RESUMEN
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 PolimerasaAsunto(s)
Anticuerpos Monoclonales Humanizados/uso terapéutico , Antineoplásicos/uso terapéutico , Leucemia-Linfoma de Células T del Adulto/tratamiento farmacológico , Adulto , Enfermedad Crónica , Ensayos Clínicos como Asunto , Humanos , Leucemia-Linfoma de Células T del Adulto/patología , Pronóstico , Inducción de RemisiónRESUMEN
Human T-cell leukemia virus type 1 (HTLV-1) propagates within and between individuals via cell-to-cell transmission, and primary infection typically occurs across juxtaposed mucosal surfaces during breastfeeding or sexual intercourse. It is therefore likely that dendritic cells (DCs) are among the first potential targets for HTLV-1. However, it remains unclear how DCs contribute to virus transmission and dissemination in the early stages of infection. We show that an HTLV-1-infected cell line (MT-2) and naturally infected CD4+ T cells transfer p19+ viral particles to the surface of allogeneic DCs via cell-to-cell contacts. Similarly organized cell-to-cell contacts also facilitate DC-mediated transfer of HTLV-1 to autologous CD4+ T cells. These findings shed light on the cellular structures involved in anterograde and retrograde transmission and suggest a key role for DCs in the natural history and pathogenesis of HTLV-1 infection.
Asunto(s)
Linfocitos T CD4-Positivos/virología , Células Dendríticas/virología , Virus Linfotrópico T Tipo 1 Humano/fisiología , Leucemia de Células T/patología , Replicación Viral , Linfocitos T CD4-Positivos/inmunología , Células Dendríticas/metabolismo , Células Dendríticas/ultraestructura , Humanos , Leucemia de Células T/metabolismo , Leucemia de Células T/virología , Microscopía Electrónica de Rastreo , Células Tumorales CultivadasRESUMEN
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
Simian T-Leukemia Virus type 1 and Simian Foamy Virus infect non-human primates. While STLV-1, as HTLV-1, causes Adult T-cell Leukemia/lymphoma, SFV infection is asymptomatic. Both retroviruses can be transmitted from NHPs to humans through bites that allow contact between infected saliva and recipient blood. Because both viruses infect CD4+ T-cells, they might interfere with each other replication, and this might impact viral transmission. Impact of STLV-1 co-infection on SFV replication was analyzed in 18 SFV-positive/STLV-1-negative and 18 naturally SFV/STLV-1 co-infected Papio anubis. Even if 9 animals were found STLV-1-positive in saliva, STLV-1 PVL was much higher in the blood. SFV proviruses were detected in the saliva of all animals. Interestingly, SFV proviral load was much higher in the blood of STLV-1/SFV co-infected animals, compared to STLV-1-negative animals. Given that soluble Tax protein can enter uninfected cells, we tested its effect on foamy virus promoter and we show that Tax protein can transactivate the foamy LTR. This demonstrates that true STLV-1 co-infection or Tax only has an impact on SFV replication and may influence the ability of the virus to be zoonotically transmitted as well as its ability to promote hematological abnormalities.
