RESUMEN
We cloned and sequenced gp41 HIV-1 from plasma of AIDS patients under HAART and T-20 (enfuvirtide, Fuzeon) therapy and revealed several T-20 resistance-associated mutations. Two mutations, a single V38A and a double N43T-N44K were the most frequent; however, they were not found together in one clone. We anticipated that simultaneous mutations of these three residues might play a vital role in the viral life cycle. To address this problem, we introduced N43T-N44K and V38M + N43T-N44K substitutions to a cloned gp41 and introduced modified gp41 into the pNL4-3 molecular clone. HEK293T cells were transfected with the obtained vectors and released viruses were examined for reverse transcriptase (RT) activity, infectivity on reporter TZM-bl cells, and in Western blotting. Nearly equal RT activity was demonstrated in viruses with and without mutations. However, viruses with the V38M + N43T-N44K mutations were not infectious and, as shown by Western blotting, gPr160 cleavage was impaired. These data suggest that V38M + N43T-N44K mutations perturbed the natural conformation of gPr160 in a way that access of furin to the cleavage site (REKR) was blocked. Therefore, the residues V38 + N43-N44 retain the gPr160 conformation in proximity to the furin cleavage site and, as a consequence, are critical for virus infectivity. These data may explain why viruses with V38M + N43T-N44K mutations were not previously detected in the plasma of T-20-experienced patients.
Asunto(s)
Sustitución de Aminoácidos/genética , Farmacorresistencia Viral , Proteínas gp160 de Envoltorio del VIH/metabolismo , Proteína gp41 de Envoltorio del VIH/genética , Infecciones por VIH/virología , VIH-1/aislamiento & purificación , Mutación Missense , Secuencia de Aminoácidos , Terapia Antirretroviral Altamente Activa , Línea Celular , Furina/metabolismo , Infecciones por VIH/tratamiento farmacológico , VIH-1/genética , Humanos , Hidrólisis , Datos de Secuencia Molecular , Procesamiento Proteico-PostraduccionalRESUMEN
Jaagsiekte retrovirus is an exogenous (exJSRV) beta-retrovirus with a simple genome. It causes lower airway epithelial cell tumors in small ruminants. Endogenous (enJSRV) counterparts of exJSRV are present in different copy numbers in numerous Bovidae family members. This work has focused on enJSRV in Simmental (Germany) and Limousine (France) beef breeds of domestic cattle and domestic goat. Of the enJSRV sequences in cattle, the orf-x sequences were about 99% identical, the LTR sequences were about 97% identical and the env sequences were nearly 95% identical to the corresponding endogenous sequences in sheep. A significant polymorphism of the proviral sequences between the cattle breeds was noted. Clonal analyses of the amplicons suggest two enJSRV proviruses in cattle genome. The endogenous sequences revealed in goat were closer to enzootic nasal tumor virus (ENTV) from goat rather than to enJSRV from sheep. The expression of enJSRV in cattle was partial (env only) and detected exclusively in bone marrow.
Asunto(s)
Animales Domésticos/virología , Bovinos/virología , Retrovirus Endógenos , Retrovirus Ovino Jaagsiekte , Análisis de Secuencia de ADN , Transcripción Genética , Secuencia de Aminoácidos , Animales , Retrovirus Endógenos/genética , Retrovirus Endógenos/metabolismo , Cabras/virología , Humanos , Retrovirus Ovino Jaagsiekte/genética , Retrovirus Ovino Jaagsiekte/metabolismo , Datos de Secuencia Molecular , Ovinos/virología , Sus scrofa/virologíaRESUMEN
Jaagsiekte retrovirus (JSRV) causes ovine pulmonary adenomatosis (OPA) that resembles bronchioloalveolar carcinoma (BAC) in humans. To test the possible role of JSRV in human diseases, DNA specimens from 103 individuals either healthy or suffering from lung carcinomas were analyzed for JSRV sequences. orf-x sequences were detected in 19 of 64 samples and gag-prt sequences in 4 of 38 samples, predominantly in individuals from Africa. Sequences obtained from orf-x amplimers varied in-between each other and differed from control endogenous ovine JSRV sequence. No association with lung cancer was found. This is the first report of JSRV-like sequences detected in humans.
