-19/virologia , Epilepsia/virologia , Flaviviridae/metabolismo , Síndrome de Guillain-Barré/virologia , Herpesviridae/metabolismo , Humanos , Esclerose Múltipla/virologia , Doenças do Sistema Nervoso/patologia , Orthomyxoviridae/metabolismo , Doença de Parkinson/virologia , Retroviridae/metabolismo , SARS-CoV-2/metabolismo
7.
Viruses ; 13(1)2021 Jan 08.
Artigo em Inglês | MEDLINE | ID: mdl-33429848

RESUMO

Sleeping Beauty (SB) is a transposon system that has been widely used as a genetic engineering tool. Central to the development of any transposon as a research tool is the ability to integrate a foreign piece of DNA into the cellular genome. Driven by the need for efficient transposon-based gene vector systems, extensive studies have largely elucidated the molecular actors and actions taking place during SB transposition. Close transposon relatives and other recombination enzymes, including retroviral integrases, have served as useful models to infer functional information relevant to SB. Recently obtained structural data on the SB transposase enable a direct insight into the workings of this enzyme. These efforts cumulatively allowed the development of novel variants of SB that offer advanced possibilities for genetic engineering due to their hyperactivity, integration deficiency, or targeting capacity. However, many aspects of the process of transposition remain poorly understood and require further investigation. We anticipate that continued investigations into the structure-function relationships of SB transposition will enable the development of new generations of transposition-based vector systems, thereby facilitating the use of SB in preclinical studies and clinical trials.


Assuntos
Elementos de DNA Transponíveis/genética , Engenharia Genética , Transposases/genética , Transposases/metabolismo , Animais , Humanos , Recombinação Genética , Retroviridae/genética , Retroviridae/metabolismo , Relação Estrutura-Atividade
8.
Viruses ; 12(10)2020 10 21.
Artigo em Inglês | MEDLINE | ID: mdl-33096825

RESUMO

Most cells can release extracellular vesicles (EVs), membrane vesicles containing various proteins, nucleic acids, enzymes, and signaling molecules. The exchange of EVs between cells facilitates intercellular communication, amplification of cellular responses, immune response modulation, and perhaps alterations in viral pathogenicity. EVs serve a dual role in inhibiting or enhancing viral infection and pathogenesis. This review examines the current literature on EVs to explore the complex role of EVs in the enhancement, inhibition, and potential use as a nanotherapeutic against clinically relevant viruses, focusing on neurotropic viruses: Zika virus (ZIKV) and human immunodeficiency virus (HIV). Overall, this review's scope will elaborate on EV-based mechanisms, which impact viral pathogenicity, facilitate viral spread, and modulate antiviral immune responses.


Assuntos
Vesículas Extracelulares/metabolismo , Viroses/metabolismo , Antivirais/farmacologia , Comunicação Celular/fisiologia , Coronavirus/metabolismo , Coronavirus/patogenicidade , Exossomos/metabolismo , HIV/metabolismo , HIV/patogenicidade , Infecções por HIV/metabolismo , Humanos , Retroviridae/metabolismo , Simplexvirus/metabolismo , Terapêutica/métodos , Viroses/tratamento farmacológico , Viroses/virologia , Zika virus/metabolismo , Zika virus/patogenicidade , Infecção por Zika virus/metabolismo
9.
Viruses ; 12(10)2020 10 20.
Artigo em Inglês | MEDLINE | ID: mdl-33092109

RESUMO

One of the most important steps in any viral lifecycle is the production of progeny virions. For retroviruses as well as other viruses, this step is a highly organized process that occurs with exquisite spatial and temporal specificity on the cellular plasma membrane. To facilitate this process, retroviruses encode short peptide motifs, or L domains, that hijack host factors to ensure completion of this critical step. One such cellular machinery targeted by viruses is known as the Endosomal Sorting Complex Required for Transport (ESCRTs). Typically responsible for vesicular trafficking within the cell, ESCRTs are co-opted by the retroviral Gag polyprotein to assist in viral particle assembly and release of infectious virions. This review in the Viruses Special Issue "The 11th International Retroviral Nucleocapsid and Assembly Symposium", details recent findings that shed light on the molecular details of how ESCRTs and the ESCRT adaptor protein ALIX, facilitate retroviral dissemination at sites of viral assembly.


