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1.
Vet Pathol ; 52(3): 566-72, 2015 May.
Artículo en Inglés | MEDLINE | ID: mdl-25248517

RESUMEN

Lymphomas are common spontaneous tumors in nonhuman primates but remain poorly characterized in Japanese macaques (Macaca fuscata). This study examined 5 cases of spontaneous malignant lymphoma in Japanese macaques, focusing on the immunophenotypes and presence of simian lymphocryptoviruses, which are Epstein-Barr virus-related herpesviruses in nonhuman primates. The macaques with lymphoma were 5 to 28 years old, indicating that lymphomas develop over a wide age range. The common macroscopic findings were splenomegaly and enlargement of lymph nodes. Histologic and immunohistochemical analyses revealed that all cases were non-Hodgkin type and exhibited a T-cell phenotype, positive for CD3 but negative for CD20 and CD79α. The lymphomas exhibited diverse cellular morphologies and were subdivided into 3 types according to the World Health Organization classification. These included 3 cases of peripheral T-cell lymphoma, not otherwise specified; 1 case of T-cell prolymphocytic leukemia; and 1 case of an unclassifiable T-cell lymphoma. Positive signals were detected by in situ hybridization in 2 of the 4 examined cases using probes for the Epstein-Barr virus-encoded small RNA (EBER). Furthermore, the presence of M. fuscata lymphocryptovirus 2, a macaque homolog of Epstein-Barr virus, was demonstrated in EBER-positive cases by polymerase chain reaction amplification followed by direct sequencing. Immunohistochemistry using antibody to the Epstein-Barr virus-encoded nuclear antigen 2 was negative, even in the EBER-positive cases. The present study suggests that T-cell lymphoma is more common than B-cell lymphoma in Japanese macaques and that M. fuscata lymphocryptovirus 2 is present in some cases.


Asunto(s)
Linfoma/veterinaria , Enfermedades de los Monos/patología , Animales , Femenino , Infecciones por Herpesviridae/diagnóstico , Infecciones por Herpesviridae/patología , Infecciones por Herpesviridae/veterinaria , Hibridación in Situ/veterinaria , Leucemia Prolinfocítica de Células T/diagnóstico , Leucemia Prolinfocítica de Células T/patología , Leucemia Prolinfocítica de Células T/veterinaria , Leucemia Prolinfocítica de Células T/virología , Ganglios Linfáticos/patología , Lymphocryptovirus , Linfoma/complicaciones , Linfoma/patología , Linfoma/virología , Linfoma de Células T/diagnóstico , Linfoma de Células T/patología , Linfoma de Células T/veterinaria , Linfoma de Células T/virología , Linfoma de Células T Periférico/diagnóstico , Linfoma de Células T Periférico/patología , Linfoma de Células T Periférico/veterinaria , Linfoma de Células T Periférico/virología , Macaca , Masculino , Enfermedades de los Monos/diagnóstico , Enfermedades de los Monos/virología , Reacción en Cadena en Tiempo Real de la Polimerasa/veterinaria , Esplenomegalia/etiología , Esplenomegalia/patología , Esplenomegalia/veterinaria , Infecciones Tumorales por Virus/diagnóstico , Infecciones Tumorales por Virus/patología , Infecciones Tumorales por Virus/veterinaria , Infecciones Tumorales por Virus/virología
2.
Vet Pathol ; 42(3): 391-6, 2005 May.
Artículo en Inglés | MEDLINE | ID: mdl-15872392

RESUMEN

Multifocal submucosal stromal tumors were diagnosed in a 5.5-year-old rhesus macaque (Macaca mulatta) experimentally infected with simian immunodeficiency virus, strain SIVsmE660, and CD4+ T cell depleted. The animal was negative for simian retroviruses, SRV-1, -2, and -5. Polymerase chain reaction analysis of DNA from tumor and spleen tissue revealed abundant, preferential presence of retroperitoneal fibromatosis herpesvirus, the macaque homologue of the Kaposi sarcoma-associated herpesvirus (human herpesvirus-8), in the tumors. This was corroborated by demonstration of viral latent nuclear antigen-1 in the nuclei of a majority of the spindeloid tumor cells. Low levels of an additional macaque herpesvirus, rhesus rhadinovirus, were also detected in the spleen and tumor tissues. The spindeloid cells labeled positively for vimentin and CD117 but were negative for CD31, CD68, desmin, and smooth muscle cell actin. Collectively, these findings suggest a relation to but not absolute identity with simian mesenchymoproliferative disorders (MPD) or typical gastrointestinal stromal tumors (GISTs).