Asunto(s)
Adenocarcinoma Bronquioloalveolar/virología , Secuencia de Bases , Betaretrovirus/genética , Retrovirus Ovino Jaagsiekte/genética , Neoplasias Pulmonares/virología , Ovinos/virología , Animales , Camerún , Carcinoma de Células Pequeñas/virología , Humanos , Datos de Secuencia Molecular , Nigeria , Adenomatosis Pulmonar Ovina/virología , Alineación de Secuencia , Análisis de Secuencia de ADNRESUMEN
Mother-to-child transmission can occur in utero, mainly intrapartum and postpartum in case of breastfeeding. In utero transmission is highly restricted and results in selection of viral variant from the mother to the child. We have developed an in vitro system that mimics the interaction between viruses, infected cells present in maternal blood, and the trophoblast, the first barrier protecting the fetus. Trophoblastic BeWo cells were grown as a tight polarized monolayer in a two-chamber system. Cell-free virions applied to the apical pole neither crossed the barrier nor productively infected BeWo cells. In contrast, apical contact with human immunodeficiency virus (HIV)-infected peripheral blood mononuclear cells (PBMCs) resulted in transcytosis of infectious virus across the trophoblastic monolayer and in productive infection correlating with the fusion of HIV-infected PBMCs with trophoblasts. We showed that viral variants are selected during these two steps and that in one case of in utero transmission, the predominant maternal viral variant characterized after transcytosis was phylogenetically indistinguishable from the predominant child's virus. Hence, the first steps of transmission of HIV-1 in utero appear to involve the interaction between HIV type 1-infected cells and the trophoblastic layer, resulting in the passage of infectious HIV by transcytosis and by fusion/infection, both leading to a selection of virus quasispecies.
Asunto(s)
Endocitosis/fisiología , VIH-1/fisiología , Transmisión Vertical de Enfermedad Infecciosa , Uniones Intercelulares/virología , Trofoblastos/virología , Comunicación Celular , Fusión Celular , Polaridad Celular , Femenino , Variación Genética , VIH-1/metabolismo , Humanos , Uniones Intercelulares/fisiología , Leucocitos Mononucleares/fisiología , Leucocitos Mononucleares/virología , Receptores CCR5/metabolismo , Receptores CXCR4/metabolismo , Trofoblastos/fisiología , Células Tumorales Cultivadas , Replicación ViralRESUMEN
Human T-cell leukemia virus type 1 (HTLV-1) is the etiologic agent of adult T-cell leukemia/lymphoma (ATLL), HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP), and other diseases. The mechanisms of virus pathogenesis are still obscure. The occurrence of defective proviruses in HTLV-1-infected cell lines and the peripheral blood mononuclear cells (PBMC) of infected individuals is a frequent feature of virus infection. We detected defective proviruses with large internal deletions in PBMC from ATLL and HAM/TSP patients and in asymptomatic HTLV-1 carriers. Seventeen PCR-amplified defective proviruses were sequenced, and three types of deletions were found. Besides truncated MA and the 5' end of the genome, truncated CA, truncated SU, and more frequently truncated TM linked to the pX region were detected. Reverse transcription-PCR analysis of PBMC from ATLL patients and asymptomatic carriers also revealed RNA transcripts with large internal deletions. Analysis of two RT-PCR cDNA clones confirmed a Gag-TM-pX structure of the transcripts. Most defective proviruses contained numerous internal stop codons, but some were capable of coding for the truncated MA linked to a variable out-of-frame peptide. Cloned defective proviruses with long open reading frames were subjected to in vitro transcription-translation followed by radioimmunoprecipitation, which showed expression of chimeric proteins between 8 and 12 kDa. Possible roles of defective proviruses and chimeric proteins are discussed, although there is no firm association with pathogenesis.