Assuntos
Complexos Endossomais de Distribuição Requeridos para Transporte/metabolismo , Retroviridae , Montagem de Vírus/fisiologia , Liberação de Vírus/fisiologia , HIV-1/metabolismo , Nucleocapsídeo/metabolismo , Retroviridae/crescimento & desenvolvimento , Retroviridae/metabolismo , Ribonucleoproteínas/metabolismo , Produtos do Gene gag do Vírus da Imunodeficiência Humana/metabolismo
10.
J Vis Exp ; (159)2020 05 31.
Artigo em Inglês | MEDLINE | ID: mdl-32538902

RESUMO

As cell function is influenced by niche-specific factors in the cellular microenvironment, methods to dissect cell localization and migration can provide further insight on cell function. B-1a cells are a unique B cell subset in mice that produce protective natural IgM antibodies against oxidation-specific epitopes that arise during health and disease. B-1a cell IgM production differs depending on B-1a cell location, and therefore it becomes useful from a therapeutic standpoint to target B-1a localization to niches supportive of high antibody production. Here we describe a method to target B-1a cell migration to the bone marrow by retroviral-mediated overexpression of the C-X-C motif chemokine receptor 4 (CXCR4). Gene induction in primary murine B cells can be challenging and typically yields low transfection efficiencies of 10-20% depending on technique. Here we demonstrate that retroviral transduction of primary murine B-1a cells results in 30-40% transduction efficiency. This method utilizes adoptive cell transfer of transduced B-1a cells into B cell-deficient recipient mice so that donor B-1a cell migration and localization can be visualized. This protocol can be modified for other retroviral constructs and can be used in diverse functional assays post-adoptive transfer, including analysis of donor cell or host cell phenotype and function, or analysis of soluble factors secreted post B-1a cell transfer. The use of distinct donor and recipient mice differentiated by CD45.1 and CD45.2 allotype and the presence of a GFP reporter within the retroviral plasmid could also enable detection of donor cells in other, immune-sufficient mouse models containing endogenous B cell populations.


Assuntos
Transferência Adotiva , Subpopulações de Linfócitos B/imunologia , Células da Medula Óssea/imunologia , Movimento Celular , Receptores CXCR4/metabolismo , Retroviridae/metabolismo , Animais , Formação de Anticorpos , Linfócitos B/imunologia , Diferenciação Celular , Imunoglobulina M/imunologia , Antígenos Comuns de Leucócito , Camundongos , Receptores CXCR4/genética , Transdução de Sinais
11.
Biochim Biophys Acta Gene Regul Mech ; 1863(9): 194583, 2020 09.
Artigo em Inglês | MEDLINE | ID: mdl-32450258

RESUMO

Retroviruses are a unique family of RNA viruses that utilize a virally encoded reverse transcriptase (RT) to replicate their genomic RNA (gRNA) through a proviral DNA intermediate. The provirus is permanently integrated into the host cell chromosome and is expressed by the host cell transcription, RNA processing, and translation machinery. Retroviral messenger RNAs (mRNAs) entirely resemble a cellular mRNA as they have a 5'cap structure, 5'untranslated region (UTR), an open reading frame (ORF), 3'UTR, and a 3'poly(A) tail. The primary transcription product interacts with the cellular RNA processing machinery and is spliced, exported to the cytoplasm, and translated. However, a proportion of the pre-mRNA subverts typical RNA processing giving rise to the full-length RNA. In the cytoplasm, the full-length retroviral RNA fulfills a dual role acting as mRNA and as the gRNA. Simple retroviruses generate two pools of full-length RNA, one for each purpose. However, complex retroviruses have a single pool of full-length RNA, which is destined for translation or encapsidation. As for eukaryotic mRNAs, translational control of retroviral protein synthesis is mostly exerted at the step of initiation. Interestingly, some retroviral mRNAs, both simple and complex, use a dual mechanism to initiate protein synthesis, a cap-dependent initiation mechanism, or via internal initiation using an internal ribosome entry site (IRES). In this review, we describe and discuss data regarding the molecular mechanism driving the canonical cap-dependent and IRES-mediated translation initiation for retroviral mRNA, focusing the discussion mainly on the most studied retroviral mRNA, the HIV-1 mRNA.