Asunto(s)
Tumores del Estroma Gastrointestinal/veterinaria , Macaca mulatta , Enfermedades de los Monos/patología , Enfermedades de los Monos/virología , Síndrome de Inmunodeficiencia Adquirida del Simio/complicaciones , Virus de la Inmunodeficiencia de los Simios , Animales , Antígenos Nucleares/metabolismo , Cartilla de ADN , Tumores del Estroma Gastrointestinal/complicaciones , Tumores del Estroma Gastrointestinal/patología , Herpesvirus Humano 8/metabolismo , Inmunohistoquímica/veterinaria , Reacción en Cadena de la Polimerasa/veterinaria , Retrovirus de los Simios/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa/veterinaria , Vimentina/metabolismo
3.
Leukemia ; 17(1): 185-95, 2003 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-12529677

RESUMEN

Real-time quantitative polymerase chain reaction (qPCR) was used to quantify viral loads of human herpesviruses (HHVs) at diagnosis in 61 samples of malignant B cells: 21 chronic lymphocytic leukemia (B-CLL), 29 acute lymphoblastic leukemia (B-ALL) and 11 multiple myeloma (MM); control samples were blasts from 16 acute myeloid leukemia (AML) and 24 blood or bone marrow samples from healthy donors. The majority of samples from healthy donors and patients (B-ALL, B-CLL or AML, but not MM) was positive for EBV and contained <100 ebv copies/100 ng dna. ebv loads were occasionally high (>500 copies/100 ng DNA) in B-ALL (2/16) and in B-CLL (2/21) samples. The fractions of samples positive for HHV-8 and HHV-6A, less than 10% for MM patients, were high for adults with B-ALL (18.8% HHV-8+, 43.8% HHV-6A+) or B-CLL (28.6% HHV-8+, 52.4% HHV-6A+). B-ALL, B-CLL and MM samples were rarely positive for HHV-6B and HHV-7. Lastly, 75% of B-ALL samples were positive for CMV, and CMV loads were significantly higher in B-ALL samples than in MM, B-CLL or AML samples. We also used PCR with consensus-degenerate hybrid oligonucleotide primers (CODEHOP) to look for novel HHVs in B cell samples: no sequence indicative of a new HHV was detected. Altogether, the data indicate that the presence of multiple HHVs, including EBV and CMV at high loads, is not rare in B-ALL and B-CLL cell samples. Future prospective studies should determine whether patients with high EBV/CMV loads at diagnosis in tumor samples face a higher risk of delayed hematological recovery, virus-related complications or relapse.


Asunto(s)
Linfoma de Burkitt/virología , ADN Viral/análisis , Herpesvirus Humano 6/aislamiento & purificación , Herpesvirus Humano 7/aislamiento & purificación , Herpesvirus Humano 8/aislamiento & purificación , Leucemia Linfocítica Crónica de Células B/virología , Mieloma Múltiple/virología , Reacción en Cadena de la Polimerasa/métodos , Adolescente , Adulto , Anciano , Secuencia de Aminoácidos , Secuencia de Bases , Estudios de Casos y Controles , Niño , Preescolar , Citomegalovirus/genética , Citomegalovirus/aislamiento & purificación , Cartilla de ADN/genética , Femenino , Herpesvirus Humano 4/genética , Herpesvirus Humano 4/aislamiento & purificación , Herpesvirus Humano 6/genética , Herpesvirus Humano 7/genética , Herpesvirus Humano 8/genética , Humanos , Lactante , Masculino , Persona de Mediana Edad , Datos de Secuencia Molecular , Reacción en Cadena de la Polimerasa/normas , Reacción en Cadena de la Polimerasa/estadística & datos numéricos , Sensibilidad y Especificidad , Homología de Secuencia de Aminoácido , Homología de Secuencia de Ácido Nucleico , Carga Viral
4.
J Bacteriol ; 182(24): 7067-9, 2000 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-11092870

RESUMEN

Dental plaque is a complex biofilm that accretes in a series of discrete steps proceeding from a gram-positive streptococcus-rich biofilm to a structure rich in gram-negative anaerobes. This study investigated information flow between two unrelated plaque bacteria, Streptococcus cristatus and Porphyromonas gingivalis. A surface protein of S. cristatus caused repression of the P. gingivalis fimbrial gene (fimA), as determined by a chromosomal fimA promoter-lacZ reporter construct and by reverse transcription-PCR. Signaling activity was associated with a 59-kDa surface protein of S. cristatus and showed specificity for the fimA gene. Furthermore, P. gingivalis was unable to form biofilm microcolonies with S. cristatus. Thus, S. cristatus is capable of modulating virulence gene expression in P. gingivalis, consequently influencing the development of pathogenic plaque.


Asunto(s)
Proteínas Bacterianas/metabolismo , Biopelículas/crecimiento & desarrollo , Placa Dental/microbiología , Proteínas Fimbrias , Porphyromonas gingivalis/genética , Porphyromonas gingivalis/patogenicidad , Streptococcus/genética , Antibiosis , Proteínas Bacterianas/genética , Infecciones por Bacteroidaceae/microbiología , Regulación Bacteriana de la Expresión Génica , Porphyromonas gingivalis/metabolismo , Transducción de Señal , Streptococcus/metabolismo , Virulencia
5.
Biochem Cell Biol ; 78(4): 437-45, 2000.
Artículo en Inglés | MEDLINE | ID: mdl-11012082

