1,720,961 research outputs found
Wilms' tumor suppressor gene (WT1) loss in T-cell acute lymphoblastic leukemia promotes cell survival and resistance to DNA damage
Full text linkT-cell acute lymphoblastic leukemia (T-ALL) is an aggressive hematologic tumor, resulting from the transformation of T-cell progenitors. Thanks to advances in molecular techniques, many alterations have been identified in T-ALL cells opening new opportunities for targeted therapy. WT1 gene deletions and mutations have been reported in 10-12% of T-ALL patients, but the mechanisms downstream of WT1 alterations in T-ALL have not been elucidated. The WT1 gene encodes a zinc-finger transcription factor which is characterized by multiple alternative isoforms. The isoforms that lack the three amino acids lysine-threonine-serine (KTS') between zinc finger 3 and 4, are conserved throughout vertebrate evolution and have high DNA-binding affinity and transcriptional activity. Most of WT1 mutations found in T-ALL are heterozygous frameshifts in exon 7 predicted to produce a truncated protein which lacks the DNA binding domain. Our main hypothesis is that WT1 acts as a haplo-insufficient tumor suppressor gene in T-ALL and that WT1 loss in T-ALL leads to de-regulation of pathway in T-ALL. In this study, we first analyzed the effects of full-length and mutant WT1 isoform over-expression on the survival and proliferation of T-ALL cells. We observed that only the (KTS') isoforms negatively affected growth of T-ALL and impaired colony formation in soft-agar. Importantly, the truncated WT1 proteins, derived from a characteristic frameshift mutation in exon 7 (E384Stop), had no effects. In parallel, we also analyzed the effects of WT1 loss in T-ALL cells. We found that WT1 knockdown in MOLT4 cells significantly increased the number of colonies in clonogenic assays in comparison with control cells. Overall these results indicated that WT1 most probably works as an haplo-insufficient tumor suppressor gene in T-ALL. In order to evaluate if mutations in WT1 locus are most likely responsible for an impaired transcriptional program we mainly focused on the analysis of WT1 deregulated targets following WT1 loss in T-ALL cells. To define the structure of the transcriptional network activated by loss of function of WT1, we performed ChIP-chip and gene expression analysis in MOLT4 T-ALL cells. ChIP-chip analysis showed that WT1 direct targets were enriched in pathways responsible for cellular response to stress, such as p53, nucleotide excision repair and Mitogen-Activated Protein Kinases (MAPK) signalling pathways. Integration of ChIP-chip data with gene expression analysis performed under WT1 loss of function conditions in MOLT4 cells provided an enrichment in the MAPK pathway. Stemming from these results, we finally evaluated if the loss of WT1 conferred increased survival after DNA damage, such as ionizing radiation or chemotherapeutic drugs, in MOLT4 T-ALL cells and primary T-ALL xenografts. Analysis of cell viability and apoptosis showed that WT1 alterations induced increased survival following DNA-damaging conditions, mainly affecting directly the transcription of important mediators of p53 apoptotic response. A master regulator of these effects was BBC3/PUMA, whose induction was augmented in the presence of both WT1 and p53 proteins. In conclusion, analyzing WT1 loss in T-ALL cells we determined a deregulation of several genes involved in the pathogenesis of T-ALL, in particular genes responsible for cellular response to stress, strongly suggesting WT1 acts as tumor suppressor gene in T-ALL cells
Calcineurin complex isolated from T-cell acute lymphoblastic leukemia (T-ALL) cells identifies new signaling pathways including mTOR/AKT/S6K whose inhibition synergize with Calcineurin inhibition to promote T-ALL cell death
No full textCalcineurin (Cn) is a calcium activated protein phosphatase involved in many aspects of normal T cell physiology, however the role of Cn and/or its downstream targets in leukemogenesis are still ill-defined. In order to identify putative downstream targets/effectors involved in the pro-oncogenic activity of Cn in T-cell acute lymphoblastic leukemia (T-ALL) we used tandem affinity chromatography, followed by mass spectrometry to purify novel Cn-interacting partners. We found the Cn-interacting proteins to be part of numerous cellular signaling pathways including eIF2 signaling and mTOR signaling. Coherently, modulation of Cn activity in T-ALL cells determined alterations in the phosphorylation status of key molecules implicated in protein translation such as eIF-2α and ribosomal protein S6. Joint targeting of PI3K-mTOR, eIF-2α and 14-3-3 signaling pathways with Cn unveiled novel synergistic pro-apoptotic drug combinations. Further analysis disclosed that the synergistic interaction between PI3K-mTOR and Cn inhibitors was prevalently due to AKT inhibition. Finally, we showed that the synergistic pro-apoptotic response determined by jointly targeting AKT and Cn pathways was linked to down-modulation of key anti-apoptotic proteins including Mcl-1, Claspin and XIAP. In conclusion, we identify AKT inhibition as a novel promising drug combination to potentiate the pro-apoptotic effects of Cn inhibitors
Time course and temperature dependence of the membrane translocation of tetanus and botulinum neurotoxins C and D in neurons