Assuntos
Regulação Viral da Expressão Gênica , Iniciação Traducional da Cadeia Peptídica , Capuzes de RNA , Precursores de RNA/genética , Splicing de RNA , RNA Viral , Retroviridae/genética , Animais , Humanos , Sítios Internos de Entrada Ribossomal , Conformação de Ácido Nucleico , Precursores de RNA/química , Precursores de RNA/metabolismo , RNA Mensageiro/química , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Retroviridae/metabolismo
12.
J Virol ; 94(11)2020 05 18.
Artigo em Inglês | MEDLINE | ID: mdl-32188730

RESUMO

Koala retrovirus (KoRV) is of an interest to virologists due to its currently active endogenization into the koala (Phascolarctos cinereus) genome. Although KoRV has frequently been isolated in wild and captive koala populations, its pathogenesis and transmission remain to be fully characterized, and most previous research has concentrated on adult koalas rather than on joeys. Here, we characterized KoRV isolates obtained from a deceased male joey and its parents (animals reared in a Japanese zoo) to investigate KoRV transmission mode and pathogenesis. We sequenced the KoRV long terminal repeat (LTR) and envelope genes isolated from the joey and its parents and found KoRV-A and KoRV-C in genomic DNA from both the parents and the joey. Notably, both parents were also positive for KoRV-B, whereas the joey was KoRV-B negative, further confirming that KoRV-B is an exogenous strain. The KoRV LTR sequence of the joey was considerably closer to that of its sire than its dam. For further characterization, total KoRV, KoRV-A, KoRV-B, and KoRV-C proviral loads were quantified in peripheral blood mononuclear cells from the parents and in blood samples from the joey. Total KoRV, KoRV-A, and KoRV-C proviral loads were also quantified for different tissues (bone, liver, kidney, lung, spleen, heart, and muscle) from the joey, revealing differences suggestive of a distinct tissue tropism (highest total KoRV proviral load in the spleen and lowest in bone). The amount of KoRV-C in the parents was less than that in the joey. Our findings contribute to an improved understanding of KoRV pathogenesis and transmission mode and highlight useful areas for future research.IMPORTANCE KoRV is unique among retroviruses in that one strain (KoRV-A) is undergoing endogenization, whereas the other main subtype (KoRV-B) and another subtype (KoRV-C) are reportedly exogenous strains. Its transmission and pathogenesis are of interest in the study of retroviruses and are crucial for any conservation strategy geared toward koala health. This study provides new evidence on the modes of KoRV transmission from parent koalas to their joey. We found vertical transmission of KoRV-A, confirming its endogenization, but with closer conservation between the joey and its sire than its dam (previous reports on joeys are rare but have postulated dam-to-joey vertical transmission). This is also the first report of a KoRV-B-negative joey from KoRV-B-positive parents, contrasting with the few previous reports of 100% transmission of KoRV-B from dams to joeys. Thus, the results in this study give some novel insights for the transmission mode of KoRV.


Assuntos
Evolução Molecular , Phascolarctidae/virologia , Infecções por Retroviridae , Retroviridae , Sequências Repetidas Terminais , Animais , Feminino , Japão , Leucócitos Mononucleares/metabolismo , Leucócitos Mononucleares/virologia , Masculino , Retroviridae/genética , Retroviridae/metabolismo , Infecções por Retroviridae/genética , Infecções por Retroviridae/metabolismo , Infecções por Retroviridae/transmissão , Infecções por Retroviridae/veterinária
13.
Bull Exp Biol Med ; 168(4): 566-573, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-32157511

RESUMO

The use of induced pluripotent stem cells (IPSC) is a promising approach to the therapy of CNS diseases. The undeniable advantage of IPSC technology is the possibility of obtaining practically all types of somatic cells for autologous transplantation bypassing bioethical problems. The review presents integrative and non-integrative methods for obtaining IPSC and the ways of their in vitro and in vivo application for the study and treatment of neurological diseases.