RESUMEN

Transforming infection of Go/G1-arrested primary mouse kidney cell cultures with simian virus 40 (SV40) induces cells to re-enter the S-phase of the cell cycle. In Go-arrested cells, no p53 is detected, whereas in cells induced to proliferate by infection, a gradual accumulation of p53 complexed to SV40 large T-antigen is observed in the nucleus. Heat treatment of actively proliferating SV40-infected cells leads to inhibition of DNA synthesis and growth arrest. To determine the fate of p53 after heat treatment, proliferating infected cells were exposed to mild heat (42.5 degrees C) for increasing lengths of time. The results presented here show that after ninety minutes of treatment, the arrest of DNA synthesis by heat correlates with the disruption of the p53/LT-antigen complex. Longer treatments induce, in addition, a reduction in the solubility of p53, which was recovered tightly associated with the nuclear fraction. This contrasted with large T-antigen, whose solubility remained unaffected by heat treatment. Although the total amount of p53 in the nucleus remained constant, as shown by immunoblot analyses, p53 was no longer detectable after immunoprecipitation or by immunofluorescent staining techniques. These results suggest that heat treatment had either induced conformational changes in its antigenic sites, or had sequestered the sites through aggregation or binding to insoluble nuclear components.


Asunto(s)
Antígenos Transformadores de Poliomavirus/metabolismo , Transformación Celular Viral , ADN/biosíntesis , Calor , Fase S , Virus 40 de los Simios/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Animales , Antígenos Transformadores de Poliomavirus/química , Ciclo Celular , Núcleo Celular/metabolismo , Células Cultivadas , Electroforesis en Gel de Poliacrilamida , Células Epiteliales , Proteínas de Choque Térmico/biosíntesis , Immunoblotting , Riñón/citología , Ratones , Virus 40 de los Simios/genética , Solubilidad , Proteína p53 Supresora de Tumor/química
6.
J Virol ; 74(10): 4919-28, 2000 May.
Artículo en Inglés | MEDLINE | ID: mdl-10775636

RESUMEN

We have cloned and characterized the entire DNA polymerase gene and flanking regions from Kaposi's sarcoma-associated herpesvirus (KSHV) and two closely related macaque homologs of KSHV, retroperitoneal fibromatosis-associated herpesvirus-Macaca nemestrina (RFHVMn) and -Macaca mulatta (RFHVMm). We have also identified and partially characterized the corresponding genomic region of another KSHV-like herpesvirus, provisionally named "M. nemestrina rhadinovirus type 2 (MneRV-2)," with close similarity to rhesus rhadinovirus (RRV). A sequence comparison of these four macaque viruses and two KSHV-like gammaherpesviruses recently identified in African green monkeys, Chlorocebus rhadinovirus types 1 and 2 (ChRV-1 and ChRV-2) reveals the presence of two distinct lineages of KSHV-like rhadinoviruses in Old World primates. The first rhadinovirus lineage consists of KSHV and its closely related homologs RFHVMn, RFHVMm, and ChRV-1, while the second more distantly related lineage consists of RRV, MneRV-2, and ChRV-2. Our findings raise the possibility of the existence of another human KSHV-like herpesvirus belonging to the second rhadinovirus lineage.


Asunto(s)
Gammaherpesvirinae/genética , Herpesvirus Humano 8/genética , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Clonación Molecular , ADN Polimerasa Dirigida por ADN/química , ADN Polimerasa Dirigida por ADN/genética , Evolución Molecular , Gammaherpesvirinae/clasificación , Humanos , Macaca mulatta , Macaca nemestrina , Datos de Secuencia Molecular , Enfermedades de los Monos/virología , Sistemas de Lectura Abierta , Filogenia , Fibrosis Retroperitoneal/veterinaria , Fibrosis Retroperitoneal/virología , Sarcoma de Kaposi/virología , Proteínas del Envoltorio Viral/química , Proteínas del Envoltorio Viral/genética
7.
Virology ; 260(1): 116-24, 1999 Jul 20.
Artículo en Inglés | MEDLINE | ID: mdl-10405363

RESUMEN

We have identified a novel lentivirus prevalent in talapoin monkeys (Myopithecus talapoin), extending previous observations of human immunodeficiency virus-1 cross-reactive antibodies in the serum of these monkeys. We obtained a virus isolate from one of three seropositive monkeys initially available to us. The virus was tentatively named simian immunodeficiency virus from talapoin monkeys (SIVtal). Despite the difficulty of isolating this virus, it was readily passed between monkeys in captivity through unknown routes of transmission. The virus could be propagated for short terms in peripheral blood mononuclear cells of talapoin monkeys but not in human peripheral blood mononuclear cells or human T cell lines. The propagated virus was used to infect a naive talapoin monkey, four rhesus macaques (M. mulatta), and two cynomolgus macaques (M. fascicularis). All animals seroconverted and virus could be reisolated during a short period after experimental infection. A survey of SIVtal-infected captive talapoin monkeys revealed a relative decrease in CD4(+) cell numbers in chronically (>2 years) infected animals. No other signs of immunodeficiency were observed in any of the infected animals. PCR amplification followed by DNA sequencing of two fragments of the polymerase gene revealed that SIVtal is different from the presently known lentiviruses and perhaps most related to the SIV from Sykes monkeys.


