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Wikipedia's influence in shaping public perceptions of science underscores the significance of scientists being recognized on the platform, as it can impact their careers. Although Wikipedia offers guidelines for determining when a scientist qualifies for their own article, it currently lacks guidance regarding whether a scientist should be acknowledged in articles related to the innovation processes to which they have contributed. To explore how Wikipedia addresses this issue of scientific "micro-notability," we introduce a digital method called Name Edit Analysis, enabling us to quantitatively and qualitatively trace mentions of scientists within Wikipedia's articles. We study two CRISPR-related Wikipedia articles and find dynamic negotiations of micro-notability as well as a surprising tension between Wikipedia's principle of safeguarding against self-promotion and the scholarly norm of "due credit." To reconcile this tension, we propose that Wikipedians and scientists collaborate to establish specific micro-notability guidelines that acknowledge scientific contributions while preventing excessive self-promotion.
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This paper analyzes Wikipedia's representation of the Nobel Prize winning CRISPR/Cas9 technology, a method for gene editing. We propose and evaluate different heuristics to match publications from several publication corpora against Wikipedia's central article on CRISPR and against the complete Wikipedia revision history in order to retrieve further Wikipedia articles relevant to the topic and to analyze Wikipedia's referencing patterns. We explore to what extent the selection of referenced literature of Wikipedia's central article on CRISPR adheres to scientific standards and inner-scientific perspectives by assessing its overlap with (1) the Web of Science (WoS) database, (2) a WoS-based field-delineated corpus, (3) highly-cited publications within this corpus, and (4) publications referenced by field-specific reviews. We develop a diachronic perspective on citation latency and compare the delays with which publications are cited in relevant Wikipedia articles to the citation dynamics of these publications over time. Our results confirm that a combination of verbatim searches by title, DOI, and PMID is sufficient and cannot be improved significantly by more elaborate search heuristics. We show that Wikipedia references a substantial amount of publications that are recognized by experts and highly cited, but that Wikipedia also cites less visible literature, and, to a certain degree, even not strictly scientific literature. Delays in occurrence on Wikipedia compared to the publication years show (most pronounced in case of the central CRISPR article) a dependence on the dynamics of both the field and the editor's reaction to it in terms of activity.
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Carcinoma Epitelial de Ovario , Neoplasias Ováricas , Femenino , Humanos , Incidencia , Sistema de Registros , SuizaRESUMEN
OBJECTIVE: The methods used to diagnose and classify ovarian cancer have changed over the past decade. We used hospital registry data to assess the incidence, treatment durations and hospital costs of ovarian cancer in Switzerland. METHODS: We carried out a retrospective analysis of a hospital registry covering all inpatient care episodes in Switzerland between 1998 and 2012. Ovarian cancer incidence was assessed by identifying patients with a first ovarian cancer diagnosis as the main reason for hospital stay after an event-free period. We assessed the duration and cost of ovarian cancer treatment sequences as well as the evolution of hospital patient volume over time. RESULTS: The average age-adjusted incidence rate was 14.6 per 100,000 women per year between 2004 and 2012. This rate is substantially higher (+35.5%) than the corresponding rate published by the National Institute for Cancer Epidemiology and Registration (NICER). Hospital patient volume was low in most cases, with more than 40% of patients treated in hospitals with fewer than 20 cases per year. However, the share of patients treated in hospitals with more than 30 cases per year has increased substantially since 2009. CONCLUSIONS: We found a substantial difference between the ovarian cancer incidence estimate based on hospital registry data and the corresponding estimate by NICER. The reasons for this substantial difference should be carefully explored. A case-wise comparison could determine whether the difference is due to over- or under-reporting in one of the two registries. The low ovarian cancer patient volume in many hospitals is in conflict with the numbers required for certified specialised cancer centres. The recent increase in patient volume in specialised cancer centres, however, might reflect a growing understanding of the needs and requirements of comprehensive cancer care.