Asunto(s)
Virus Defectuosos/genética , Productos del Gen gag/genética , Infecciones por HTLV-I/virología , Virus Linfotrópico T Tipo 1 Humano/genética , Provirus/genética , Proteínas Oncogénicas de Retroviridae/genética , Eliminación de Secuencia , Factores de Transcripción , Proteínas del Envoltorio Viral/genética , Regiones no Traducidas 5' , Secuencia de Aminoácidos , Secuencia de Bases , Southern Blotting/métodos , Línea Celular Transformada , ADN Viral , Genoma Viral , Humanos , Leucocitos Mononucleares/virología , Datos de Secuencia Molecular , Reacción en Cadena de la Polimerasa/métodos , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/métodos , Proteínas de la Matriz Viral/genética , Proteínas Reguladoras y Accesorias ViralesRESUMEN
To determine the mechanisms by which human immunodeficiency virus type 1 (HIV-1) crosses the placenta into the fetal blood, 12 matched samples of serial maternal blood, term placentas, and infant blood obtained from a cohort of pregnant women in Cameroon identified as predominantly infected by subtype A viruses were studied. HIV-1 env sequences were detected by polymerase chain reaction (PCR) in both chorionic villi and enriched trophoblastic cells of all 12 placentas but at variable rates of detection. Heteroduplex mobility assay analysis showed the presence of multiple HIV-1 env quasispecies in sequential maternal peripheral blood mononuclear cell samples, but only a small number of env variants were found in chorionic villi and enriched trophoblastic cells. These data indicate that HIV-1 env sequences are always present in term placentas of seropositive women, contrasting with the low frequency at which infection is diagnosed by PCR in neonates with tat, gag, and env primers. Maternal HIV-1 variants appear to undergo a strong negative selection by different cell populations within the placental villi.
Asunto(s)
Infecciones por VIH/complicaciones , Infecciones por VIH/transmisión , VIH-1/clasificación , VIH-1/genética , Transmisión Vertical de Enfermedad Infecciosa , Placenta/virología , Complicaciones Infecciosas del Embarazo/virología , Secuencia de Bases , Camerún , Vellosidades Coriónicas/virología , Estudios de Cohortes , Cartilla de ADN/genética , ADN Viral/sangre , ADN Viral/genética , ADN Viral/aislamiento & purificación , Femenino , Sangre Fetal/virología , Genes env , Variación Genética , Infecciones por VIH/virología , VIH-1/aislamiento & purificación , Humanos , Recién Nacido , Repeticiones de Minisatélite , Reacción en Cadena de la Polimerasa , Embarazo , Selección Genética , Trofoblastos/virologíaRESUMEN
Human immunodeficiency virus, generated during contact between HIV-infected cells and the apical surface of an epithelial cell, can cross a tight epithelial barrier by transcytosis. We show that transcytosis of primary HIV isolates is blocked by dimeric IgA or IgM against HIV envelope proteins. Neutralization occurs intracellularly within the apical recycling endosome, and immune complexes are specifically recycled to the mucosal surface. One epitope involved in neutralization is a conserved sequence of the gp41 HIV envelope protein subunit. Finally, transcytosis also occurs across functional human mucosal tissue in a process inhibited by a serosal internalization of IgM against the HIV envelope protein. These results suggest that induction of mucosal immunity to HIV envelope proteins may impair the transcytotic route of HIV mucosal transmission.