Asunto(s)
Cercopithecidae/virología , VIH-1/inmunología , Virus de la Inmunodeficiencia de los Simios/aislamiento & purificación , Animales , Antígenos CD4/análisis , Antígenos CD8/análisis , Células Cultivadas , Cercopithecidae/inmunología , Células Clonales , Reacciones Cruzadas , Transmisión de Enfermedad Infecciosa , Anticuerpos Anti-VIH/inmunología , Seropositividad para VIH/inmunología , Vivienda para Animales , Humanos , Leucocitos Mononucleares/virología , Macaca mulatta , Microscopía Electrónica , Datos de Secuencia Molecular , Síndrome de Inmunodeficiencia Adquirida del Simio/inmunología , Síndrome de Inmunodeficiencia Adquirida del Simio/transmisión , Virus de la Inmunodeficiencia de los Simios/clasificación , Virus de la Inmunodeficiencia de los Simios/genética , Virus de la Inmunodeficiencia de los Simios/inmunología , Cultivo de Virus/métodos
8.
Am J Respir Cell Mol Biol ; 20(2): 327-31, 1999 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-9922225

RESUMEN

Fibroblasts are the major cell type responsible for synthesizing matrix constituents in lung and other connective tissues. Evidence indicates that fibroblasts are heterogeneous, and that subpopulations with some distinct properties are clonally selected and expanded in fibrotic diseases. However, few distinct markers capable of demonstrating the presence of fibroblast subpopulations in tissues have been isolated so far. With the objective of identifying proteins that could detect fibroblast subpopulations, we compared the messenger RNA (mRNA) expression of two cultured human lung fibroblast subpopulations by differential display. Total RNA was obtained, complementary DNA (cDNA) was synthesized, and the polymerase chain reaction (PCR) products obtained with several primer pairs were compared. One 724-bp product, which was strongly expressed by one human lung fibroblast subpopulation, was identified and cloned. This product was poorly expressed by the other lung fibroblast subpopulation. The mRNA for the gene encoding this product was not detectable in human smooth-muscle cells, endothelial cells, or epithelial cells, although it was present in dermal fibroblasts. The mRNA was detected in normal and fibrotic human lungs. Search of the National Center for Biotechnology (NCBI) GenBank DNA database with the sequence obtained from this clone revealed no significant matches. However, a search of the NCBI database of expressed sequence tags (dBEST) revealed five different human expressed sequence tag (EST) clones corresponding to the LR8 cDNA sequence. Six additional mouse and one pig EST clones were identified that showed significant similarity to the human fibroblast cDNA. Composites of the entire coding sequences for the human fibroblast gene product and the mouse homologue were assembled from the respective overlapping EST sequences. The open reading frame identified for each composite sequence predicted protein products of 270 and 263 amino acids for the human and mouse sequences, respectively, which were 52% identical, with three gaps. At the amino acid level, no significant sequence similarity was detected with any other sequences in exhaustive searches of the NCBI DNA and protein databases or the Blocks databases. A PCR product with predicted length and sequence was obtained by using a sense primer upstream to LR8 and an antisense primer within LR8. Our results indicate that this differentially displayed product represents a previously undescribed protein that could be useful for distinguishing fibroblasts, and possibly fibroblast subpopulations, from other cell types in lungs and other tissues.


Asunto(s)
Proteínas Aviares , Lipoproteínas/genética , Pulmón/metabolismo , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Clonación Molecular , ADN Complementario , Fibroblastos/metabolismo , Humanos , Pulmón/citología , Ratones , Datos de Secuencia Molecular , Sistemas de Lectura Abierta , ARN Mensajero/genética , ARN Mensajero/metabolismo , Homología de Secuencia de Aminoácido , Homología de Secuencia de Ácido Nucleico , Porcinos
10.
Nucleic Acids Res ; 26(7): 1628-35, 1998 Apr 01.
Artículo en Inglés | MEDLINE | ID: mdl-9512532

RESUMEN

We describe a new primer design strategy for PCR amplification of unknown targets that are related to multiply-aligned protein sequences. Each primer consists of a short 3' degenerate core region and a longer 5' consensus clamp region. Only 3-4 highly conserved amino acid residues are necessary for design of the core, which is stabilized by the clamp during annealing to template molecules. During later rounds of amplification, the non-degenerate clamp permits stable annealing to product molecules. We demonstrate the practical utility of this hybrid primer method by detection of diverse reverse transcriptase-like genes in a human genome, and by detection of C5DNA methyltransferase homologs in various plant DNAs. In each case, amplified products were sufficiently pure to be cloned without gel fractionation. This COnsensus-DEgenerate Hybrid Oligonucleotide Primer (CODEHOP) strategy has been implemented as a computer program that is accessible over the World Wide Web (http://blocks.fhcrc.org/codehop.html) and is directly linked from the BlockMaker multiple sequence alignment site for hybrid primer prediction beginning with a set of related protein sequences.