Asunto(s)
Coinfección/virología , Infecciones por Deltaretrovirus/virología , Infecciones por Retroviridae/virología , Virus Linfotrópico T Tipo 1 de los Simios/aislamiento & purificación , Virus Espumoso de los Simios/aislamiento & purificación , Carga Viral , Animales , Sangre/virología , Infecciones por Deltaretrovirus/complicaciones , Transmisión de Enfermedad Infecciosa , Papio anubis , Provirus/aislamiento & purificación , Infecciones por Retroviridae/complicaciones , Saliva/virología , Replicación ViralRESUMEN
BACKGROUND: The Human T-Lymphotropic Virus type 1c subtype (HTLV-1c) is highly endemic to central Australia where the most frequent complication of HTLV-1 infection in Indigenous Australians is bronchiectasis. We carried out a prospective study to quantify the prognosis of HTLV-1c infection and chronic lung disease and the risk of death according to the HTLV-1c proviral load (pVL). METHODOLOGY/PRINCIPAL FINDINGS: 840 Indigenous adults (discharge diagnosis of bronchiectasis, 154) were recruited to a hospital-based prospective cohort. Baseline HTLV-1c pVL were determined and the results of chest computed tomography and clinical details reviewed. The odds of an association between HTLV-1 infection and bronchiectasis or bronchitis/bronchiolitis were calculated, and the impact of HTLV-1c pVL on the risk of death was measured. Radiologically defined bronchiectasis and bronchitis/bronchiolitis were significantly more common among HTLV-1-infected subjects (adjusted odds ratio = 2.9; 95% CI, 2.0, 4.3). Median HTLV-1c pVL for subjects with airways inflammation was 16-fold higher than that of asymptomatic subjects. There were 151 deaths during 2,140 person-years of follow-up (maximum follow-up 8.13 years). Mortality rates were higher among subjects with HTLV-1c pVL ≥1000 copies per 105 peripheral blood leukocytes (log-rank χ2 (2df) = 6.63, p = 0.036) compared to those with lower HTLV-1c pVL or uninfected subjects. Excess mortality was largely due to bronchiectasis-related deaths (adjusted HR 4.31; 95% CI, 1.78, 10.42 versus uninfected). CONCLUSION/SIGNIFICANCE: Higher HTLV-1c pVL was strongly associated with radiologically defined airways inflammation and with death due to complications of bronchiectasis. An increased risk of death due to an HTLV-1 associated inflammatory disease has not been demonstrated previously. Our findings indicate that mortality associated with HTLV-1c infection may be higher than has been previously appreciated. Further prospective studies are needed to determine whether these results can be generalized to other HTLV-1 endemic areas.
Asunto(s)
Infecciones por HTLV-I/etnología , Infecciones por HTLV-I/virología , Virus Linfotrópico T Tipo 1 Humano/fisiología , Enfermedades Pulmonares/etnología , Nativos de Hawái y Otras Islas del Pacífico , Provirus/fisiología , Carga Viral , Adulto , Anciano , Australia/epidemiología , Bronquiectasia/epidemiología , Bronquiectasia/etnología , Bronquiectasia/virología , Bronquiolitis/epidemiología , Bronquiolitis/etnología , Bronquiolitis/virología , Bronquitis/epidemiología , Bronquitis/etnología , Bronquitis/virología , Enfermedad Crónica/epidemiología , Estudios de Cohortes , Supervivencia sin Enfermedad , Femenino , Infecciones por HTLV-I/epidemiología , Infecciones por HTLV-I/mortalidad , Virus Linfotrópico T Tipo 1 Humano/clasificación , Virus Linfotrópico T Tipo 1 Humano/genética , Virus Linfotrópico T Tipo 1 Humano/aislamiento & purificación , Humanos , Enfermedades Pulmonares/diagnóstico por imagen , Enfermedades Pulmonares/epidemiología , Enfermedades Pulmonares/virología , Masculino , Persona de Mediana Edad , Pronóstico , Estudios Prospectivos , Provirus/aislamiento & purificación , Factores de Riesgo , Tomografía Computarizada de EmisiónRESUMEN
Since the identification of the antisense protein of HTLV-2 (APH-2) and the demonstration that APH-2 mRNA is expressed in vivo in most HTLV-2 carriers, much effort has been dedicated to the elucidation of similarities and/or differences between APH-2 and HBZ, the antisense protein of HTLV-1. Similar to HBZ, APH-2 negatively regulates HTLV-2 transcription. However, it does not promote cell proliferation. In contrast to HBZ, APH-2 half-life is very short. Here, we show that APH-2 is addressed to PML nuclear bodies in T-cells, as well as in different cell types. Covalent SUMOylation of APH-2 is readily detected, indicating that APH-2 might be addressed to the PML nuclear bodies in a SUMO-dependent manner. We further show that silencing of PML increases expression of APH-2, while expression of HBZ is unaffected. On the other hand, SUMO-1 overexpression leads to a specific loss of APH-2 expression that is restored upon proteasome inhibition. Furthermore, the carboxy-terminal LAGLL motif of APH-2 is responsible for both the targeting of the protein to PML nuclear bodies and its short half-life. Taken together, these observations indicate that natural APH-2 targeting to PML nuclear bodies induces proteasomal degradation of the viral protein in a SUMO-dependent manner. Hence, this study deciphers the molecular and cellular bases of APH-2 short half-life in comparison to HBZ and highlights key differences in the post-translational mechanisms that control the expression of both proteins.
