RESUMEN
Human T cell leukemia virus type 1 (HTLV-1), also known as human T lymphotropic virus type 1, was the first exogenous human retrovirus discovered. Unlike the distantly related lentivirus HIV-1, HTLV-1 causes disease in only 5-10% of infected people, depending on their ethnic origin. But whereas HIV-1 infection and the consequent diseases can be efficiently contained in most cases by antiretroviral drug treatment, there is no satisfactory treatment for the malignant or inflammatory diseases caused by HTLV-1. The purpose of the present article is to review recent advances in the understanding of the mechanisms by which the virus persists in vivo and causes disabling or fatal diseases.
Asunto(s)
Infecciones por HTLV-I/inmunología , Infecciones por HTLV-I/virología , Virus Linfotrópico T Tipo 1 Humano/fisiología , Animales , Susceptibilidad a Enfermedades , Infecciones por HTLV-I/complicaciones , Infecciones por HTLV-I/epidemiología , Interacciones Huésped-Patógeno/inmunología , Humanos , Inmunidad , Inmunidad Celular , Interferón Tipo I/metabolismo , Leucemia-Linfoma de Células T del Adulto/etiología , Leucemia-Linfoma de Células T del Adulto/patología , Subgrupos de Linfocitos T/inmunología , Subgrupos de Linfocitos T/metabolismo , Proteínas Reguladoras y Accesorias Virales/genética , Proteínas Reguladoras y Accesorias Virales/inmunología , Proteínas Reguladoras y Accesorias Virales/metabolismo , Latencia del Virus/inmunologíaRESUMEN
A typical HTLV-1-infected individual carries >104 different HTLV-1-infected T cell clones, each with a single-copy provirus integrated in a unique genomic site. We previously showed that the HTLV-1 provirus causes aberrant transcription in the flanking host genome and, by binding the chromatin architectural protein CTCF, forms abnormal chromatin loops with the host genome. However, it remained unknown whether these effects were exerted simply by the presence of the provirus or were induced by its transcription. To answer this question, we sorted HTLV-1-infected T-cell clones into cells positive or negative for proviral plus-strand expression, and then quantified host and provirus transcription using RNA-seq, and chromatin looping using quantitative chromosome conformation capture (q4C), in each cell population. We found that proviral plus-strand transcription induces aberrant transcription and splicing in the flanking genome but suppresses aberrant chromatin loop formation with the nearby host chromatin. Reducing provirus-induced host transcription with an inhibitor of transcriptional elongation allows recovery of chromatin loops in the plus-strand-expressing population. We conclude that aberrant host transcription induced by proviral expression causes temporary, reversible disruption of chromatin looping in the vicinity of the provirus.
Asunto(s)
Virus Linfotrópico T Tipo 1 Humano , Virus Linfotrópico T Tipo 1 Humano/genética , Virus Linfotrópico T Tipo 1 Humano/metabolismo , Cromatina/genética , Cromatina/metabolismo , Provirus/genética , Linfocitos TRESUMEN
Infections by Human T cell Leukaemia Virus type 1 (HTLV-1) persist for the lifetime of the host by integrating into the genome of CD4+ T cells. Proviral gene expression is essential for proviral survival and the maintenance of the proviral load, through the pro-proliferative changes it induces in infected cells. Despite their role in HTLV-1 infection and a persistent cytotoxic T lymphocyte response raised against the virus, proviral transcripts from the sense-strand are rarely detected in fresh cells extracted from the peripheral blood, and have recently been found to be expressed intermittently by a small subset of cells at a given time. Ex vivo culture of infected cells prompts synchronised proviral expression in infected cells from peripheral blood, allowing the study of factors involved in reactivation in primary cells. Here, we used bulk RNA-seq to examine the host transcriptome over six days in vitro, following proviral reactivation in primary peripheral CD4+ T cells isolated from subjects with non-malignant HTLV-1 infection. Infected cells displayed a conserved response to reactivation, characterised by discrete stages of gene expression, cell division and subsequently horizontal transmission of the virus. We observed widespread changes in Polycomb gene expression following reactivation, including an increase in PRC2 transcript levels and diverse changes in the expression of PRC1 components. We hypothesize that these transcriptional changes constitute a negative feedback loop that maintains proviral latency by re-deposition of H2AK119ub1 following the end of proviral expression. Using RNAi, we found that certain deubiquitinases, BAP1, USP14 and OTUD5 each promote proviral transcription. These data demonstrate the detailed trajectory of HTLV-1 proviral reactivation in primary HTLV-1-carrier lymphocytes and the impact on the host cell.