Asunto(s)
Metilasas de Modificación del ADN/química , Cartilla de ADN , Evolución Molecular , Filogenia , ADN Polimerasa Dirigida por ARN/química , Secuencia de Aminoácidos , Animales , Artritis Reumatoide/genética , Secuencia de Bases , Codón , Redes de Comunicación de Computadores , Secuencia de Consenso , Secuencia Conservada , Metilasas de Modificación del ADN/genética , Humanos , Datos de Secuencia Molecular , Hibridación de Ácido Nucleico , Reacción en Cadena de la Polimerasa/métodos , ADN Polimerasa Dirigida por ARN/genética , Sarcoma de Kaposi/genética , Alineación de Secuencia , Homología de Secuencia de Aminoácido , Homología de Secuencia de Ácido Nucleico , Programas Informáticos
11.
J Virol ; 72(4): 3082-7, 1998 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-9525633

RESUMEN

As part of the evaluation of porcine cells, tissues, and organs intended for transplantation into humans, we investigated the conditions required to induce expression and release of porcine endogenous retrovirus (PoEV) from primary cells. Pigs contain endogenous retroviral sequences encoding infectious retrovirus, yet little is known about the conditions required to activate the expression and release of PoEV from primary cells. We show here that mitogenic activation of peripheral blood mononuclear cells (PBMC) isolated from the National Institutes of Health (NIH) miniature pig and the Yucatan pig resulted in the activation and release of an infectious type C retrovirus. Coculture of activated porcine PBMC with pig or human cell lines resulted in the transfer and expression of PoEV-specific sequences and the establishment of a productive infection. Sequence comparison of portions of the PoEV pol gene expressed in pig cell lines productively infected with virus derived from NIH miniature pig and Yucatan pig PBMC revealed marked similarity, suggesting that one or a few loci may be capable of being activated to yield an infectious virus. These findings demonstrate that the presence of endogenous viruses in source animals needs to be carefully considered when the infectious disease potential of xenotransplantation is being assessed.


Asunto(s)
Gammaretrovirus/fisiología , Porcinos Enanos/virología , Porcinos/virología , Animales , Secuencia de Bases , Línea Celular , Quirópteros , Cricetinae , ADN Viral , Células HeLa , Humanos , Riñón/citología , Leucocitos Mononucleares/efectos de los fármacos , Leucocitos Mononucleares/virología , Visón , Mitógenos/farmacología , Datos de Secuencia Molecular , Fitohemaglutininas/farmacología , Ratas , Acetato de Tetradecanoilforbol/farmacología
12.
Hum Mol Genet ; 6(12): 2051-60, 1997 Nov.
Artículo en Inglés | MEDLINE | ID: mdl-9328468

RESUMEN

Fragile X mental retardation syndrome is associated with an expansion of a CGG repeat within the 5'UTR of the first exon of the FMR1 gene, abnormal methylation of the CpG island in the promoter region, and a transcriptional silencing of this gene. We studied transcriptional regulation of the FMR1 gene using protein footprint analysis of the active and inactive gene in vivo . We identified four footprints within the FMR1 promoter region which correspond to consensus binding sites of known transcription factors, alpha-PAL/NRF1, Sp1, H4TF1/Sp1-like and c-myc. These footprints were present in normal cells with a transcriptionally active FMR1 gene. The same footprints were present in different cell types: primary fibroblasts, lymphoblastoid cells and peripheral lymphocytes. However, for the 1.1 kb region analyzed, no footprints were detected in a variety of cell types derived from patients with fragile X syndrome which have a transcriptionally inactive FMR1 gene. A BLAST nucleotide search identified sequence similarities between the region of the FMR1 gene containing the footprints and an analogous region within the promoter region of the gene for the heterogeneous nuclear ribonucleoprotein (hnRNP) A2, a member of a family of ribonucleoproteins implicated in mRNA processing and nuclear-cytoplasm transport. The nucleotide sequences identified in the hnRNP-A2 promoter region correspond to the same consensus binding sites showing DNA-protein interactions in the FMR1 gene. Our previous functional studies and the studies of others demonstrate that FMR proteins, like hnRNP-A2, are also ribonucleoproteins which appear to be involved in mRNA transport. The results from our footprint studies suggest that the expression of the FMR1 gene is regulated by the binding of specific transcription factors to sequence elements in the 5' region of the gene and that this expression may be regulated by elements in common with the hnRNP-A2 gene. Common regulation of these two genes might play an important role in the cooperative processing and transport of mRNA from the nucleus to the translation machinery.


Asunto(s)
Ribonucleoproteína Heterogénea-Nuclear Grupo A-B , Proteínas del Tejido Nervioso/química , Proteínas del Tejido Nervioso/genética , Regiones Promotoras Genéticas/fisiología , Proteínas de Unión al ARN , Ribonucleoproteínas/química , Ribonucleoproteínas/genética , Femenino , Fibroblastos/metabolismo , Proteína de la Discapacidad Intelectual del Síndrome del Cromosoma X Frágil , Síndrome del Cromosoma X Frágil/genética , Ribonucleoproteínas Nucleares Heterogéneas , Humanos , Activación de Linfocitos/genética , Linfocitos/metabolismo , Masculino , Datos de Secuencia Molecular , Proteínas del Tejido Nervioso/metabolismo , Unión Proteica/genética , Secuencias Reguladoras de Ácidos Nucleicos , Ribonucleoproteínas/metabolismo , Relación Estructura-Actividad
13.
J Infect Dis ; 176(3): 775-7, 1997 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-9291331