Asunto(s)
Duodeno/citología , Proteínas gp160 de Envoltorio del VIH/inmunología , Proteína gp41 de Envoltorio del VIH/inmunología , VIH/inmunología , Líquido Intracelular/inmunología , Complejo Antígeno-Anticuerpo/metabolismo , Biopsia , Comunicación Celular , Duodeno/patología , Epítopos/inmunología , Humanos , Inmunoglobulina A/sangre , Inmunoglobulina A/inmunología , Inmunoglobulina M/sangre , Inmunoglobulina M/inmunología , Leucocitos Mononucleares/citología , Leucocitos Mononucleares/virología , Movimiento/fisiología , Membrana Mucosa/inmunología , Membrana Mucosa/virologíaRESUMEN
Sixteen matching sera and DNA samples from healthy African blood donors living in rural areas of Guinea were analysed for the presence of type D retrovirus markers. Screening for the antibodies against structural proteins of Mason-Pfizer monkey virus (M-PMV) was carried out by Western blot with a purified M-PMV as an antigen. Eight out of 16 sera samples were found to contain antibodies against at least two gag gene-coded proteins, and three of these were weakly positive against env gene-coded protein. Using PCR amplification and Southern hybridization, we detected M-PMV-like gag sequences in 11 out of 16 samples and env-related sequences in 8 out of 16 samples. Six DNAs were found to contain both M-PMV gag- and env-related sequences. Restriction endonuclease analysis of the PCR-amplified gag sequences from two individuals and direct DNA sequencing analysis of the amplimers confirmed their M-PMV-like origin. Detection of antibodies and M-PMV-related sequences in blood donors from Guinea, but not in French or Algerian blood donors, indicated exogenous SRV infection in humans from certain geographic areas of Western Africa.
Asunto(s)
Virus del Mono Mason-Pfizer/aislamiento & purificación , Adolescente , Adulto , Anticuerpos Antivirales/análisis , Biomarcadores , Southern Blotting , Western Blotting , Línea Celular , Niño , Enzimas de Restricción del ADN , ADN Viral/análisis , Femenino , Guinea , Humanos , Masculino , Proyectos Piloto , Reacción en Cadena de la Polimerasa , Infecciones por Retroviridae/virología , Infecciones Tumorales por Virus/virología , Proteínas Estructurales Virales/análisisRESUMEN
In this article, we describe the effect of indoleamines: serotonin (5-HT) and synthetic soluble melanin, on the multiplication of HIV-1 in T4 lymphocytic cell lines. The results show that viral production is increased when infected CEM-11 cells are incubated with 5-HT (10(-7) M and 10(-8) M) for 72 hours, whereas at higher doses (10(-3) M and 10(-4) M), there is an inhibition of viral multiplication. As well, when infected CEM cells were cultured in the presence of 5-HT at 10(-4) M, during 15 days, virus production, syncytia formation and cytolytic effect were drastically inhibited. Melanin also inhibits HIV-1 cytopathic effect on MT-2 cells, without cell toxicity, at concentrations of 0.2-10 micrograms/ml. Syncytium formation and cell lysis were also blocked by melanin at concentrations of 0.1 to 10 micrograms/ml, when uninfected MT-2 cells were mixed with HIV-1 chronically infected CEM-11 cells.
Asunto(s)
Antivirales/farmacología , VIH-1/efectos de los fármacos , Melaninas/farmacología , Serotonina/farmacología , Replicación Viral/efectos de los fármacos , Linfocitos T CD4-Positivos/efectos de los fármacos , Linfocitos T CD4-Positivos/virología , Células Cultivadas , Efecto Citopatogénico Viral/efectos de los fármacos , VIH-1/fisiología , HumanosRESUMEN
Viruses, and more particularly retroviruses, have been postulated to play a role in the pathogenesis of autoimmune diseases. In a search for spumaretrovirus infection markers, we screened a group of 29 patients with Graves disease and a representative healthy population (23 subjects) as a control. Southern blot hybridization under stringent conditions, of patients' DNA extracted from peripheral blood lymphocytes, with a spumaretrovirus-specific genomic probe derived from the human spumaretrovirus (HSRV) prototype, gave a positive signal in 10 cases. Moreover, by PCR, HSRV-related sequences were detected in the DNA of 19 patients (66%). Positive DNA samples in Southern blots were also positive in PCR for all regions tested (gag, bel1, bel2, long terminal repeat). Amplified (gag and bel2) products were cloned and sequenced; they showed high homology with HSRV. On the other hand, all 23 control subjects were negative by both procedures. Sera from both populations were examined for the presence of antibodies reactive with antigens of the spumaretrovirus family. These sera were negative by several immunodetection techniques: ELISA, indirect immunofluorescence, serum neutralization, and Western blotting. These results strongly suggest the existence of an association between Graves disease and the presence of HSRV-related infection markers.
