The Oncoprotein EVI1 and the DNA Methyltransferase Dnmt3 Co-Operate in Binding and De Novo Methylation of Target DNA

EVI1 has pleiotropic functions during murine embryogenesis and its targeted disruption leads to prenatal death by severely affecting the development of virtually all embryonic organs. However, its functions in adult tissues are still unclear. When inappropriately expressed, EVI1 becomes one of the most aggressive oncogenes associated with human hematopoietic and solid cancers. The mechanisms by which EVI1 transforms normal cells are unknown, but we showed recently that EVI1 indirectly upregulates self-renewal and cell-cycling genes by inappropriate methylation of CpG dinucleotides in the regulatory regions of microRNA-124-3 (miR-124-3), leading to the repression of this small gene that controls normal differentiation and cell cycling of somatic cells. We used the regulatory regions of miR-124-3 as a read-out system to investigate how EVI1 induces de novo methylation of DNA. Here we show that EVI1 physically interacts with DNA methyltransferases 3a and 3b (Dnmt3a/b), which are the only de novo DNA methyltransferases identified to date in mouse and man, and that it forms an enzymatically active protein complex that induces de novo DNA methylation in vitro. This protein complex targets and binds to a precise region of miR-124-3 that is necessary for repression of a reporter gene by EVI1. Based on our findings, we propose that in cooperation with Dnmt3a/b EVI1 regulates the methylation of DNA as a sequence-specific mediator of de novo DNA methylation and that inappropriate EVI1 expression contributes to carcinogenesis through improper DNA methylation

Amyloid-Mediated Sequestration of Essential Proteins Contributes to Mutant Huntingtin Toxicity in Yeast

BACKGROUND: Polyglutamine expansion is responsible for several neurodegenerative disorders, among which Huntington disease is the most well-known. Studies in the yeast model demonstrated that both aggregation and toxicity of a huntingtin (htt) protein with an expanded polyglutamine region strictly depend on the presence of the prion form of Rnq1 protein ([PIN+]), which has a glutamine/asparagine-rich domain. PRINCIPAL FINDINGS: Here, we showed that aggregation and toxicity of mutant htt depended on [PIN+] only quantitatively: the presence of [PIN+] elevated the toxicity and the levels of htt detergent-insoluble polymers. In cells lacking [PIN+], toxicity of mutant htt was due to the polymerization and inactivation of the essential glutamine/asparagine-rich Sup35 protein and related inactivation of another essential protein, Sup45, most probably via its sequestration into Sup35 aggregates. However, inhibition of growth of [PIN+] cells depended on Sup35/Sup45 depletion only partially, suggesting that there are other sources of mutant htt toxicity in yeast. CONCLUSIONS: The obtained data suggest that induced polymerization of essential glutamine/asparagine-rich proteins and related sequestration of other proteins which interact with these polymers represent an essential source of htt toxicity

Exclusion of mutations in the PRNP, JPH3, TBP, ATN1, CREBBP, POU3F2 and FTL genes as a cause of disease in Portuguese patients with a Huntington-like phenotype

Huntington disease (HD) is an autosomal dominant neurodegenerative disorder characterised by chorea, cognitive impairment, dementia and personality changes, caused by the expansion of a CAG repeat in the HD gene. Often, patients with a similar clinical presentation do not carry expansions of the CAG repeat in this gene [Huntington disease-like (HDL) patients]. We report the genetic analysis of 107 Portuguese patients with an HDL phenotype. The HDL genes PRNP and JPH3, encoding the prion protein and junctophilin-3, respectively, were screened for repeat expansions in these patients. Given the partial clinical overlap of SCA17, DRPLA and neuroferritinopathy with HD, their causative genes (TBP, ATN1, and FTL, respectively) were also analysed. Finally, repeat expansions in two candidate genes, CREBBP and POU3F2, which encode the nuclear transcriptional coactivator CREB-binding protein and the CNS-specific transcription factor N-Oct-3, respectively, were also studied. Expansions of the repetitive tracts of the PRNP, JPH3, TBP, ATN1, CREBBP and POU3F2 genes were excluded in all patients, as were sequence alterations in the FTL gene. Since none of the genes already included in the differential diagnosis of HD was responsible for the disease in our sample, the genetic heterogeneity of the HDL phenotype is still open for investigation.Fundação para a Ciência e a Tecnologia (FCT) and FEDER (grant CBO/33485/99). BIC included in grant CBO/33485/99, respectivel