Asunto(s)
Infecciones por HTLV-I , Virus Linfotrópico T Tipo 1 Humano , Humanos , Virus Linfotrópico T Tipo 1 Humano/fisiología , Provirus/genética , Transcriptoma , Linfocitos T CD4-Positivos , Carga Viral , Ubiquitina Tiolesterasa/metabolismoRESUMEN
Human T-cell leukemia virus type 1 (HTLV-1), also known as human T-lymphotropic virus type 1, causes the aggressive malignancy known as adult T-cell leukemia/lymphoma (ATL) in 5% of infected people and a chronic progressive inflammatory disease of the central nervous system, HTLV-1-associated myelopathy, in â¼0.3% to 4% of them, varying between regions where it is endemic. Reliable treatments are lacking for both conditions, although there have been promising recent advances in the prevention and treatment of ATL. Because ATL typically develops after several decades of infection, it is necessary to understand how the virus persists in the host despite a strong immune response, and how this persistence results in oncogenesis.
Asunto(s)
Virus Linfotrópico T Tipo 1 Humano , Leucemia-Linfoma de Células T del Adulto , Linfoma , Adulto , Humanos , Carcinogénesis , Transformación Celular Neoplásica , Linfoma/complicacionesRESUMEN
Expression of the transcriptional transactivator protein Tax, encoded on the proviral plus-strand of human T-cell leukaemia virus type 1 (HTLV-1), is crucial for the replication of the virus, but Tax-expressing cells are rarely detected in fresh blood ex vivo. The dynamics and consequences of the proviral plus-strand transcriptional burst remain insufficiently characterised. We combined time-lapse live-cell imaging, single-cell tracking and mathematical modelling to study the dynamics of Tax expression at single-cell resolution in two naturally-infected, non-malignant T-cell clones transduced with a short-lived enhanced green fluorescent protein (d2EGFP) Tax reporter system. Five different patterns of Tax expression were observed during the 30-hour observation period; the distribution of these patterns differed between the two clones. The mean duration of Tax expression in the two clones was 94 and 417 hours respectively, estimated from mathematical modelling of the experimental data. Tax expression was associated with a transient slowing in cell-cycle progression and proliferation, increased apoptosis, and enhanced activation of the DNA damage response pathways. Longer-term follow-up (14 days) revealed an increase in the proportion of proliferating cells and a decrease in the fraction of apoptotic cells as the cells ceased Tax expression, resulting in a greater net expansion of the initially Tax-positive population. Time-lapse live-cell imaging showed enhanced cell-to-cell adhesion among Tax-expressing cells, and decreased cell motility of Tax-expressing cells at the single-cell level. The results demonstrate the within-clone and between-clone heterogeneity in the dynamics and patterns of HTLV-1 plus-strand transcriptional bursts and the balance of positive and negative consequences of the burst for the host cell.
Asunto(s)
Virus Linfotrópico T Tipo 1 Humano , Provirus , Humanos , Provirus/genética , Virus Linfotrópico T Tipo 1 Humano/genéticaRESUMEN
The human T-cell leukemia virus type 1 (HTLV-1) transactivator protein Tax has pleiotropic functions in the host cell affecting cell-cycle regulation, DNA damage response pathways and apoptosis. These actions of Tax have been implicated in the persistence and pathogenesis of HTLV-1-infected cells. It is now known that tax expression occurs in transcriptional bursts of the proviral plus-strand, but the effects of the burst on host transcription are not fully understood. We carried out RNA sequencing of two naturally-infected T-cell clones transduced with a Tax-responsive Timer protein, which undergoes a time-dependent shift in fluorescence emission, to study transcriptional changes during successive phases of the HTLV-1 plus-strand burst. We found that the transcriptional regulation of genes involved in the NF-κB pathway, cell-cycle regulation, DNA damage response and apoptosis inhibition were immediate effects accompanying the plus-strand burst, and are limited to the duration of the burst. The results distinguish between the immediate and delayed effects of HTLV-1 reactivation on host transcription, and between clone-specific effects and those observed in both clones. The major transcriptional changes in the infected host T-cells observed here, including NF-κB, are transient, suggesting that these pathways are not persistently activated at high levels in HTLV-1-infected cells. The two clones diverged strongly in their expression of genes regulating the cell cycle. Up-regulation of senescence markers was a delayed effect of the proviral plus-strand burst and the up-regulation of some pro-apoptotic genes outlasted the burst. We found that activation of the aryl hydrocarbon receptor (AhR) pathway enhanced and prolonged the proviral burst, but did not increase the rate of reactivation. Our results also suggest that sustained plus-strand expression is detrimental to the survival of infected cells.