Asunto(s)
ADN Viral/sangre , ADN/sangre , Genes Virales , Enfermedad de Graves/microbiología , Leucocitos/microbiología , Spumavirus/genética , Spumavirus/aislamiento & purificación , Animales , Secuencia de Bases , Southern Blotting , Clonación Molecular , Codón/genética , ADN/genética , ADN/aislamiento & purificación , ADN Viral/genética , ADN Viral/aislamiento & purificación , Enfermedad de Graves/sangre , Humanos , Datos de Secuencia Molecular , Oligodesoxirribonucleótidos , Reacción en Cadena de la Polimerasa/métodos , Secuencias Repetitivas de Ácidos NucleicosRESUMEN
Analysis was made of serum anti-HTLV-I antibodies, virus-specific proteins in peripheral blood lymphocytes (PBL) and proviruses in lymphocyte DNA of a patient with adult T-cell leukemia (ATL), Kaposi's sarcoma, and chronic myelopathy. Using Western blot and PCR (with HIV-1 specific primers), it was shown that Kaposi's sarcoma was not linked to HIV infection. Western blot analysis of serum revealed antibodies against p19, p24 and Pr 53 of HTLV-I. Examination of proteins in fresh PBL by Western blot revealed a high level of HTLV-I specific protein expression. Southern blot analysis of the patient's DNA revealed two different sites for HTLV-I provirus integration.
Asunto(s)
Virus Linfotrópico T Tipo 1 Humano/metabolismo , Leucemia de Células T/microbiología , Paraparesia Espástica Tropical/microbiología , Sarcoma de Kaposi/microbiología , Proteínas Virales/metabolismo , Anciano , Southern Blotting , ADN Viral/análisis , Anticuerpos Antideltaretrovirus/análisis , Femenino , Virus Linfotrópico T Tipo 1 Humano/genética , Humanos , Provirus/genética , Mapeo RestrictivoRESUMEN
Using Southern-Western and zinc-blotting techniques, the interactions of single-stranded DNA and zinc with structural proteins of type B, C, and D retroviruses were examined. Besides nucleic capsid proteins of retroviruses known to form complexes with nucleic acids, some other core proteins of type C retroviruses were shown to bind nucleic acids. All nucleic capsid proteins of the examined retroviruses appeared to be also zinc binding. In the present report, we propose two protein-blotting techniques that could be performed on a single nitrocellulose membrane. A first technique allows to detect zinc- and DNA-binding proteins immobilized on the membrane; a second (a modification of Southern-Western blotting) makes it possible to detect DNA-binding proteins followed by immunological reprobing.
Asunto(s)
Southern Blotting/métodos , Western Blotting/métodos , Retroviridae/química , Proteínas Virales/análisis , Proteínas de Unión al ADN/análisis , Zinc/metabolismo , Dedos de ZincRESUMEN
Relationships with retroviruses have recently been found in different human pathologies as autoimmune diseases which would be associated with the presence and eventually the expression of retroviral sequences. Detection of the presence of HTLV-1 and HIV-1 homologous sequences and their expression was realised on lymphocytes of 14 patients with polyendrocrinopathies (Basedow-Graves' disease and insulin-dependent diabetes) and four relatives of one index case. No antibodies to HTLV-1 and HIV-1 could be detected by Western blot and Elisa tests. HTLV-1 related sequences were revealed by Southern blot (SB) in 5 out of 18 subjects' DNA. Analyses of all DNA were performed by polymerase chain reaction (PCR). Seven DNA, including the 5 previously positive in SB, and two relatives (father and grandfather), negative in SB, contained HTLV-1-gag related sequences, but neither pol nor pX regions. Concerning HIV-1, all 18 DNA examined were negative by both methods. DNA of ten clinically healthy donors were found to be negative with the same tests.