Assuntos
Células-Tronco Pluripotentes Induzidas/transplante , Células-Tronco Neurais/transplante , Doenças Neurodegenerativas/terapia , Medicina de Precisão/métodos , Transplante de Células-Tronco/métodos , Animais , Diferenciação Celular , Vetores Genéticos/química , Vetores Genéticos/metabolismo , Humanos , Células-Tronco Pluripotentes Induzidas/citologia , Células-Tronco Pluripotentes Induzidas/metabolismo , Fatores de Transcrição Kruppel-Like/genética , Fatores de Transcrição Kruppel-Like/metabolismo , Lentivirus/genética , Lentivirus/metabolismo , Células-Tronco Neurais/citologia , Células-Tronco Neurais/metabolismo , Doenças Neurodegenerativas/genética , Doenças Neurodegenerativas/metabolismo , Doenças Neurodegenerativas/patologia , Fator 3 de Transcrição de Octâmero/genética , Fator 3 de Transcrição de Octâmero/metabolismo , Proteínas Proto-Oncogênicas c-myc/genética , Proteínas Proto-Oncogênicas c-myc/metabolismo , Retroviridae/genética , Retroviridae/metabolismo , Fatores de Transcrição SOXB1/genética , Fatores de Transcrição SOXB1/metabolismo , Transdução Genética/métodos
14.
Cell Death Dis ; 11(1): 5, 2020 01 02.
Artigo em Inglês | MEDLINE | ID: mdl-31919392

RESUMO

T cell homeostasis is critical for the proper function of the immune system. Following the sharp expansion upon pathogen infection, most T cells die in order to keep balance in the immune system, a process which is controlled by death receptors during the early phase and Bcl-2 proteins in the later phase. It is still highly debated whether the apoptosis of T cells is determined from the beginning, upon activation, or determined later during the contraction. MCL1, a Bcl-2 family member, plays a pivotal role in T cell survival. As a fast turnover protein, MCL1 levels are tightly regulated by the 26S proteasome-controlled protein degradation process. In searching for regulatory factors involved in the actions of MCL1 during T cell apoptosis, we found that ALG-2 was critical for MCL1 stability, a process mediated by a direct interaction between ALG-2 and Rpn3, a key component of the 26S proteasome. As a critical calcium sensor, ALG-2 regulated the activity of the 26S proteasome upon increases to cytosolic calcium levels following T cell activation, this consequently influenced the stability of MCL1 and accelerated the T cell "death" process, leading to T cell contraction and restoration of immune homeostasis. Our study provides support for the notion that T cells are destined for apoptosis after activation, and echoes the previous study about the function of ALG-2 in T cell death.


Assuntos
Proteínas Reguladoras de Apoptose/metabolismo , Apoptose/genética , Proteínas de Ligação ao Cálcio/metabolismo , Proteína de Sequência 1 de Leucemia de Células Mieloides/metabolismo , Complexo de Endopeptidases do Proteassoma/metabolismo , Linfócitos T/metabolismo , Proteínas Reguladoras de Apoptose/genética , Cálcio/metabolismo , Proteínas de Ligação ao Cálcio/genética , Linhagem Celular Tumoral , Cromatografia Líquida , Vetores Genéticos , Homeostase , Humanos , Proteína de Sequência 1 de Leucemia de Células Mieloides/genética , Complexo de Endopeptidases do Proteassoma/genética , Ligação Proteica , Proteólise , Retroviridae/genética , Retroviridae/metabolismo , Linfócitos T/imunologia , Espectrometria de Massas em Tandem
15.
Curr Issues Mol Biol ; 35: 1-16, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31422930

RESUMO

Small ubiquitin-like modifier (SUMO)ylation is a crucial post-translational modification that controls functions of a wide collection of proteins and biological processes. Hence, given its pleiotropic role, viruses have developed many approaches to usurp SUMO conjugation to exploit the cellular host environment for their own benefit. Consistently, cancer cells also frequently impact on SUMO to force cellular transformation, underlining the importance of SUMO in health and diseases. Therefore, after a brief introduction to the multistep SUMOylation pathway, in this review we will focus our attention on several examples of strategies adopted by oncogenic viruses to hijack SUMOylation in order to promote infection, persistence and malignant transformation of host cells.