Asunto(s)
Virus Linfotrópico T Tipo 1 Humano , Productos del Gen tax/genética , Productos del Gen tax/metabolismo , Virus Linfotrópico T Tipo 1 Humano/fisiología , Humanos , FN-kappa B/metabolismo , Provirus , Activación TranscripcionalRESUMEN
BACKGROUND: Coinfection with human immunodeficiency virus type 1 (HIV-1) and human T-cell leukemia virus type 1 (HTLV-1) diminishes the value of the CD4+ T-cell count in diagnosing AIDS, and increases the rate of HTLV-1-associated myelopathy. It remains elusive how HIV-1/HTLV-1 coinfection is related to such characteristics. We investigated the mutual effect of HIV-1/HTLV-1 coinfection on their integration sites (ISs) and clonal expansion. METHODS: We extracted DNA from longitudinal peripheral blood samples from 7 HIV-1/HTLV-1 coinfected, and 12 HIV-1 and 13 HTLV-1 monoinfected individuals. Proviral loads (PVL) were quantified using real-time polymerase chain reaction (PCR). Viral ISs and clonality were quantified by ligation-mediated PCR followed by high-throughput sequencing. RESULTS: PVL of both HIV-1 and HTLV-1 in coinfected individuals was significantly higher than that of the respective virus in monoinfected individuals. The degree of oligoclonality of both HIV-1- and HTLV-1-infected cells in coinfected individuals was also greater than in monoinfected subjects. ISs of HIV-1 in cases of coinfection were more frequently located in intergenic regions and transcriptionally silent regions, compared with HIV-1 monoinfected individuals. CONCLUSIONS: HIV-1/HTLV-1 coinfection makes an impact on the distribution of viral ISs and clonality of virus-infected cells and thus may alter the risks of both HTLV-1- and HIV-1-associated disease.
Asunto(s)
Coinfección , Infecciones por VIH/complicaciones , VIH-1 , Infecciones por HTLV-I/complicaciones , Virus Linfotrópico T Tipo 1 Humano , Paraparesia Espástica Tropical/epidemiología , Recuento de Linfocito CD4 , Infecciones por VIH/epidemiología , VIH-1/genética , VIH-1/aislamiento & purificación , Infecciones por HTLV-I/epidemiología , Secuenciación de Nucleótidos de Alto Rendimiento , Virus Linfotrópico T Tipo 1 Humano/genética , Virus Linfotrópico T Tipo 1 Humano/aislamiento & purificación , Humanos , Paraparesia Espástica Tropical/diagnóstico , Provirus/genética , Reacción en Cadena en Tiempo Real de la PolimerasaRESUMEN
Adult T-cell leukemia/lymphoma (ATL) is an aggressive hematological malignancy caused by human T-cell leukemia virus type-1 (HTLV-1). ATL is preceded by decades of chronic HTLV-1 infection, and the tumors carry both somatic mutations and proviral DNA integrated into the tumor genome. In order to gain insight into the oncogenic process, we used targeted sequencing to track the evolution of the malignant clone in 6 individuals, 2 to 10 years before the diagnosis of ATL. Clones of premalignant HTLV-1-infected cells bearing known driver mutations were detected in the blood up to 10 years before individuals developed acute and lymphoma subtype ATL. Six months before diagnosis, the total number and variant allele fraction of mutations increased in the blood. Peripheral blood mononuclear cells from premalignant cases (1 year prediagnosis) had significantly higher mutational burden in genes frequently mutated in ATL than did high-risk, age-matched HTLV-1 carriers who remained ATL-free after a median of 10 years of follow-up. These data show that HTLV-1-infected T-cell clones carrying key oncogenic driver mutations can be detected in cases of ATL years before the onset of symptoms. Early detection of such mutations may enable earlier and more effective intervention to prevent the development of ATL.