RESUMEN

The prevalence of human herpesvirus 8 (HHV-8)/Kaposi's sarcoma (KS)-associated herpesvirus was investigated in the semen of 99 human immunodeficiency virus (HIV)-infected men (median CD4 cell count, 357/mm3) by use of a polymerase chain reaction (PCR) assay capable of detecting <10 copies of HHV-8 DNA. Of the subjects, 95 (96%) self-identified as men who have sex with men (MSM), and 3 had a history of clinical KS. Seminal cell specimens were negative for HHV-8 in 98 subjects. None of the 26 without KS (27.1% of 96 tested) who were seropositive for HHV-8 by IFA for latency-associated nuclear antigens had HHV-8 detected in their semen. The only subject with any evidence for seminal HHV-8 DNA was seropositive for HHV-8 and had active KS. HHV-8 was detected in 10 (10.4%) of 96 peripheral blood mononuclear cell specimens. The prevalence of HHV-8 DNA by PCR in semen of HIV-infected MSM without KS is low.


Asunto(s)
Infecciones Oportunistas Relacionadas con el SIDA/virología , Herpesvirus Humano 8/aislamiento & purificación , Sarcoma de Kaposi/virología , Semen/virología , Animales , ADN Viral/análisis , Drosophila melanogaster , Humanos , Masculino , Reacción en Cadena de la Polimerasa , Prevalencia , Sarcoma de Kaposi/epidemiología
14.
J Virol ; 71(5): 4138-44, 1997 May.
Artículo en Inglés | MEDLINE | ID: mdl-9094697

RESUMEN

Simian retroperitoneal fibromatosis (RF) is a vascular fibroproliferative neoplasm which has many morphological and histological similarities to human Kaposi's sarcoma (KS). Like epidemic KS in AIDS patients, RF is highly associated with an immunodeficiency syndrome (simian acquired immunodeficiency syndrome [SAIDS]) caused by a retrovirus infection. Recently, a new gammaherpesvirus, called Kaposi's sarcoma-associated herpesvirus (KSHV) or human herpesvirus 8 (HHV8), has been identified in KS tumors, suggesting that KS has a viral etiology. Our previous experimental transmission studies and epidemiological data suggest that RF also has an infectious etiology. In order to determine whether a similar virus is also associated with RF, we have assayed for the presence of an unknown herpesvirus using degenerate PCR primers targeting the highly conserved DNA polymerase genes of the herpesvirus family. Here we provide DNA sequence evidence for two new herpesviruses closely related to KSHV from RF tissues of two macaque species, Macaca nemestrina and Macaca mulatta. Our data suggest that KSHV and the putative macaque herpesviruses define a new group within the subfamily Gammaherpesvirinae whose members are implicated in the pathogenesis of KS and KS-like neoplasms in different primate species.


Asunto(s)
Fibroma/veterinaria , Herpesvirus Humano 8/aislamiento & purificación , Macaca mulatta/virología , Macaca nemestrina/virología , Enfermedades de los Monos/virología , Neoplasias Retroperitoneales/veterinaria , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Fibroma/virología , Gammaherpesvirinae/clasificación , Herpesvirus Humano 8/clasificación , Datos de Secuencia Molecular , Neoplasias Retroperitoneales/virología
15.
Yeast ; 13(15): 1409-21, 1997 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-9434347

RESUMEN

Saccharomyces carlsbergensis is an amphiploid, and it has previously been suggested that the genomes of S. carlsbergensis originate from S. cerevisiae and S. monacensis. We have cloned the ACB1 genes encoding the acyl-CoA binding protein (ACBP) from S. carlsbergensis, S. cerevisiae and S. monacensis. Two genes were found in S. carlsbergensis and named ACB1 type 1 and type 2, respectively. The type 1 gene is identical to the S. cerevisiae ACB1 gene except for three substitutions, one single base pair deletion and one double base pair insertion, all located in the promoter region. The type 2 gene is completely identical to the S. monacensis ACB1 gene. These findings substantiate the notion that S. carlsbergensis is a hybrid between S. cerevisiae and S. monacensis. Both ACB1 type 1 and type 2 are actively transcribed in S. carlsbergensis and transcription is initiated at sites identical to those used for transcriptional initiation of the ACB1 genes in S. cerevisiae and S. monacensis, respectively. Two polyadenylation sites, spaced 225 bp apart, are present in the S. cerevisiae ACB1 gene. The upstream polyadenylation site is used exclusively during exponential growth, whereas both sites are utilized during later stages of growth.