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Carcinoma Epitelial de Ovario/epidemiología , Carcinoma Epitelial de Ovario/cirugía , Tiempo de Internación/estadística & datos numéricos , Neoplasias Ováricas/epidemiología , Neoplasias Ováricas/cirugía , Sistema de Registros , Carcinoma Epitelial de Ovario/terapia , Quimioterapia Adyuvante , Femenino , Hospitales/estadística & datos numéricos , Humanos , Incidencia , Tiempo de Internación/economía , Persona de Mediana Edad , Neoplasias Ováricas/terapia , Estudios Retrospectivos , Suiza/epidemiologíaRESUMEN
Retractions of scientific articles are becoming the most relevant institution for making sense of scientific misconduct. An increasing number of retracted articles, mainly attributed to misconduct, is currently providing a new empirical basis for research about scientific misconduct. This article reviews the relevant research literature from an interdisciplinary context. Furthermore, the results from these studies are contextualized sociologically by asking how scientific misconduct is made visible through retractions. This study treats retractions as an emerging institution that renders scientific misconduct visible, thus, following up on the sociology of deviance and its focus on visibility. The article shows that retractions, by highlighting individual cases of misconduct and general policies for preventing misconduct while obscuring the actors and processes through which retractions are effected, produce highly fragmented patterns of visibility. These patterns resemble the bifurcation in current justice systems.
Le retrait d'articles scientifiques après publication est devenu le principal instrument pour mesurer l'ampleur de la fraude scientifique. L'augmentation des cas de retrait d'article, essentiellement pour des raisons de fraude, fournit une nouvelle base empirique pour analyser la fraude scientifique. Cet article se propose de passer en revue la littérature scientifique traitant ce sujet dans un contexte interdisciplinaire. Il contextualise les résultats de cette étude dans le champ sociologique en s'interrogeant sur le mécanisme de dévoilement de la fraude. Il considère les retraits d'article comme un nouvel instrument de révélation de la fraude qui insiste sur la notion de visibilité dans une perspective sociologique de la déviance. En mettant l'accent sur les cas individuels et les politiques de prévention des fraudes tout en faisant l'impasse sur les acteurs et les procédures de retrait des articles, ce processus produit un espace fragmenté de visibilité. En cela, il s'apparente à la séparation des questions judiciaires (bifurcation) dans les décisions de justice.
Las retracciones de artículos científicos se están convirtiendo en la institución más relevante para dar sentido a la mala conducta científica. Un número creciente de artículos retractados, mayormente debido a la mala conducta, está proporcionando una nueva base empírica para la investigación sobre la mala conducta científica. Este artículo revisa la literatura de investigación relevante desde un contexto interdisciplinario. Además, los resultados de estos estudios se contextualizan sociológicamente preguntando cómo la mala conducta científica se hace visible a través de retracciones. Estamos tratando a retracciones como institución emergente que vuelve visible a la mala conducta científica, por lo tanto, seguimos a la sociología de la desviación y su enfoque en la visibilidad. Mostramos que las retracciones, al iluminar los casos individuales de mala conducta y las políticas generales para evitarla, oscurecen los actores y los procesos mediante los cuales se efectúan las retracciones, produciendo patrones altamente fragmentadas de visibilidad. Estos patrones se asemejan a la bifurcación en los sistemas de justicia actuales.
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The design of Si-(B)-C materials is investigated, with detailed insight into the precursor chemistry and processing, the precursor-to-ceramic transformation, and the ceramic microstructural evolution at high temperatures. In the early stage of the process, the reaction between allylhydridopolycarbosilane (AHPCS) and borane dimethyl sulfide is achieved. This is investigated in detail through solid-state NMR and FTIR spectroscopy and elemental analyses for Si/B ratios ranging from 200 to 30. Boron-based bridges linking AHPCS monomeric fragments act as crosslinking units, extending the processability range of AHPCS and suppressing the distillation of oligomeric fragments during the low-temperature pyrolysis regime. Polymers with low boron contents display appropriate requirements for facile processing in solution, leading to the design of monoliths with hierarchical porosity, significant pore volume, and high specific surface area after pyrolysis. Polymers with high boron contents are more appropriate for the preparation of dense ceramics through direct solid shaping and pyrolysis. We provide a comprehensive study of the thermal decomposition mechanisms, and a subsequent detailed study of the high-temperature behavior of the ceramics produced at 1000 °C. The nanostructure and microstructure of the final SiC-based ceramics are intimately linked to the boron content of the polymers. B4 C/C/SiC nanocomposites can be obtained from the polymer with the highest boron content.