Asunto(s)
Virus Linfotrópico T Tipo 2 Humano/metabolismo , Cuerpos de Inclusión Intranucleares/metabolismo , Proteína de la Leucemia Promielocítica/metabolismo , Proteínas Virales/química , Proteínas Virales/metabolismo , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/metabolismo , Regulación Viral de la Expresión Génica , Semivida , Células HeLa , Virus Linfotrópico T Tipo 2 Humano/genética , Humanos , Células Jurkat , Complejo de la Endopetidasa Proteasomal/metabolismo , Proteolisis , Proteínas de los Retroviridae/metabolismo , Proteína SUMO-1/metabolismo , Sumoilación , Transcripción GenéticaAsunto(s)
Infecciones por HTLV-I/diagnóstico , Dermatosis del Cuero Cabelludo/diagnóstico , Enfermedades Cutáneas Bacterianas/diagnóstico , Oído , Femenino , Infecciones por HTLV-I/complicaciones , Humanos , Lactante , Cuello , Dermatosis del Cuero Cabelludo/microbiología , Enfermedades Cutáneas Bacterianas/tratamiento farmacológico , Enfermedades Cutáneas Bacterianas/microbiología , Australia del SurRESUMEN
A 36-year-old Caucasian male presented with adult T-cell leukemia/lymphoma (ATL). HTLV-1 contamination was attributed to a neonatal exchange transfusion. Remission was achieved but 11 years later he presented with symptoms suggesting ATL relapse. Molecular studies of T-cell clonality and virus integration sites revealed a clonal disease, distinct from the first tumor.
Asunto(s)
Protocolos de Quimioterapia Combinada Antineoplásica/administración & dosificación , Recambio Total de Sangre , Virus Linfotrópico T Tipo 1 Humano , Leucemia-Linfoma de Células T del Adulto/tratamiento farmacológico , Adulto , Femenino , Humanos , Leucemia-Linfoma de Células T del Adulto/etiología , Masculino , Embarazo , Inducción de RemisiónRESUMEN
HTLV-1 causes Adult T cell Leukemia/Lymphoma (ATLL) in humans. We describe an ATL-like disease in a 9 year-old female baboon naturally infected with STLV-1 (the simian counterpart of HTLV-1), with a lymphocyte count over 1010/L, lymphocytes with abnormal nuclear morphology, and pulmonary and skin lesions. The animal was treated with a combination of AZT and alpha interferon. Proviral load (PVL) was measured every week. Because the disease continued to progress, the animal was euthanized. Abnormal infiltrates of CD3+CD25+ lymphocytes and Tax-positive cells were found by histological analyses in both lymphoid and non-lymphoid organs. PVL was measured and clonal diversity was assessed by LM-PCR (Ligation-Mediated Polymerase Chain Reaction) and high throughput sequencing, in blood during treatment and in 14 different organs. The highest PVL was found in lymph nodes, spleen and lungs. One major clone and a number of intermediate abundance clones were present in blood throughout the course of treatment, and in organs. These results represent the first multi-organ clonality study in ATLL. We demonstrate a previously undescribed clonal complexity in ATLL. Our data reinforce the usefulness of natural STLV-1 infection as a model of ATLL.
Asunto(s)
Infecciones por Deltaretrovirus/veterinaria , Enfermedades de los Monos/patología , Virus Linfotrópico T Tipo 1 de los Simios , Animales , Infecciones por Deltaretrovirus/tratamiento farmacológico , Infecciones por Deltaretrovirus/patología , Infecciones por Deltaretrovirus/virología , Modelos Animales de Enfermedad , Femenino , Interferón-alfa/farmacología , Leucemia-Linfoma de Células T del Adulto/patología , Linfocitos/patología , Enfermedades de los Monos/tratamiento farmacológico , Enfermedades de los Monos/virología , Papio , Carga Viral , Zidovudina/farmacologíaRESUMEN
The therapeutic efficacy of the AZT and IFN combination in ATL presumably reflects the inhibition of RT-related functions.HTLV-1-RT activity from short-term cultured PBMCs may represent a predictive correlate of clinical response to AZT/IFN in ATL patients.