Asunto(s)
Proteínas Portadoras/genética , Proteínas Fúngicas/genética , Genes Fúngicos , Saccharomyces/genética , Secuencia de Bases , Clonación Molecular , Inhibidor de la Unión a Diazepam , Datos de Secuencia Molecular , Familia de Multigenes , Poli A/biosíntesis , Procesamiento Postranscripcional del ARN , ARN de Hongos/biosíntesis , ARN Mensajero/biosíntesis , Saccharomyces cerevisiae/genética , Análisis de Secuencia de ADN , Homología de Secuencia de Ácido Nucleico , Especificidad de la Especie , Transcripción Genética
16.
J Clin Microbiol ; 34(7): 1666-71, 1996 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-8784566

RESUMEN

A consensus primer PCR method which amplifies a region of herpesviral DNA-directed DNA polymerase (EC 2.7.7.7) and which uses degenerate primers in a nested format was developed. Primers were designed to target sequences coding for highly conserved amino acid motifs covering a region of approximately 800 bp. The assay was applied to 22 species of herpesviruses (8 human and 14 animal viruses), with PCR products obtained for 21 of 22 viruses. In the process, 14 previously unreported amino acid-coding sequences from herpesviral DNA polymerases were obtained, including regions of human herpesviruses 7 and 8. The 50 to 60 amino acid-coding sequences recovered in the present study were determined to be unique to each viral species studied, with very little sequence variation between strains of a single species when studied. Template dilution studies in the presence of human carrier DNA demonstrated that six human herpesviruses (herpesviruses 1, 2, 3, 4, 5, and 6B) could be detected at levels at or below 100 genome equivalents per 100 ng of carrier DNA. These data suggest that consensus primer PCR targeted to herpesviral DNA polymerase may prove to be useful in the detection and identification of known herpesviruses in clinical samples and the initial characterization of new herpesviral genomes.


Asunto(s)
Herpesviridae/genética , Reacción en Cadena de la Polimerasa/métodos , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Secuencia de Consenso , Cartilla de ADN/genética , ADN Viral/genética , ADN Polimerasa Dirigida por ADN/genética , Estudios de Evaluación como Asunto , Herpesviridae/clasificación , Herpesviridae/enzimología , Humanos , Datos de Secuencia Molecular , Filogenia , Homología de Secuencia de Aminoácido , Especificidad de la Especie
17.
Proc Natl Acad Sci U S A ; 92(16): 7440-4, 1995 Aug 01.
Artículo en Inglés | MEDLINE | ID: mdl-7638210

RESUMEN

Representational difference analysis was used to search for pathogens in multiple sclerosis brains. We detected a 341-nucleotide fragment that was 99.4% identical to the major DNA binding protein gene of human herpesvirus 6 (HHV-6). Examination of 86 brain specimens by PCR demonstrated that HHV-6 was present in > 70% of MS cases and controls and is thus a commensal virus of the human brain. By DNA sequencing, 36/37 viruses from MS cases and controls were typed as HHV-6 variant B group 2. Other herpesviruses, retroviruses, and measles virus were detected infrequently or not at all. HHV-6 expression was examined by immunocytochemistry with monoclonal antibodies against HHV-6 virion protein 101K and DNA binding protein p41. Nuclear staining of oligodendrocytes was observed in MS cases but not in controls, and in MS cases it was observed around plaques more frequently than in uninvolved white matter. MS cases showed prominent cytoplasmic staining of neurons in gray matter adjacent to plaques, although neurons expressing HHV-6 were also found in certain controls. Since destruction of oligodendrocytes is a hallmark of MS, these studies suggest an association of HHV-6 with the etiology or pathogenesis of MS.


Asunto(s)
Herpesvirus Humano 6/genética , Herpesvirus Humano 6/aislamiento & purificación , Esclerosis Múltiple/virología , Adulto , Antígenos Virales/metabolismo , Secuencia de Bases , Cartilla de ADN/genética , ADN Viral/genética , ADN Viral/aislamiento & purificación , Infecciones por Herpesviridae/complicaciones , Infecciones por Herpesviridae/patología , Infecciones por Herpesviridae/virología , Herpesvirus Humano 6/patogenicidad , Humanos , Inmunohistoquímica , Datos de Secuencia Molecular , Esclerosis Múltiple/etiología , Esclerosis Múltiple/patología , Oligodendroglía/virología , Reacción en Cadena de la Polimerasa
18.
Genomics ; 25(2): 469-76, 1995 Jan 20.
Artículo en Inglés | MEDLINE | ID: mdl-7789980

RESUMEN

The acyl-CoA binding protein (ACBP) and the diazepam binding inhibitor (DBI) or endozepine are independent isolates of a single 86-amino-acid, 10-kDa protein. ACBP/DBI is highly conserved between species and has been identified in several diverse organisms, including human, cow, rat, frog, duck, insects, plants, and yeast. Although the genomic locus has not yet been cloned in humans, complementary DNA clones with different 5' ends have been isolated and characterized. These cDNA clones appear to be encoded by a single gene. However, Southern blot analyses, in situ hybridizations, and somatic cell hybrid chromosomal mapping all suggest that there are multiple ACBP/DBI-related sequences in the genome. To identify potential members of this gene family, degenerate oligonucleotides corresponding to highly conserved regions of ACBP/DBI were used to screen a human genomic DNA library using the polymerase chain reaction. A novel gene, DBIP1, that is closely related to ACBP/DBI but is clearly distinct was identified. DBIP1 bears extensive sequence homology to ACBP/DBI but lacks the introns predicted by rat and duck genomic sequence studies. A 1-base deletion in the coding region results in a frameshift and, along with the absence of introns and the lack of a detectable transcript, suggests that DBIP1 is a pseudogene. ACBP/DBI has previously been mapped to chromosome 2, although this was recently disputed, and a chromosome 6 location was suggested. We show that ACBP/DBI is correctly placed on chromosome 2 and that the gene identified on chromosome 6 is DBIP1.