No full textCalcineurin and GSK-3 inhibition sensitizes T-cell acute lymphoblastic leukemia cells to apoptosis through X-linked inhibitor of apoptosis protein degradation
No full textThe calcineurin (Cn)-nuclear factor of activated T cells signaling pathway is critically involved in many aspects of normal T-cell physiology; however, its direct implication in leukemogenesis is still ill-defined. Glycogen synthase kinase-3 beta (GSK-3 beta) has recently been reported to interact with Cn in neuronal cells and is implicated in MLL leukemia. Our biochemical studies clearly demonstrated that Cn was able to interact with GSK-3 beta in T-cell acute lymphoblastic leukemia (T-ALL) cells, and that this interaction was direct, leading to an increased catalytic activity of GSK-3 beta, possibly through autophosphorylation of Y216. Sensitivity to GSK-3 inhibitor treatment correlated with altered GSK-3 beta phosphorylation and was more prominent in T-ALL with Pre/Pro immunophenotype. In addition, dual Cn and GSK-3 inhibitor treatment in T-ALL cells promoted sensitization to apoptosis through proteasomal degradation of X-linked inhibitor of apoptosis protein (XIAP). Consistently, resistance to drug treatments in primary samples was strongly associated with higher XIAP protein levels. Finally, we showed that dual Cn and GSK-3 inhibitor treatment in vitro and in vivo is effective against available models of T-ALL, indicating an insofar untapped therapeutic opportunity
The thioredoxin reductase-thioredoxin system is involved in the entry of tetanus and botulinum neurotoxins in the cytosol of nerve terminals
No full textTetanus and botulinum neurotoxins cause paralysis by cleaving SNARE proteins within the cytosol of nerve terminals. They are endocytosed inside acidic vesicles and the pH gradient across the membrane drives the translocation of their metalloprotease L domain in the cytosol. This domain is linked to the rest of the molecule by a single interchain disulfide bridge that has to be reduced on the cytosolic side of the membrane to free its enzymatic activity. By using specific inhibitors of the various cytosolic protein disulfides reducing systems, we show here that the NADPH-thioredoxin reductase-thioredoxin redox system is the main responsible for this disulfide reduction. In addition, we indicate auranofin, as a possible basis for the design of novel inhibitors of these neurotoxins. © 2012 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved
Going Beyond Counting First Authors in Author Co-citation Analysis
Full text linkThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
Variations on the Author
Full text link“Variations on the Author” discusses two of Eduardo Coutinho’s recent films (Um Dia na Vida, from 2010, and Últimas Conversas, posthumously released in 2015) and their contribution to the general question of documentary authorship. The director’s filmography is characterized by a consistent yet self-effacing form of authorial self-inscription: Coutinho often features as an interviewer that rather than express opinions propels discourses; an interviewer that is good at listening. This mode of self-inscription characterizes him as an author who is not expressive but who is nonetheless markedly present on the screen. In Um Dia na Vida, however, Coutinho is completely absent form the image, while Últimas Conversas, on the contrary, includes a confessional prologue that moves the director from the margins to the center of his films. This article examines the ways in which these works stand out in the filmography of a director who offers new insights into the notion of cinematic authorship
Appropriate Similarity Measures for Author Cocitation Analysis
Full text linkWe provide a number of new insights into the methodological discussion about author cocitation analysis. We first argue that the use of the Pearson correlation for measuring the similarity between authors’ cocitation profiles is not very satisfactory. We then discuss what kind of similarity measures may be used as an alternative to the Pearson correlation. We consider three similarity measures in particular. One is the well-known cosine. The other two similarity measures have not been used before in the bibliometric literature. Finally, we show by means of an example that our findings have a high practical relevance.information science;Pearson correlation;cosine;similarity measure;author cocitation analysis
Dispelling the Myths Behind First-author Citation Counts
Full text linkWe conducted a full-scale evaluative citation analysis study of scholars in the XML research field to explore just how different from each other author rankings resulting from different citation counting methods actually are, and to demonstrate the capability of emerging data and tools on the Web in supporting more realistic citation counting methods. Our results contest some common arguments for the continued
use of first-author citation counts in the evaluation of scholars, such as high correlations between author rankings by first-author citation counts and other citation
counting methods, and high costs of using more realistic citation counting methods that are not well-supported by the ISI databases. It is argued that increasingly available digital full text research papers make it possible for citation analysis studies to go beyond what the ISI databases have directly supported and to employ more
sophisticated methods
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