Asunto(s)
Células Clonales/patología , Evolución Molecular , Infecciones por HTLV-I/complicaciones , Virus Linfotrópico T Tipo 1 Humano/aislamiento & purificación , Leucemia-Linfoma de Células T del Adulto/diagnóstico , Leucocitos Mononucleares/patología , Linfocitos T/patología , Células Clonales/virología , Humanos , Leucemia-Linfoma de Células T del Adulto/epidemiología , Leucemia-Linfoma de Células T del Adulto/virología , Leucocitos Mononucleares/virología , Estudios Longitudinales , Linfocitos T/virología , Reino Unido/epidemiologíaRESUMEN
The human T cell leukemia virus HTLV-1 establishes a persistent infection in vivo in which the viral sense-strand transcription is usually silent at a given time in each cell. However, cellular stress responses trigger the reactivation of HTLV-1, enabling the virus to transmit to a new host cell. Using single-molecule RNA FISH, we measured the kinetics of the HTLV-1 transcriptional reactivation in peripheral blood mononuclear cells (PBMCs) isolated from HTLV-1+ individuals. The abundance of the HTLV-1 sense and antisense transcripts was quantified hourly during incubation of the HTLV-1-infected PBMCs ex vivo. We found that, in each cell, the sense-strand transcription occurs in two distinct phases: the initial low-rate transcription is followed by a phase of rapid transcription. The onset of transcription peaked between 1 and 3 hours after the start of in vitro incubation. The variance in the transcription intensity was similar in polyclonal HTLV-1+ PBMCs (with tens of thousands of distinct provirus insertion sites), and in samples with a single dominant HTLV-1+ clone. A stochastic simulation model was developed to estimate the parameters of HTLV-1 proviral transcription kinetics. In PBMCs from a leukemic subject with one dominant T-cell clone, the model indicated that the average duration of HTLV-1 sense-strand activation by Tax (i.e. the rapid transcription) was less than one hour. HTLV-1 antisense transcription was stable during reactivation of the sense-strand. The antisense transcript HBZ was produced at an average rate of ~0.1 molecules per hour per HTLV-1+ cell; however, between 20% and 70% of HTLV-1-infected cells were HBZ-negative at a given time, the percentage depending on the individual subject. HTLV-1-infected cells are exposed to a range of stresses when they are drawn from the host, which initiate the viral reactivation. We conclude that whereas antisense-strand transcription is stable throughout the stress response, the HTLV-1 sense-strand reactivation is highly heterogeneous and occurs in short, self-terminating bursts.
Asunto(s)
Infecciones por HTLV-I/virología , Virus Linfotrópico T Tipo 1 Humano/fisiología , Leucocitos Mononucleares/virología , Análisis de la Célula Individual/métodos , Proteínas Virales/genética , Activación Viral/genética , Latencia del Virus/genética , Células Cultivadas , Regulación Viral de la Expresión Génica , Infecciones por HTLV-I/genética , Humanos , Hibridación Fluorescente in Situ , Cinética , Procesos Estocásticos , Replicación ViralRESUMEN
Human T-lymphotropic virus type-1 (HTLV-1) persists within hosts via infectious spread (de novo infection) and mitotic spread (infected cell proliferation), creating a population structure of multiple clones (infected cell populations with identical genomic proviral integration sites). The relative contributions of infectious and mitotic spread to HTLV-1 persistence are unknown, and will determine the efficacy of different approaches to treatment. The prevailing view is that infectious spread is negligible in HTLV-1 persistence beyond early infection. However, in light of recent high-throughput data on the abundance of HTLV-1 clones, and recent estimates of HTLV-1 clonal diversity that are substantially higher than previously thought (typically between 104 and 105 HTLV-1+ T cell clones in the body of an asymptomatic carrier or patient with HTLV-1-associated myelopathy/tropical spastic paraparesis), ongoing infectious spread during chronic infection remains possible. We estimate the ratio of infectious to mitotic spread using a hybrid model of deterministic and stochastic processes, fitted to previously published HTLV-1 clonal diversity estimates. We investigate the robustness of our estimates using three alternative estimators. We find that, contrary to previous belief, infectious spread persists during chronic infection, even after HTLV-1 proviral load has reached its set point, and we estimate that between 100 and 200 new HTLV-1 clones are created and killed every day. We find broad agreement between all estimators. The risk of HTLV-1-associated malignancy and inflammatory disease is strongly correlated with proviral load, which in turn is correlated with the number of HTLV-1-infected clones, which are created by de novo infection. Our results therefore imply that suppression of de novo infection may reduce the risk of malignant transformation.