Asunto(s)
ADN/química , Diabetes Mellitus Tipo 1/complicaciones , Genes gag/genética , Enfermedad de Graves/complicaciones , Virus Linfotrópico T Tipo 1 Humano/genética , Anciano , Secuencia de Bases , Southern Blotting , Diabetes Mellitus Tipo 1/sangre , Enfermedad de Graves/sangre , Humanos , Leucocitos , Persona de Mediana Edad , Datos de Secuencia MolecularRESUMEN
Twelve long-term cell lines were established from peripheral blood mononuclear cells (PBMC) or cerebrospinal fluid cells of patients with human T lymphotropic virus type I (HTLV-1) seropositive tropical spastic paraparesis (TSP) originating from the French West Indies, French Guyana or the Central African Republic. Most of these long-term interleukin-2-dependent cell lines exhibited a pattern characteristic of CD4(+)-activated T cells with high expression of CD2, CD3 and CD4 antigens, associated with a strong density of TAC and DR molecules. Nevertheless, in five cases CD8 expression was present at a significant level. HTLV-I antigens were never detected in uncultured PBMC, but they were expressed in a few cells after short-term culture and after 4 months the majority of the cells were HTLV-I positive, as demonstrated by indirect immunofluorescence (IF) using polyclonal or monoclonal anti-p19 and anti-p24 antibodies. Low and variable levels of reverse transcriptase activity were detected in supernatant fluids of these cell lines only after 4 months of culture, when at least 50% of the cells exhibited HTLV-I antigens by IF. However, numerous type C HTLV-I-like viral particles were detected, mostly in the extracellular spaces, with rare budding particles. Similar findings were found in three T cell lines derived from West Indian and African patients with adult T-cell leukaemia/lymphoma (ATLL). Differences in high Mr polypeptides were detected by Western blot in cell lysates when comparing TSP- or ATLL-derived T cell lines. Thus a signal of 62K was easily detectable in all the TSP lines, but not in the ATLL lines. In all cell lines bands corresponding to p53, p24 and p19 viral core polypeptides were present, as was the env gene-coded protein p46.
Asunto(s)
Antígenos HTLV-I/análisis , Virus Linfotrópico T Tipo 1 Humano/inmunología , Paraparesia Espástica Tropical/inmunología , Linfocitos T/inmunología , Adulto , Antígenos CD/análisis , Western Blotting , División Celular , Línea Celular , República Centroafricana , Femenino , Guyana Francesa , Virus Linfotrópico T Tipo 1 Humano/crecimiento & desarrollo , Virus Linfotrópico T Tipo 1 Humano/ultraestructura , Humanos , Masculino , Microscopía Electrónica , Persona de Mediana Edad , Paraparesia Espástica Tropical/sangre , Paraparesia Espástica Tropical/líquido cefalorraquídeo , Fenotipo , Linfocitos T/microbiología , Linfocitos T/ultraestructura , Proteínas del Núcleo Viral/análisis , Indias OccidentalesRESUMEN
In the course of a systematic screening of sera from dogs suffering from autoimmune disorders, three sera were shown by indirect immunofluorescence to characteristically label the nucleoli and nucleoplasm of human cell lines (Hep-2 and HeLa). This pattern of staining persisted throughout the cell cycle, except for mitosis when the fluorescence was localized in extrachromosomal areas. By immunoblotting nuclear and subnuclear fractions, three polypeptides of 110,000, 95,000, and 45,000 Da apparent molecular weight were identified, which reacted with all three sera. By means of affinity purification, it was shown that an antibody specific for any one of the three proteins also reacts with the two others. This antigenic cross-reactivity suggested regions of structural homology shared by the three proteins. Indeed, treatment of nucleoli with high concentrations of DNase I containing residual proteolytic activity resulted in the disappearance of the 110- and 95-kDa proteins and the concomitant appearance of a doublet of 45-kDa proteins. Subnuclear localization studies indicated that all three polypeptides were located in both nucleoli and nucleoplasm. Significantly, the 110-kDa protein differs from the major nucleolar protein, nucleolin, by its electrophoretic mobility in two-dimensional gels, its location in nucleoli and in nucleoplasm, its absence in nucleolar organizer regions of chromosomes, and its differential solubility of DNase I. Therefore, the three antigenically related species reported in this study constitute a new class of nucleolar proteins.