RESUMEN
UNLABELLED: Human T-cell lymphotropic virus type 1 (HTLV-1) and HTLV-2 encode auxiliary proteins that play important roles in viral replication, viral latency, and immune escape. The presence of auxiliary protein-encoding open reading frames (ORFs) in HTLV-3, the latest HTLV to be discovered, is unknown. Simian T-cell lymphotropic virus type 3 (STLV-3) is almost identical to HTLV-3. Given the lack of HTLV-3-infected cell lines, we took advantage of STLV-3-infected cells and of an STLV-3 molecular clone to search for the presence of auxiliary transcripts. Using reverse transcriptase PCR (RT-PCR), we first uncovered the presence of three unknown viral mRNAs encoding putative proteins of 5, 8, and 9 kDa and confirmed the presence of the previously reported RorfII transcript. The existence of these viral mRNAs was confirmed by using splice site-specific RT-PCR with ex vivo samples. We showed that p5 is distributed throughout the cell and does not colocalize with a specific organelle. The p9 localization is similar to that of HTLV-1 p12 and induced a strong decrease in the calreticulin signal, similarly to HTLV-1 p12. Although p8, RorfII, and Rex-3 share an N-terminal sequence that is predicted to contain a nucleolar localization signal (NoLS), only p8 is found in the nucleolus. The p8 location in the nucleolus is linked to a bipartite NoLS. p8 and, to a lesser extent, p9 repressed viral expression but did not alter Rex-3-dependent mRNA export. Using a transformation assay, we finally showed that none of the STLV-3 auxiliary proteins had the ability to induce colony formation, while both Tax-3 and antisense protein of HTLV-3 (APH-3) promoted cellular transformation. Altogether, these results complete the characterization of the newly described primate T-lymphotropic virus type 3 (PTLV-3). IMPORTANCE: Together with their simian counterparts, HTLVs form the primate T-lymphotropic viruses. HTLVs arose from interspecies transmission between nonhuman primates and humans. HTLV-1 and HTLV-2 encode auxiliary proteins that play important roles in viral replication, viral latency, and immune escape. The presence of ORFs encoding auxiliary proteins in HTLV-3 or STLV-3 genomes was unknown. Using in silico analyses, ex vivo samples, or in vitro experiments, we have uncovered the presence of 3 previously unknown viral mRNAs encoding putative proteins and confirmed the presence of a previously reported viral transcript. We characterized the intracellular localization of the four proteins. We showed that two of these proteins repress viral expression but that none of them have the ability to induce colony formation. However, both Tax and the antisense protein APH-3 promote cell transformation. Our results allowed us to characterize 4 new retroviral proteins for the first time.
Asunto(s)
Perfilación de la Expresión Génica , Virus Linfotrópico T Tipo 3 de los Simios/genética , Virus Linfotrópico T Tipo 3 de los Simios/fisiología , Proteínas Virales/análisis , Proteínas Virales/genética , Animales , Línea Celular , Núcleo Celular/química , Citosol/química , Humanos , Peso Molecular , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Proteínas Virales/químicaRESUMEN
BACKGROUND: Interferon induced transmembrane proteins 1, 2 and 3 (IFITMs) belong to a family of highly related antiviral factors that have been shown to interfere with a large spectrum of viruses including Filoviruses, Coronaviruses, Influenza virus, Dengue virus and HIV-1. In all these cases, the reported mechanism of antiviral inhibition indicates that the pool of IFITM proteins present in target cells blocks incoming viral particles in endosomal vesicles where they are subsequently degraded. RESULTS: In this study, we describe an additional mechanism through which IFITMs block HIV-1. In virus-producing cells, IFITMs coalesce with forming virions and are incorporated into viral particles. Expression of IFITMs during virion assembly leads to the production of virion particles of decreased infectivity that are mostly affected during entry in target cells. This mechanism of inhibition is exerted against different retroviruses and does not seem to be dependent on the type of Envelope present on retroviral particles. CONCLUSIONS: The results described here identify a novel mechanism through which IFITMs affect HIV-1 infectivity during the late phases of the viral life cycle. Put in the context of data obtained by other laboratories, these results indicate that IFITMs can target HIV at two distinct moments of its life cycle, in target cells as well as in virus-producing cells. These results raise the possibility that IFITMs could similarly affect distinct steps of the life cycle of a number of other viruses.