Asunto(s)
Infecciones por HTLV-I , Interacciones Huésped-Patógeno , Virus Linfotrópico T Tipo 1 Humano , Linfocitos T CD4-Positivos/virología , Infecciones por HTLV-I/fisiopatología , Infecciones por HTLV-I/virología , Interacciones Huésped-Patógeno/genética , Interacciones Huésped-Patógeno/fisiología , Virus Linfotrópico T Tipo 1 Humano/clasificación , Virus Linfotrópico T Tipo 1 Humano/genética , Virus Linfotrópico T Tipo 1 Humano/patogenicidad , Humanos , Mitosis/genética , Mitosis/fisiología , Modelos Biológicos , Provirus/genética , Provirus/patogenicidad , Carga Viral/genética , Integración Viral/genéticaRESUMEN
BACKGROUND: Adult T-cell leukemia-lymphoma (ATL) is an aggressive mature lymphoid proliferation associated with poor prognosis. Standard of care includes chemotherapy and/or the combination of zidovudine and interferon-alpha. However, most patients experience relapse less than 6 months after diagnosis. Allogeneic stem cell transplantation is the only curative treatment, but is only feasible in a minority of cases. We previously showed in a mouse model that Arsenic trioxide (As2O3) targets ATL leukemia initiating cells. RESULTS: As2O3 consolidation was given in 9 patients with ATL (lymphoma n = 4; acute n = 2; and indolent n = 3), who were in complete (n = 4) and partial (n = 3) remission, in stable (n = 1) and in progressive (n = 1) disease. Patients received up to 8 weeks of As2O3 at the dose of 0.15 mg/kg/day intravenously in combination with zidovudine and interferon-alpha. One patient in progression died rapidly. Of the remaining eight patients, three with indolent ATL subtype showed overall survivals of 48, 53 and 97 months, and duration of response to As2O3 of 22, 25 and 73 months. The other 5 patients with aggressive ATL subtype had median OS of 36 months and a median duration of response of 10 months. Side effects were mostly hematological and cutaneous (one grade 3) and reversible with dose reduction of AZT/IFN and/or As2O3 discontinuation. The virus integration analysis revealed the regression of the predominant malignant clone in one patient with a chronic subtype. CONCLUSION: These results suggest that consolidation with As2O3 could be an option for patients with ATL in response after induction therapy and who are not eligible for allogeneic stem cell transplantation.
Asunto(s)
Trióxido de Arsénico/uso terapéutico , Leucemia-Linfoma de Células T del Adulto/tratamiento farmacológico , Administración Intravenosa , Adulto , Trióxido de Arsénico/administración & dosificación , Quimioterapia Combinada , Femenino , Virus Linfotrópico T Tipo 1 Humano/efectos de los fármacos , Humanos , Interferón-alfa/uso terapéutico , Estudios Longitudinales , Masculino , Persona de Mediana Edad , Estudios Retrospectivos , Zidovudina/uso terapéuticoRESUMEN
The prevalence of human T-cell lymphotropic virus type 1 (HTLV-1) and hepatitis B virus (HBV) coinfection is high in certain Indigenous Australian populations, but its impact on HTLV-1 has not been described. We compared 2 groups of Indigenous adults infected with HTLV-1, either alone or coinfected with HBV. The 2 groups had a similar HTLV-1 proviral load, but there was a significant increase in clonal expansion of HTLV-1-infected lymphocytes in coinfected asymptomatic individuals. The degree of clonal expansion was correlated with the titer of HBV surface antigen. We conclude that HTLV-1/HBV coinfection may predispose to HTLV-1-associated malignant disease.