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When, in 1928, the Clarke School for the Deaf in Northampton, Massachusetts, opened a psychological research division, it was nothing unusual in a time fascinated with the sciences of education. Yet with its longstanding ties to Northampton's Smith College, the school was able to secure the collaboration of eminent Gestalt psychologist Kurt Koffka, who, in turn, engaged 2 more German-speaking emigrants, Margarete Eberhardt and social psychologist Fritz Heider, and Heider's American wife Grace Moore Heider. This collaboration has seen little attention from historians, who have treated Koffka's and Heider's time in Northampton as a transitory phase. I argue, however, that their research on deafness adds to the history of emigration and knowledge transfer between European and American Schools of psychology, and to historical understanding of the interrelation of Gestalt, child, and social psychology. Professionals in child studies and developmental psychology were keenly interested in the holistic and introspective approach Gestalt psychology offered. Deaf children were considered a particularly fascinating research population for exploring the relationship between thought and language, perception and development, Gestalt, and reality. At the Clarke School, Grace Moore Heider was among the first Americans to apply Gestalt principles to child psychology. In a time in which pejorative eugenic beliefs dominated professional perceptions of disability, the Heiders' groundbreaking work defined the deaf as a social and phenomenological minority. This was in opposition to dominant beliefs in deaf education, yet it points to early roots of a social model of deafness and disability, which historians usually locate in 1960s and '70s activism. (PsycINFO Database Record
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A series of boron-modified polyorganosilazanes was synthesized from a poly(vinylmethyl-co-methyl)silazane and controlled amounts of borane dimethyl sulfide. The role of the chemistry behind their synthesis has been studied in detail by using solid-state NMR spectroscopy, FTIR spectroscopy, and elemental analysis. The intimate relationship between the chemistry and the processability of these polymers is discussed. Polymers with low boron contents displayed appropriate requirements for facile processing in solution, such as impregnation of host carbon materials, which resulted in the design of mesoporous monoliths with a high specific surface area after pyrolysis. Polymers with high boron content are more appropriate for solid-state processing to design mechanically robust monolith-type macroporous and dense structures after pyrolysis. Boron acts as a crosslinking element, which offers the possibility to extend the processability of polyorganosilazanes and suppress the distillation of oligomeric fragments in the low-temperature region of their thermal decomposition (i.e., pyrolysis) at 1000 °C under nitrogen. Polymers with controlled and high ceramic yields were generated. We provide a comprehensive mechanistic study of the two-step thermal decomposition based on a combination of thermogravimetric experiments coupled with elemental analysis, solid-state NMR spectroscopy, and FTIR spectroscopy. Selected characterization tools allowed the investigation of specific properties of the monolith-type SiBCN materials.
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Elevated proteasome activity extends lifespan in model organisms such as yeast, worms and flies. This pro-longevity effect might be mediated by improved protein homeostasis, as this protease is an integral module of the protein homeostasis network. Proteasomes also regulate cellular processes through temporal and spatial degradation of signaling pathway components. Here we demonstrate that the regulatory function of the proteasome plays an essential role in aging cells and that the beneficial impact of elevated proteasome capacity on lifespan partially originates from deregulation of the AMPK signaling pathway. Proteasome-mediated lifespan extension activity was carbon-source dependent and cells with enhancement proteasome function exhibited increased respiratory activity and oxidative stress response. These findings suggested that the pro-aging impact of proteasome upregulation might be related to changes in the metabolic state through a premature induction of respiration. Deletion of yeast AMPK, SNF1, or its activator SNF4 abrogated proteasome-mediated lifespan extension, supporting this hypothesis as the AMPK pathway regulates metabolism. We found that the premature induction of respiration in cells with increased proteasome activity originates from enhanced turnover of Mig1, an AMPK/Snf1 regulated transcriptional repressor that prevents the induction of genes required for respiration. Increasing proteasome activity also resulted in partial relocation of Mig1 from the nucleus to the mitochondria. Collectively, the results argue for a model in which elevated proteasome activity leads to the uncoupling of Snf1-mediated Mig1 regulation, resulting in a premature activation of respiration and thus the induction of a mitohormetic response, beneficial to lifespan. In addition, we observed incorrect Mig1 localization in two other long-lived yeast aging models: cells that overexpress SIR2 or deleted for the Mig1-regulator HXK2. Finally, compromised proteasome function blocks lifespan extension in both strains. Thus, our findings suggest that proteasomes, Sir2, Snf1 and Hxk2 form an interconnected aging network that controls metabolism through coordinated regulation of Mig1.