Asunto(s)
Antígenos de Diferenciación/metabolismo , VIH-1/inmunología , VIH-1/fisiología , Proteínas de la Membrana/metabolismo , Proteínas de Unión al ARN/metabolismo , Ensamble de Virus , Internalización del Virus , Antivirales/metabolismo , VIH-1/crecimiento & desarrollo , Interacciones Huésped-Patógeno , HumanosRESUMEN
Efficient HTLV-1 viral transmission occurs through cell-to-cell contacts. The Tax viral transcriptional activator protein facilitates this process. Using a comparative transcriptomic analysis, we recently identified a series of genes up-regulated in HTLV-1 Tax expressing T-lymphocytes. We focused our attention towards genes that are important for cytoskeleton dynamic and thus may possibly modulate cell-to-cell contacts. We first demonstrate that Gem, a member of the small GTP-binding proteins within the Ras superfamily, is expressed both at the RNA and protein levels in Tax-expressing cells and in HTLV-1-infected cell lines. Using a series of ChIP assays, we show that Tax recruits CREB and CREB Binding Protein (CBP) onto a c-AMP Responsive Element (CRE) present in the gem promoter. This CRE sequence is required to drive Tax-activated gem transcription. Since Gem is involved in cytoskeleton remodeling, we investigated its role in infected cells motility. We show that Gem co-localizes with F-actin and is involved both in T-cell spontaneous cell migration as well as chemotaxis in the presence of SDF-1/CXCL12. Importantly, gem knock-down in HTLV-1-infected cells decreases cell migration and conjugate formation. Finally, we demonstrate that Gem plays an important role in cell-to-cell viral transmission.
Asunto(s)
Citoesqueleto/metabolismo , Virus Linfotrópico T Tipo 1 Humano , Proteínas de Unión al GTP Monoméricas/metabolismo , Linfocitos T/metabolismo , Linfocitos T/virología , Línea Celular , Quimiotaxis de Leucocito/fisiología , Inmunoprecipitación de Cromatina , Técnica del Anticuerpo Fluorescente , Regulación Viral de la Expresión Génica/fisiología , Productos del Gen tax/metabolismo , Immunoblotting , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Linfocitos T/patología , Activación Transcripcional/fisiología , Transducción GenéticaRESUMEN
Non-human primates are considered to be likely sources of viruses that can infect humans and thus pose a significant threat to human population. This is well illustrated by some retroviruses, as the simian immunodeficiency viruses and the simian T lymphotropic viruses, which have the ability to cross-species, adapt to a new host and sometimes spread. This leads to a pandemic situation for HIV-1 or an endemic one for HTLV-1. Here, we present the available data on the discovery, epidemiology, cross-species transmission and molecular virology of the recently discovered HTLV-3 and HTLV-4 deltaretroviruses, as well as the simian foamy retroviruses present in different human populations at risk, especially in central African hunters. We discuss also the natural history in humans of these retroviruses of zoonotic origin (magnitude and geographical distribution, possible inter-human transmission). In Central Africa, the increase of the bushmeat trade during the last decades has opened new possibilities for retroviral emergence in humans, especially in immuno-compromised persons.
Asunto(s)
Infecciones por Deltaretrovirus/epidemiología , Infecciones por Deltaretrovirus/veterinaria , VIH-2/fisiología , Virus Linfotrópico T Tipo 3 Humano/fisiología , Virus Espumoso de los Simios/fisiología , África Central/epidemiología , Animales , Asia/epidemiología , Infecciones por Deltaretrovirus/transmisión , Infecciones por Deltaretrovirus/virología , Transmisión de Enfermedad Infecciosa/prevención & control , Productos del Gen tax/genética , VIH-2/patogenicidad , Haplorrinos , Especificidad del Huésped , Virus Linfotrópico T Tipo 3 Humano/patogenicidad , Humanos , Isoformas de Proteínas/genética , Virus Espumoso de los Simios/patogenicidad , Zoonosis/epidemiología , Zoonosis/transmisión , Zoonosis/virologíaRESUMEN
Monocyte-derived dendritic cells (MDDCs) play a key role in the regulation of the immune system and are the target of numerous gene therapy applications. The genetic modification of MDDCs is possible with human immunodeficiency virus type 1 (HIV-1)-derived lentiviral vectors (LVs) but requires high viral doses to bypass their natural resistance to viral infection, and this in turn affects their physiological properties. To date, a single viral protein is able to counter this restrictive phenotype, Vpx, a protein derived from members of the HIV-2/simian immunodeficiency virus SM lineage that counters at least two restriction factors present in myeloid cells. By tagging Vpx with a short heterologous membrane-targeting domain, we have obtained HIV-1 LVs incorporating high levels of this protein (HIV-1-Src-Vpx). These vectors efficiently transduce differentiated MDDCs and monocytes either as previously purified populations or as populations within unsorted peripheral blood mononuclear cells (PBMCs). In addition, these vectors can be efficiently pseudotyped with receptor-specific envelopes, further restricting their cellular tropism almost uniquely to MDDCs. Compared to conventional HIV-1 LVs, these novel vectors allow for an efficient genetic modification of MDDCs and, more importantly, do not cause their maturation or affect their survival, which are unwanted side effects of the transduction process. This study describes HIV-1-Src-Vpx LVs as a novel potent tool for the genetic modification of differentiated MDDCs and of circulating monocyte precursors with strong potential for a wide range of gene therapy applications.