Asunto(s)
Coinfección/virología , Infecciones por HTLV-I/complicaciones , Infecciones por HTLV-I/virología , Hepatitis B/complicaciones , Virus Linfotrópico T Tipo 1 Humano/clasificación , Virus Linfotrópico T Tipo 1 Humano/aislamiento & purificación , Adulto , Anciano , Anciano de 80 o más Años , Australia , Femenino , Humanos , Linfocitos/virología , Masculino , Persona de Mediana Edad , Grupos de Población , Provirus/genética , Provirus/aislamiento & purificación , Carga ViralRESUMEN
HIV-1 integrates more frequently into transcribed genes, however the biological significance of HIV-1 integration targeting has remained elusive. Using a selective high-throughput chemical screen, we discovered that the cardiac glycoside digoxin inhibits wild-type HIV-1 infection more potently than HIV-1 bearing a single point mutation (N74D) in the capsid protein. We confirmed that digoxin repressed viral gene expression by targeting the cellular Na+/K+ ATPase, but this did not explain its selectivity. Parallel RNAseq and integration mapping in infected cells demonstrated that digoxin inhibited expression of genes involved in T-cell activation and cell metabolism. Analysis of >400,000 unique integration sites showed that WT virus integrated more frequently than N74D mutant within or near genes susceptible to repression by digoxin and involved in T-cell activation and cell metabolism. Two main gene networks down-regulated by the drug were CD40L and CD38. Blocking CD40L by neutralizing antibodies selectively inhibited WT virus infection, phenocopying digoxin. Thus the selectivity of digoxin depends on a combination of integration targeting and repression of specific gene networks. The drug unmasked a functional connection between HIV-1 integration and T-cell activation. Our results suggest that HIV-1 evolved integration site selection to couple its early gene expression with the status of target CD4+ T-cells, which may affect latency and viral reactivation.
Asunto(s)
Fármacos Anti-VIH/farmacología , Linfocitos T CD4-Positivos/inmunología , Digoxina/farmacología , Infecciones por VIH/inmunología , Infecciones por VIH/virología , VIH-1/fisiología , Integración Viral/efectos de los fármacos , Linfocitos T CD4-Positivos/efectos de los fármacos , Células Cultivadas , Infecciones por VIH/tratamiento farmacológico , VIH-1/efectos de los fármacos , VIH-1/genética , Humanos , Activación de Linfocitos/efectos de los fármacos , Latencia del Virus/efectos de los fármacosRESUMEN
Human T-lymphotropic virus type 1 (HTLV-1) is a retrovirus that causes malignant and inflammatory diseases in â¼10% of infected people. A typical host has between 10(4) and 10(5) clones of HTLV-1-infected T lymphocytes, each clone distinguished by the genomic integration site of the single-copy HTLV-1 provirus. The HTLV-1 bZIP (HBZ) factor gene is constitutively expressed from the minus strand of the provirus, whereas plus-strand expression, required for viral propagation to uninfected cells, is suppressed or intermittent in vivo, allowing escape from host immune surveillance. It remains unknown what regulates this pattern of proviral transcription and latency. Here, we show that CTCF, a key regulator of chromatin structure and function, binds to the provirus at a sharp border in epigenetic modifications in the pX region of the HTLV-1 provirus in T cells naturally infected with HTLV-1. CTCF is a zinc-finger protein that binds to an insulator region in genomic DNA and plays a fundamental role in controlling higher order chromatin structure and gene expression in vertebrate cells. We show that CTCF bound to HTLV-1 acts as an enhancer blocker, regulates HTLV-1 mRNA splicing, and forms long-distance interactions with flanking host chromatin. CTCF-binding sites (CTCF-BSs) have been propagated throughout the genome by transposons in certain primate lineages, but CTCF binding has not previously been described in present-day exogenous retroviruses. The presence of an ectopic CTCF-BS introduced by the retrovirus in tens of thousands of genomic locations has the potential to cause widespread abnormalities in host cell chromatin structure and gene expression.