Assuntos
Neoplasias/metabolismo , Neoplasias/virologia , Retroviridae/metabolismo , Proteínas Modificadoras Pequenas Relacionadas à Ubiquitina/metabolismo , Sumoilação , Cromatina/genética , Cromatina/metabolismo , Hepacivirus/genética , Hepacivirus/metabolismo , Hepacivirus/patogenicidade , Vírus da Hepatite B/genética , Vírus da Hepatite B/metabolismo , Vírus da Hepatite B/patogenicidade , Herpesvirus Humano 4/genética , Herpesvirus Humano 4/metabolismo , Herpesvirus Humano 4/patogenicidade , Herpesvirus Humano 8/genética , Herpesvirus Humano 8/metabolismo , Herpesvirus Humano 8/patogenicidade , Vírus Linfotrópico T Tipo 1 Humano/genética , Vírus Linfotrópico T Tipo 1 Humano/metabolismo , Vírus Linfotrópico T Tipo 1 Humano/patogenicidade , Humanos , Poliomavírus das Células de Merkel/genética , Poliomavírus das Células de Merkel/metabolismo , Poliomavírus das Células de Merkel/patogenicidade , Neoplasias/genética , Papillomaviridae/genética , Papillomaviridae/metabolismo , Papillomaviridae/patogenicidade , Retroviridae/genética , Retroviridae/crescimento & desenvolvimento , Retroviridae/patogenicidade , Proteínas Modificadoras Pequenas Relacionadas à Ubiquitina/genética , Ubiquitina-Proteína Ligases/metabolismo
16.
Methods Mol Biol ; 2097: 223-230, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-31776929

RESUMO

CRISPR-Cas9 technology allows for rapid, targeted genome editing at nearly any loci with limited off-target effects. Here, we describe a method for using retroviral transduction to deliver single-guide RNA to primary bone marrow-derived macrophages. This protocol allows for high-throughput reverse genetics assays in primary immune cells and is also compatible with retroviral systems for transgene expression.


Assuntos
Proteína 9 Associada à CRISPR/metabolismo , Sistemas CRISPR-Cas/genética , Marcação de Genes/métodos , Macrófagos/metabolismo , Animais , Células Cultivadas , Células HEK293 , Humanos , Camundongos , Retroviridae/metabolismo , Transdução Genética
17.
Nat Commun ; 10(1): 5652, 2019 12 11.
Artigo em Inglês | MEDLINE | ID: mdl-31827084

RESUMO

Transposable elements (TEs) have been proposed to play an important role in driving the expansion of gene regulatory networks during mammalian evolution, notably by contributing to the evolution and function of long non-coding RNAs (lncRNAs). XACT is a primate-specific TE-derived lncRNA that coats active X chromosomes in pluripotent cells and may contribute to species-specific regulation of X-chromosome inactivation. Here we explore how different families of TEs have contributed to shaping the XACT locus and coupling its expression to pluripotency. Through a combination of sequence analysis across primates, transcriptional interference, and genome editing, we identify a critical enhancer for the regulation of the XACT locus that evolved from an ancestral group of mammalian endogenous retroviruses (ERVs), prior to the emergence of XACT. This ERV was hijacked by younger hominoid-specific ERVs that gave rise to the promoter of XACT, thus wiring its expression to the pluripotency network. This work illustrates how retroviral-derived sequences may intervene in species-specific regulatory pathways.


Assuntos
Retrovirus Endógenos/genética , Elementos Facilitadores Genéticos , Primatas/virologia , RNA Longo não Codificante/genética , Retroviridae/genética , Animais , Elementos de DNA Transponíveis , Retrovirus Endógenos/metabolismo , Evolução Molecular , Regulação da Expressão Gênica , Redes Reguladoras de Genes , Interações Hospedeiro-Patógeno , Humanos , Filogenia , Primatas/classificação , Primatas/genética , RNA Longo não Codificante/metabolismo , Retroviridae/metabolismo , Especificidade da Espécie
18.
Viruses ; 11(9)2019 08 22.
Artigo em Inglês | MEDLINE | ID: mdl-31443391

RESUMO

Foamy viruses (FV) are retroviruses belonging to the Spumaretrovirinae subfamily. They are non-pathogenic viruses endemic in several mammalian hosts like non-human primates, felines, bovines, and equines. Retroviral DNA integration is a mandatory step and constitutes a prime target for antiretroviral therapy. This activity, conserved among retroviruses and long terminal repeat (LTR) retrotransposons, involves a viral nucleoprotein complex called intasome. In the last decade, a plethora of structural insights on retroviral DNA integration arose from the study of FV. Here, we review the biochemistry and the structural features of the FV integration apparatus and will also discuss the mechanism of action of strand transfer inhibitors.