Role of CBP and SATB-1 in Aging, Dietary Restriction, and Insulin-Like Signaling

Increased transcriptional complex activity, or pharmacological mimics of increased complex activity, predict lifespan in mice and mediate the protective effects of dietary restriction during aging

Duration of viremia and fecal shedding of the virus in hepatitis A infected children

La infección por el virus de hepatitis A (HAV) es endémica en Argentina. El uso de técnicas moleculares permitió extender la detección del RNA del HAV en suero y heces en pacientes con diferentes presentaciones clínicas. Comparamos la sensibilidad del protocolo de RT-PCR que usamos con cebadores dirigidos a distintas regiones del genoma, resultando la detección de la región VP3 C terminal la más sensible. Se obtuvieron prospectivamente muestras de suero y materia fecal de 20 niños con hepatitis aguda autolimitada por HAV. El RNA del HAV fue detectado en 18/20 niños en muestras basales y en 19/20 sumando una muestra posterior. El RNA del HAV fue detectable en 9/20 pacientes hasta 30 días en suero; en materia fecal en 2/20 hasta 60 días y en 1/20 hasta 90 días. La secuencia genómica para la región VP1/2A en 8 muestras demostró que todas pertenecían al subgenotipo IA, aunque eran diferentes entre sí. Solo en 1/11 niños con falla hepática fulminante fue posible la detección del RNA del HAV utilizando la región VP3 C terminal y el genotipo fue I. La reciente introducción de la vacunación universal en niños de 1 año de edad en Argentina podría disminuir drásticamente la circulación del virus emergiendo nuevas fuentes de infección y permitiendo la introducción de nuevos genotipos. Las técnicas moleculares aplicadas al estudio de la historia natural de la infección y a la vigilancia epidemiológica contribuyen al control y la toma de decisiones eficientes en políticas de Salud Pública.Hepatitis A virus (HAV) infection is endemic in Argentina. Molecular tools have allowed HAV RNA detection to be extent to sera and feces from patients with different clinical backgrounds. We compare the sensitivity of the RT-PCR protocol we follow using primers targeting different genomic regions and VP3 C terminal was the most sensitive. Sequential sera and fecal samples were obtained from 20 children with acute self limited Hepatitis A. HAV RNA was detectable in 18/20 children if sera and stool specimens were collected at the onset of symptoms and in 19/20 if a later sample was considered. HAV RNA was detectable in serum from 9/20 patients until day 30 and in feces from 2 patients until day 60 and until day 90 in one. Genomic sequences from VP1/2A region in 8 samples showed they all belong to subgenotype IA although they were different between them. HAV RNA was detectable only in 1/11 sera from children with acute liver failure when VP3 C terminal fragment was searched and it belonged to genotype I. Universal vaccination in one year old children was recently implemented in Argentina and it will dramatically enable the decrease of the viral circulation, making new sources of infection emerge and allowing the introduction of new genotypes. The application of molecular tools to the study of the natural history of infection and to the epidemiologic surveillance may contribute to efficient control and lead to rational decisions in public health policies.Fil: Munné, María Silvina. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Laboratorio Nacional de Referencia de Hepatitis Virales; Argentina.Fil: Cañero Velasco, María C. Hospital Municipal del Niño de San Justo. Unidad de Gastroenterología y Hepatología; Argentina.Fil: Moreiro, Rita. Hospital Nacional de Pediatría J. P. Garrahan. Laboratorio; Argentina.Fil: Vladimirsky, Sara. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Laboratorio Nacional de Referencia de Hepatitis Virales; Argentina.Fil: Otegui, Lucio. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Laboratorio Nacional de Referencia de Hepatitis Virales; Argentina.Fil: Castro, Raúl. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Laboratorio Nacional de Referencia de Hepatitis Virales; Argentina.Fil: Brajterman, Leonardo. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Laboratorio Nacional de Referencia de Hepatitis Virales; Argentina.Fil: Soto, Sonia. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Laboratorio Nacional de Referencia de Hepatitis Virales; Argentina.Fil: Mutti, Jorge. Hospital Municipal del Niño de San Justo. Unidad de Gastroenterología y Hepatología; Argentina.Fil: Nucifora, Silvia. Hospital Municipal del Niño de San Justo. Unidad de Gastroenterología y Hepatología; Argentina.Fil: Lara, Elena. Hospital Municipal del Niño de San Justo. Laboratorio Central; Argentina.Fil: Sosa, Anibal. Hospital Municipal del Niño de San Justo. Laboratorio Central; Argentina.Fil: Godoy, Patricia. Hospital Municipal del Niño de San Justo. Unidad de Gastroenterología y Hepatología; Argentina.Fil: Ciocca, Mirta. Hospital Nacional de Pediatría J. P. Garrahan. Unidad de Hígado; Argentina.Fil: Cuarterolo, Miriam. Hospital Nacional de Pediatría J. P. Garrahan. Unidad de Hígado; Argentina.Fil: Quarleri, Jorge F. Facultad de Medicina. Centro Nacional de Referencia para el SIDA; Argentina.Fil: González, Jorge E. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Laboratorio Nacional de Referencia de Hepatitis Virales; Argentina