Asunto(s)
Epigénesis Genética , Genoma Humano , Infecciones por HTLV-I/genética , Virus Linfotrópico T Tipo 1 Humano/genética , Mutagénesis Insercional/genética , Provirus/genética , Proteínas Represoras/metabolismo , Factores de Transcripción/genética , Proteínas Reguladoras y Accesorias Virales/genética , Integración Viral/genética , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/biosíntesis , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/genética , Sitios de Unión , Factor de Unión a CCCTC , Linfocitos T CD4-Positivos/virología , Cromatina/ultraestructura , Inmunoprecipitación de Cromatina , Secuencia de Consenso , ADN/genética , ADN/metabolismo , Metilación de ADN , ADN Viral/genética , ADN Viral/metabolismo , Regulación Viral de la Expresión Génica , Infecciones por HTLV-I/virología , Código de Histonas , Humanos , Unión Proteica , Proteínas de los Retroviridae/biosíntesis , Proteínas de los Retroviridae/genética , Transcripción GenéticaRESUMEN
Human T cell lymphotropic virus-1 (HTLV-1) primarily infects CD4+ T cells, causing inflammatory disorders or a T cell malignancy in 5% to 10% of carriers. The cytotoxic T lymphocyte (CTL) response is a key factor that controls the viral load and thus the risk of disease. The ability to detect the viral protein Tax in primary cells has made it possible to estimate the rate at which Tax-expressing infected cells are eliminated by CTLs in persistently infected people. However, most HTLV-1-infected cells are Tax-at a given time, and their immunophenotype is poorly defined. Here, we aimed to identify a cell-surface molecule expressed by both Tax+ and Tax-HTLV-1-infected cells and use it to analyse the CTL response in fresh peripheral blood mononuclear cells. Cell adhesion molecule 1 (CADM1/TSLC1) was the best single marker of HTLV-1 infection, identifying HTLV-1-infected cells with greater sensitivity and specificity than CD25, CCR4 or ICAM-1. CADM1+CD4+ T cells carried a median of 65% of proviral copies in peripheral blood. In a cohort of 23 individuals, we quantified the rate of CTL-mediated killing of Tax+ and Tax-CADM1+ cells. We show that CADM1 expression is associated with enhanced susceptibility of infected cells to CTL lysis: despite the immunodominance of Tax in the CTL response, Tax+CADM1- cells were inefficiently lysed by CTLs. Upregulation of the CADM1 ligand CRTAM on CD8+ T cells correlated with efficient lysis of infected cells. Tax-CADM1+ cells were lysed at a very low rate by autologous CTLs, however, were efficiently killed when loaded with exogenous peptide antigen. High expression of CADM1 on most HTLV-1-infected cells in the face of enhanced CTL counterselection implies that CADM1 confers a strong benefit on the virus.
Asunto(s)
Moléculas de Adhesión Celular/inmunología , Citotoxicidad Inmunológica/inmunología , Infecciones por HTLV-I/inmunología , Inmunoglobulinas/inmunología , Molécula 1 de Adhesión Celular , Citometría de Flujo , Productos del Gen tax/metabolismo , Virus Linfotrópico T Tipo 1 Humano/inmunología , Humanos , Reacción en Cadena en Tiempo Real de la Polimerasa , Linfocitos T Citotóxicos/inmunologíaRESUMEN
There is growing evidence that CD8+ cytotoxic T lymphocyte (CTL) responses can contribute to long-term remission of many malignancies. The etiological agent of adult T-cell leukemia/lymphoma (ATL), human T lymphotropic virus type-1 (HTLV-1), contains highly immunogenic CTL epitopes, but ATL patients typically have low frequencies of cytokine-producing HTLV-1-specific CD8+ cells in the circulation. It remains unclear whether patients with ATL possess CTLs that can kill the malignant HTLV-1 infected clone. Here we used flow cytometric staining of TCRVß and cell adhesion molecule-1 (CADM1) to identify monoclonal populations of HTLV-1-infected T cells in the peripheral blood of patients with ATL. Thus, we quantified the rate of CD8+-mediated killing of the putative malignant clone in ex vivo blood samples. We observed that CD8+ cells from ATL patients were unable to lyse autologous ATL clones when tested directly ex vivo. However, short in vitro culture restored the ability of CD8+ cells to kill ex vivo ATL clones in some donors. The capacity of CD8+ cells to lyse HTLV-1 infected cells which expressed the viral sense strand gene products was significantly enhanced after in vitro culture, and donors with an ATL clone that expressed the HTLV-1 Tax gene were most likely to make a detectable lytic CD8+ response to the ATL cells. We conclude that some patients with ATL possess functional tumour-specific CTLs which could be exploited to contribute to control of the disease.