Asunto(s)
Células Dendríticas/virología , Vectores Genéticos , VIH-1/inmunología , Monocitos/virología , Transducción Genética , Proteínas Reguladoras y Accesorias Virales/metabolismo , Células Cultivadas , VIH-1/genética , VIH-2/genética , Humanos , Virus de la Inmunodeficiencia de los Simios/genética , Proteínas Reguladoras y Accesorias Virales/genéticaRESUMEN
Permanent activation of the NF-κB pathway by the human T cell leukemia virus type 1 (HTLV-1) Tax (Tax1) viral transactivator is a key event in the process of HTLV-1-induced T lymphocyte immortalization and leukemogenesis. Although encoding a Tax transactivator (Tax2) that activates the canonical NF-κB pathway, HTLV-2 does not cause leukemia. These distinct pathological outcomes might be related, at least in part, to distinct NF-κB activation mechanisms. Tax1 has been shown to be both ubiquitinated and SUMOylated, and these two modifications were originally proposed to be required for Tax1-mediated NF-κB activation. Tax1 ubiquitination allows recruitment of the IKK-γ/NEMO regulatory subunit of the IKK complex together with Tax1 into centrosome/Golgi-associated cytoplasmic structures, followed by activation of the IKK complex and RelA/p65 nuclear translocation. Herein, we compared the ubiquitination, SUMOylation, and acetylation patterns of Tax2 and Tax1. We show that, in contrast to Tax1, Tax2 conjugation to endogenous ubiquitin and SUMO is barely detectable while both proteins are acetylated. Importantly, Tax2 is neither polyubiquitinated on lysine residues nor ubiquitinated on its N-terminal residue. Consistent with these observations, Tax2 conjugation to ubiquitin and Tax2-mediated NF-κB activation is not affected by overexpression of the E2 conjugating enzyme Ubc13. We further demonstrate that a nonubiquitinable, non-SUMOylable, and nonacetylable Tax2 mutant retains a significant ability to activate transcription from a NF-κB-dependent promoter after partial activation of the IKK complex and induction of RelA/p65 nuclear translocation. Finally, we also show that Tax2 does not interact with TRAF6, a protein that was shown to positively regulate Tax1-mediated activation of the NF-κB pathway.
Asunto(s)
Productos del Gen tax/metabolismo , Virus Linfotrópico T Tipo 2 Humano/patogenicidad , FN-kappa B/metabolismo , Proteínas Modificadoras Pequeñas Relacionadas con Ubiquitina/metabolismo , Sumoilación , Ubiquitina/metabolismo , Acetilación , Células HeLa , Humanos , Células Jurkat , Procesamiento Proteico-PostraduccionalRESUMEN
SAMHD1 is a newly identified restriction factor that targets lentiviruses in myeloid cells and is countered by the SIV(SM)/HIV-2 Vpx protein. By analyzing a large panel of Vpx mutants, we identify several residues throughout the 3-helix bundle predicted for Vpx that impair both its functionality and its ability to degrade SAMHD1. We determine that SAMHD1 is a strictly non-shuttling nuclear protein and that as expected WT Vpx localizes with it in the nucleus. However, we also identify a functional Vpx mutant with predominant cytoplasmic distribution that colocalizes with SAMHD1 in this location, suggesting that Vpx may also retain SAMHD1 in the cell cytoplasm, prior to its entry into the nucleus. Several mutations in Vpx were shown to affect the stability of Vpx, as well as Vpx:Vpx interactions. However, no strict correlation was observed between these parameters and the functionality of Vpx, implying that neither properties is absolutely required for this function and indicating that even unstable Vpx mutants may be very efficient in inducing SAMHD1 degradation. Overall, our analysis identifies several Vpx residues required for SAMHD1 degradation and points to a very efficient and plastic mechanism through which Vpx depletes this restriction factor.