Duration of viremia and fecal shedding of the virus in hepatitis A infected children

La infección por el virus de hepatitis A (HAV) es endémica en Argentina. El uso de técnicas moleculares permitió extender la detección del RNA del HAV en suero y heces en pacientes con diferentes presentaciones clínicas. Comparamos la sensibilidad del protocolo de RT-PCR que usamos con cebadores dirigidos a distintas regiones del genoma, resultando la detección de la región VP3 C terminal la más sensible. Se obtuvieron prospectivamente muestras de suero y materia fecal de 20 niños con hepatitis aguda autolimitada por HAV. El RNA del HAV fue detectado en 18/20 niños en muestras basales y en 19/20 sumando una muestra posterior. El RNA del HAV fue detectable en 9/20 pacientes hasta 30 días en suero; en materia fecal en 2/20 hasta 60 días y en 1/20 hasta 90 días. La secuencia genómica para la región VP1/2A en 8 muestras demostró que todas pertenecían al subgenotipo IA, aunque eran diferentes entre sí. Solo en 1/11 niños con falla hepática fulminante fue posible la detección del RNA del HAV utilizando la región VP3 C terminal y el genotipo fue I. La reciente introducción de la vacunación universal en niños de 1 año de edad en Argentina podría disminuir drásticamente la circulación del virus emergiendo nuevas fuentes de infección y permitiendo la introducción de nuevos genotipos. Las técnicas moleculares aplicadas al estudio de la historia natural de la infección y a la vigilancia epidemiológica contribuyen al control y la toma de decisiones eficientes en políticas de Salud Pública.Hepatitis A virus (HAV) infection is endemic in Argentina. Molecular tools have allowed HAV RNA detection to be extent to sera and feces from patients with different clinical backgrounds. We compare the sensitivity of the RT-PCR protocol we follow using primers targeting different genomic regions and VP3 C terminal was the most sensitive. Sequential sera and fecal samples were obtained from 20 children with acute self limited Hepatitis A. HAV RNA was detectable in 18/20 children if sera and stool specimens were collected at the onset of symptoms and in 19/20 if a later sample was considered. HAV RNA was detectable in serum from 9/20 patients until day 30 and in feces from 2 patients until day 60 and until day 90 in one. Genomic sequences from VP1/2A region in 8 samples showed they all belong to subgenotype IA although they were different between them. HAV RNA was detectable only in 1/11 sera from children with acute liver failure when VP3 C terminal fragment was searched and it belonged to genotype I. Universal vaccination in one year old children was recently implemented in Argentina and it will dramatically enable the decrease of the viral circulation, making new sources of infection emerge and allowing the introduction of new genotypes. The application of molecular tools to the study of the natural history of infection and to the epidemiologic surveillance may contribute to efficient control and lead to rational decisions in public health policies.Fil: Munné, María Silvina. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Laboratorio Nacional de Referencia de Hepatitis Virales; Argentina.Fil: Cañero Velasco, María C. Hospital Municipal del Niño de San Justo. Unidad de Gastroenterología y Hepatología; Argentina.Fil: Moreiro, Rita. Hospital Nacional de Pediatría J. P. Garrahan. Laboratorio; Argentina.Fil: Vladimirsky, Sara. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Laboratorio Nacional de Referencia de Hepatitis Virales; Argentina.Fil: Otegui, Lucio. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Laboratorio Nacional de Referencia de Hepatitis Virales; Argentina.Fil: Castro, Raúl. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Laboratorio Nacional de Referencia de Hepatitis Virales; Argentina.Fil: Brajterman, Leonardo. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Laboratorio Nacional de Referencia de Hepatitis Virales; Argentina.Fil: Soto, Sonia. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Laboratorio Nacional de Referencia de Hepatitis Virales; Argentina.Fil: Mutti, Jorge. Hospital Municipal del Niño de San Justo. Unidad de Gastroenterología y Hepatología; Argentina.Fil: Nucifora, Silvia. Hospital Municipal del Niño de San Justo. Unidad de Gastroenterología y Hepatología; Argentina.Fil: Lara, Elena. Hospital Municipal del Niño de San Justo. Laboratorio Central; Argentina.Fil: Sosa, Anibal. Hospital Municipal del Niño de San Justo. Laboratorio Central; Argentina.Fil: Godoy, Patricia. Hospital Municipal del Niño de San Justo. Unidad de Gastroenterología y Hepatología; Argentina.Fil: Ciocca, Mirta. Hospital Nacional de Pediatría J. P. Garrahan. Unidad de Hígado; Argentina.Fil: Cuarterolo, Miriam. Hospital Nacional de Pediatría J. P. Garrahan. Unidad de Hígado; Argentina.Fil: Quarleri, Jorge F. Facultad de Medicina. Centro Nacional de Referencia para el SIDA; Argentina.Fil: González, Jorge E. ANLIS Dr.C.G.Malbrán. Instituto Nacional de Enfermedades Infecciosas. Laboratorio Nacional de Referencia de Hepatitis Virales; Argentina

A meta-review of evidence on heart failure disease management programs: the challenges of describing and synthesizing evidence on complex interventions

Background: Despite favourable results from past meta-analyses, some recent large trials have not found Heart Failure (HF) disease management programs to be beneficial. To explore reasons for this, we evaluated evidence from existing meta-analyses. Methods: Systematic review incorporating meta-review was used. We selected meta-analyses of randomized controlled trials published after 1995 in English that examined the effects of HF disease management programs on key outcomes. Databases searched: MEDLINE, EMBASE, Cochrane Database of Systematic Reviews (CDSR), DARE, NHS EED, NHS HTA, Ageline, AMED, Scopus, Web of Science and CINAHL; cited references, experts and existing reviews were also searched. Results: 15 meta-analyses were identified containing a mean of 18.5 randomized trials of HF interventions +/- 10.1 (range: 6 to 36). Overall quality of the meta-analyses was very mixed (Mean AMSTAR Score = 6.4 +/- 1.9; range 2-9). Reporting inadequacies were widespread around populations, intervention components, settings and characteristics, comparison, and comparator groups. Heterogeneity (statistical, clinical, and methodological) was not taken into account sufficiently when drawing conclusions from pooled analyses. Conclusions: Meta-analyses of heart failure disease management programs have promising findings but often fail to report key characteristics of populations, interventions, and comparisons. Existing reviews are of mixed quality and do not adequately take account of program complexity and heterogeneity