Assuntos
Integrases , Spumavirus , Integração Viral , Animais , Antirretrovirais/química , Antirretrovirais/farmacologia , Domínio Catalítico , DNA Viral/química , Infecções por HIV/tratamento farmacológico , HIV-1/efeitos dos fármacos , Humanos , Inibidores de Integrase/química , Inibidores de Integrase/farmacologia , Integrases/química , Integrases/metabolismo , Modelos Moleculares , Nucleoproteínas/química , Nucleoproteínas/metabolismo , Retroviridae/genética , Retroviridae/metabolismo , Spumavirus/genética , Spumavirus/metabolismo , Sequências Repetidas Terminais
19.
RNA Biol ; 16(12): 1749-1763, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31469343

RESUMO

During replication of long terminal repeat (LTR)-retrotransposons, their proteins and genome (g) RNA assemble into virus-like particles (VLPs) that are not infectious but functionally related to retroviral virions. Both virions and VLPs contain gRNA in a dimeric form, but contrary to retroviruses, little is known about how gRNA dimerization and packaging occurs in LTR-retrotransposons. The LTR-retrotransposon Ty1 from Saccharomyces cerevisiae is an informative model for studying LTR-retrotransposon and retrovirus replication. Using structural, mutational and functional analyses, we explored dimerization of Ty1 genomic RNA. We provide direct evidence that interactions of self-complementary PAL1 and PAL2 palindromic sequences localized within the 5'UTR are essential for Ty1 gRNA dimer formation. Mutations disrupting PAL1-PAL2 complementarity restricted RNA dimerization in vitro and Ty1 mobility in vivo. Although dimer formation and mobility of these mutants was inhibited, our work suggests that Ty1 RNA can dimerize via alternative contact points. In contrast to previous studies, we cannot confirm a role for PAL3, tRNAiMet as well as recently proposed initial kissing-loop interactions in dimer formation. Our data also supports the critical role of Ty1 Gag in RNA dimerization. Mature Ty1 Gag binds in the proximity of sequences involved in RNA dimerization and tRNAiMet annealing, but the 5' pseudoknot in Ty1 RNA may constitute a preferred Gag-binding site. Taken together, these results expand our understanding of genome dimerization and packaging strategies utilized by LTR-retroelements.


Assuntos
RNA de Transferência/genética , RNA Viral/genética , Retroelementos , Retroviridae/genética , Saccharomyces cerevisiae/virologia , Regiões 5' não Traduzidas , Pareamento de Bases , Sequência de Bases , Dimerização , Modelos Moleculares , Mutação , Conformação de Ácido Nucleico , RNA de Transferência/química , RNA de Transferência/metabolismo , RNA Viral/química , RNA Viral/metabolismo , Retroviridae/metabolismo , Saccharomyces cerevisiae/genética , Vírion/genética , Vírion/metabolismo , Replicação Viral
20.
Sci Rep ; 9(1): 12416, 2019 08 27.
Artigo em Inglês | MEDLINE | ID: mdl-31455828

RESUMO

Koala retrovirus (KoRV) is in the process of endogenization into the koala (Phascolarctos cinereus) genome and is currently spreading through the Australian koala population. Understanding how the koala's immune system responds to KoRV infection is critical for developing an efficacious vaccine to protect koalas. To this end, we analyzed the antibody response of 235 wild koalas, sampled longitudinally over a four-year period, that harbored KoRV-A, and with or without KoRV-B. We found that the majority of the sampled koalas were able to make anti-KoRV antibodies, and that there was a linear increase in anti-KoRV IgG levels in koalas up to approximately seven years of age and then a gradual decrease thereafter. Koalas infected with both KoRV-A and KoRV-B were found to have slightly higher anti-KoRV IgG titers than koalas with KoRV-A alone and there was an inverse relationship between anti-KoRV IgG levels and circulating KoRV viral load. Finally, we identified distinct epitopes on the KoRV envelope protein that were recognized by antibodies. Together, these findings provide insight into the koala's immune response to KoRV and may be useful in the development of a therapeutic KoRV vaccine.


Assuntos
Anticorpos Antivirais/sangue , Formação de Anticorpos , Imunoglobulina G/sangue , Phascolarctidae , Retroviridae/metabolismo , Animais , Feminino , Masculino , Phascolarctidae/sangue , Phascolarctidae/virologia , Infecções por Retroviridae/sangue , Infecções por Retroviridae/veterinária , Infecções por Retroviridae/virologia
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