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Envejecimiento/genética , Hexoquinasa/genética , Proteínas Serina-Treonina Quinasas/genética , Proteínas Represoras/genética , Proteínas de Saccharomyces cerevisiae/genética , Proteínas Reguladoras de Información Silente de Saccharomyces cerevisiae/genética , Sirtuina 2/genética , Proteínas Quinasas Activadas por AMP/genética , Envejecimiento/metabolismo , Regulación Fúngica de la Expresión Génica , Hexoquinasa/metabolismo , Humanos , Mitocondrias/genética , Mitocondrias/metabolismo , Estrés Oxidativo , Fosforilación , Complejo de la Endopetidasa Proteasomal/genética , Proteínas Serina-Treonina Quinasas/metabolismo , Proteínas Represoras/metabolismo , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/crecimiento & desarrollo , Proteínas de Saccharomyces cerevisiae/metabolismo , Proteínas Reguladoras de Información Silente de Saccharomyces cerevisiae/metabolismo , Sirtuina 2/metabolismoRESUMEN
The proteasome, a validated anticancer target, participates in an array of biochemical activities, which range from the proteolysis of defective proteins to antigen presentation. We report the preparation of biochemically and photophysically distinct green, red, and far-red real-time sensors designed to simultaneously monitor the proteasome's chymotrypsin-, trypsin-, and caspase-like activities, respectively. These sensors were employed to assess the effect of simultaneous multiple active site catalysis on the kinetic properties of the individual subunits. Furthermore, we have found that the catalytic signature of the proteasome varies depending on the source, cell type, and disease state. Trypsin-like activity is more pronounced in yeast than in mammals, whereas chymotrypsin-like activity is the only activity detectable in B-cells (unlike other mammalian cells). Furthermore, chymotrypsin-like activity is more prominent in transformed B cells relative to their counterparts from healthy donors.
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Complejo de la Endopetidasa Proteasomal/metabolismo , Animales , Línea Celular , Colorantes Fluorescentes , Humanos , Estructura Molecular , Pliegue de Proteína , Conejos , Saccharomyces cerevisiae , Bibliotecas de Moléculas PequeñasRESUMEN
The proteasome cleaves intracellular proteins into peptides. Earlier studies found that treatment of human embryonic kidney 293T (HEK293T) cells with epoxomicin (an irreversible proteasome inhibitor) generally caused a decrease in levels of intracellular peptides. However, bortezomib (an antitumor drug and proteasome inhibitor) caused an unexpected increase in the levels of most intracellular peptides in HEK293T and SH-SY5Y cells. To address this apparent paradox, quantitative peptidomics was used to study the effect of a variety of other proteasome inhibitors on peptide levels in HEK293T and SH-SY5Y cells. Inhibitors tested included carfilzomib, MG132, MG262, MLN2238, AM114, and clasto-Lactacystin ß-lactone. Only MG262 caused a substantial elevation in peptide levels that was comparable to the effect of bortezomib, although carfilzomib and MLN2238 elevated the levels of some peptides. To explore off-target effects, the proteosome inhibitors were tested with various cellular peptidases. Bortezomib did not inhibit tripeptidyl peptidase 2 and only weakly inhibited cellular aminopeptidase activity, as did some of the other proteasome inhibitors. However, potent inhibitors of tripeptidyl peptidase 2 (butabindide) and cellular aminopeptidases (bestatin) did not substantially alter the peptidome, indicating that the increase in peptide levels due to proteasome inhibitors is not a result of peptidase inhibition. Although we cannot exclude other possibilities, we presume that the paradoxical increase in peptide levels upon treatment with bortezomib and other inhibitors is the result of allosteric effects of these compounds on the proteasome. Because intracellular peptides are likely to be functional, it is possible that some of the physiologic effects of bortezomib and carfilzomib arise from the perturbation of peptide levels inside the cell.