Asunto(s)
Enfermedades Autoinmunes/veterinaria , Nucléolo Celular/inmunología , Enfermedades de los Perros/inmunología , Proteínas Nucleares/inmunología , Animales , Enfermedades Autoinmunes/inmunología , Línea Celular , Nucléolo Celular/ultraestructura , Perros , Técnica del Anticuerpo Fluorescente , Células HeLa/inmunología , Células HeLa/ultraestructura , Humanos , Sueros Inmunes , Immunoblotting , Peso Molecular , Proteínas Nucleares/aislamiento & purificaciónRESUMEN
The sera of two patients with autoimmune disorders recognize in HeLa cell extracts two proteins with apparent molecular masses of 37,000 (p37) daltons and 32,000 daltons (p32). These proteins are non covalently associated with 5S RNA and sediment as 7-10 S particles in sucrose density gradients. Both proteins are antigenetically related to TFIIIA, a previously described protein of Xenopus laevis, which is known as a 5S RNA transcription factor and occurs in oocytes as a noncovalent complex with 5S RNA. Like TFIIIA, HeLa cell proteins p37 binds in vitro to 5S RNA and to cloned 5S RNA genes. These results suggest that protein p37 fulfils in HeLa cells a function similar to that of TFIIIA in amphibian oocytes, ie control of 5S RNA transcription.
Asunto(s)
Autoanticuerpos/inmunología , Enfermedades Autoinmunes/inmunología , Proteínas Nucleares/inmunología , ARN Ribosómico 5S/metabolismo , ARN Ribosómico/metabolismo , Ribonucleoproteínas/inmunología , Factores de Transcripción/inmunología , Animales , Especificidad de Anticuerpos , Compartimento Celular , Citoplasma/metabolismo , Células HeLa , Humanos , Peso Molecular , Xenopus laevisRESUMEN
PSL(p55) is a nuclear 55kD antigen present in various mammalian cell systems, which has been first identified by use of human autoimmune antibodies (Barque et al. 1983, EMBO J. 2, 743). It has been shown to be associated with interphase chromatine and to be synthesized in during the S phase of the cell cycle. In this work, we have analysed the status of PSL in promyelocytic HL-60 human cells in exponential or stationary growth, or undergoing granulocytic differentiation in presence of Retinoic acid. By use of 2-dimensional electrophoresis, PSL was found to be composed of two acidic proteins designated p55A and p55B. Unexpectedly, estimated 10-20 fold higher amounts of each species were found in cells treated for 5 days with 10(-6)M Retinoic acid, than in asynchronously growing cells or resting cells. Moreover, the p55A protein was phosphorylated during the process. On the basis of these results, PSL appears to be involved in some steps of the granulocytic differentiation process.
Asunto(s)
Ciclo Celular , Diferenciación Celular , Proteínas Nucleares/metabolismo , Tretinoina/farmacología , Diferenciación Celular/efectos de los fármacos , Humanos , Punto Isoeléctrico , Peso Molecular , Fosfoproteínas/metabolismo , Fosforilación , Células Tumorales CultivadasRESUMEN
Using the sera from two patients with autoimmune disorders, we have identified by immunoprecipitation of HeLa cell extracts two proteins with apparent molecular masses of 37 kDa (p 37) and 32 kDa (p 32). These proteins are associated with 5 S RNA. They are antigenetically related to Xenopus laevis 5 S RNA transcription factor TFIIIA, which is very abundant in early oocytes of this species. In contrast to what is observed in X. laevis oocytes, the TFIIIA-related proteins of HeLa cells are present in very small amounts. Our data suggest that proteins p 37 and p 32 are involved in the control of 5 S RNA transcription.