Asunto(s)
Proteínas Portadoras/genética , Cromosomas Humanos Par 6 , Seudogenes , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Mapeo Cromosómico , Cromosomas Humanos Par 2 , Clonación Molecular , Cricetinae , ADN Complementario/genética , Inhibidor de la Unión a Diazepam , Patos/genética , Biblioteca de Genes , Humanos , Células Híbridas , Datos de Secuencia Molecular , Reacción en Cadena de la Polimerasa , Ratas/genética , Homología de Secuencia , Especificidad de la Especie
19.
Biochem J ; 302 ( Pt 2): 479-85, 1994 Sep 01.
Artículo en Inglés | MEDLINE | ID: mdl-8093000

RESUMEN

Acyl-CoA-binding protein (ACBP) is a 10 kDa protein characterized in vertebrates. We have isolated two ACBP homologues from the yeast Saccharomyces carlsbergensis, named yeast ACBP types 1 and 2. Both proteins contain 86 amino acid residues and are identical except for four conservative substitutions. In comparison with human ACBP, yeast ACBPs exhibit 48% (type 1) and 49% (type 2) conservation of amino acid residues. The amino acid sequence of S. carlsbergensis ACBP type 1 was found to be identical with the one ACBP present in Saccharomyces cerevisiae. A recombinant form of this protein was expressed in Escherichia coli and S. cerevisiae, purified, and its acyl-CoA-binding properties were characterized by isoelectric focusing and microcalorimetric analyses. The yeast ACBP was found to bind acyl-CoA esters with high affinity (Kd 0.55 x 10(-10) M). Overexpression of yeast ACBP in S. cerevisiae resulted in a significant expansion of the intracellular acyl-CoA pool. Finally, Southern-blotting analysis of the two genes encoding ACBP types 1 and 2 in S. carlsbergensis strongly indicated that this species is a hybrid between S. cerevisiae and Saccharomyces monacensis.


Asunto(s)
Acilcoenzima A/metabolismo , Proteínas Portadoras/metabolismo , Saccharomyces/metabolismo , Secuencia de Aminoácidos , Secuencia de Bases , Southern Blotting , Calorimetría , Proteínas Portadoras/química , Proteínas Portadoras/genética , Inhibidor de la Unión a Diazepam , Espectroscopía de Resonancia por Spin del Electrón , Electroforesis en Gel de Poliacrilamida , Escherichia coli/genética , Genes Fúngicos , Genoma Fúngico , Humanos , Punto Isoeléctrico , Espectrometría de Masas , Datos de Secuencia Molecular , Proteínas Recombinantes/química , Proteínas Recombinantes/metabolismo , Saccharomyces/química , Saccharomyces/genética , Saccharomyces cerevisiae/química , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Alineación de Secuencia
20.
DNA Cell Biol ; 13(6): 669-78, 1994 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-8024707

RESUMEN

A computer-aided homology search of the GenBank nucleotide database using the amino acid sequence of human acyl CoA-binding protein (ACBP)/diazepam-binding inhibitor (DBI)-endozepine as a probe revealed that a genomic fragment containing the gene encoding the mallard duck (Anas platyrhynchos) S-acyl fatty acid synthase thioesterase also contains sequences which encode the duck homolog of ACBP/DBI. The duck ACBP/DBI gene is positioned downstream of the thioesterase gene in a tail-to-tail orientation separated from the 3' end of the thioesterase gene by only several hundred nucleotides. Three exons were identified that have strong homology to the published cDNA sequences of human and bovine ACBP/DBI. These exons define all of the coding region except for the amino-terminal domain, which was subsequently cloned by polymerase chain reaction (PCR) amplification. The encoded amino acid sequence of the duck ACBP/DBI is 62-68% homologous to mammalian ACBP/DBI sequences. While the mammalian ACBP/DBI is expressed mainly in the liver, with smaller amounts in the brain and heart, mRNA transcripts of duck ACBP/DBI were detected only in the brain with no evidence for expression in the liver or heart. The close proximity of the genes for ACBP/DBI and S-acyl fatty acid synthase thioesterase raises the possibility of co-regulation of expression.


Asunto(s)
Proteínas Portadoras/genética , Tioléster Hidrolasas/genética , Secuencia de Aminoácidos , Animales , Secuencia de Bases , Northern Blotting , Bovinos , Clonación Molecular , ADN , Inhibidor de la Unión a Diazepam , Patos , Humanos , Datos de Secuencia Molecular , Reacción en Cadena de la Polimerasa , Homología de Secuencia de Aminoácido
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