Asunto(s)
Citotoxicidad Inmunológica/inmunología , Leucemia-Linfoma de Células T del Adulto/inmunología , Receptores de Antígenos de Linfocitos T alfa-beta/inmunología , Linfocitos T Citotóxicos/inmunología , Adulto , Anciano , Femenino , Citometría de Flujo , Productos del Gen tax/inmunología , Humanos , Masculino , Persona de Mediana EdadRESUMEN
BACKGROUND: Human T-lymphotropic virus type 1 (HTLV-1) infects an estimated 10 million persons globally with transmission resulting in lifelong infection. Disease, linked to high proviral load, occurs in a minority. In established infection HTLV-1 replicates through infectious spread and clonal expansion of infected lymphocytes. Little is known about acute HTLV-1 infection. The kinetics of early HTLV-1 infection, following transplantation-acquired infection in three recipients from one HTLV-1 infected donor, is reported. The recipients were treated with two HTLV-1 enzyme inhibitors 3 weeks post exposure following the detection of HTLV-1 provirus at low level in each recipient. HTLV-1 infection was serially monitored by serology, quantification of proviral load and HTLV-1 2LTR DNA circles and by HTLV-1 unique integration site analysis. RESULTS: HTLV-1 antibodies were first detected 16-39 days post-transplantation. HTLV-1 provirus was detected by PCR on day 16-23 and increased by 2-3 log by day 38-45 with a peak proviral doubling time of 1.4 days, after which steady state was reached. The rapid proviral load expansion was associated with high frequency of HTLV-1 2LTR DNA circles. The number of HTLV-1 unique integration sites was high compared with established HTLV-1 infection. Clonal expansion of infected cells was detected as early as day 37 with high initial oligoclonality index, consistent with early mitotic proliferation. CONCLUSIONS: In recipients infected through organ transplantation HTLV-1 disseminated rapidly despite early anti-HTLV-1 treatment. Proviral load set point was reached within 6 weeks. Seroconversion was not delayed. Unique integration site analysis and HTLV-1 2LTR DNA circles indicated early clonal expansion and high rate of infectious spread.
Asunto(s)
Infecciones por HTLV-I/patología , Infecciones por HTLV-I/virología , Virus Linfotrópico T Tipo 1 Humano/aislamiento & purificación , Provirus/aislamiento & purificación , Receptores de Trasplantes , Trasplante/efectos adversos , Carga Viral , Anticuerpos Antivirales/sangre , Antivirales/uso terapéutico , ADN Viral/análisis , Virus Linfotrópico T Tipo 1 Humano/inmunología , Humanos , Reacción en Cadena de la Polimerasa , Factores de TiempoRESUMEN
Human T-lymphotropic virus type 1 (HTLV-1) and type 2 (HTLV-2) both cause lifelong persistent infections, but differ in their clinical outcomes. HTLV-1 infection causes a chronic or acute T-lymphocytic malignancy in up to 5% of infected individuals whereas HTLV-2 has not been unequivocally linked to a T-cell malignancy. Virus-driven clonal proliferation of infected cells both in vitro and in vivo has been demonstrated in HTLV-1 infection. However, T-cell clonality in HTLV-2 infection has not been rigorously characterized. In this study we used a high-throughput approach in conjunction with flow cytometric sorting to identify and quantify HTLV-2-infected T-cell clones in 28 individuals with natural infection. We show that while genome-wide integration site preferences in vivo were similar to those found in HTLV-1 infection, expansion of HTLV-2-infected clones did not demonstrate the same significant association with the genomic environment of the integrated provirus. The proviral load in HTLV-2 is almost confined to CD8+ T-cells and is composed of a small number of often highly expanded clones. The HTLV-2 load correlated significantly with the degree of dispersion of the clone frequency distribution, which was highly stable over â¼8 years. These results suggest that there are significant differences in the selection forces that control the clonal expansion of virus-infected cells in HTLV-1 and HTLV-2 infection. In addition, our data demonstrate that strong virus-driven proliferation per se does not predispose to malignant transformation in oncoretroviral infections.