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Antineoplásicos/farmacología , Ácidos Borónicos/farmacología , Oligopéptidos/farmacología , Péptidos/metabolismo , Inhibidores de Proteasoma/farmacología , Pirazinas/farmacología , Bortezomib , Línea Celular Tumoral , Células HEK293 , Humanos , Concentración 50 Inhibidora , Espacio Intracelular/efectos de los fármacos , Espacio Intracelular/metabolismo , Leupeptinas/farmacologíaRESUMEN
The proteasome core interacts with different activators and incorporates alternate active subunits, thereby generating a diverse pool of subspecies. The enzymatic properties of these different species are not well understood. In this issue of Chemistry & Biology, Raule and colleagues present a comprehensive enzymatic characterization of immunoproteasome complexes associated with the proteasome activator PA28.
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Complejo de la Endopetidasa Proteasomal/química , Complejo de la Endopetidasa Proteasomal/metabolismoRESUMEN
The conserved Blm10/PA200 activators bind to the proteasome core particle gate and facilitate turnover of peptides and unfolded proteins in vitro. We report here that Blm10 is required for the maintenance of functional mitochondria. BLM10 expression is induced 25-fold upon a switch from fermentation to oxidative metabolism. In the absence of BLM10, Saccharomyces cerevisiae cells exhibit a temperature-sensitive growth defect under oxidative growth conditions and produce colonies with dysfunctional mitochondria at high frequency. Loss of BLM10 leads to reduced respiratory capacity, increased mitochondrial oxidative damage, and reduced viability in the presence of oxidative stress or death stimuli. In the absence of BLM10, increased fragmentation of the mitochondrial network under oxidative stress is observed indicative of elevated activity of the mitochondrial fission machinery. The degradation of Dnm1, the main factor mediating mitochondrial fission, is impaired in the absence of BLM10 in vitro and in vivo. These data suggest that the mitochondrial functional and morphological changes observed are related to elevated Dnm1 levels. This hypothesis is supported by the finding that cells that constitutively overexpress DNM1 display the same mitochondrial defects as blm10Δ cells. The data are consistent with a model in which Blm10 proteasome-mediated turnover of Dnm1 is required for the maintenance of mitochondrial function and provides cytoprotection under conditions that induce increased mitochondrial damage and programmed cell death.
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GTP Fosfohidrolasas/metabolismo , Mitocondrias/metabolismo , Proteínas Mitocondriales/metabolismo , Complejo de la Endopetidasa Proteasomal/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/metabolismo , Apoptosis , Secuencia de Bases , Cartilla de ADN , GTP Fosfohidrolasas/genética , Proteínas Mitocondriales/genética , Estrés Oxidativo , Proteínas de Saccharomyces cerevisiae/genéticaRESUMEN
The ubiquitin-proteasome system (UPS) is the primary selective degradation system in the nuclei and cytoplasm of eukaryotic cells, required for the turnover of myriad soluble proteins. The hundreds of factors that comprise the UPS include an enzymatic cascade that tags proteins for degradation via the covalent attachment of a poly-ubiquitin chain, and a large multimeric enzyme that degrades ubiquitinated proteins, the proteasome. Protein degradation by the UPS regulates many pathways and is a crucial component of the cellular proteostasis network. Dysfunction of the ubiquitination machinery or the proteolytic activity of the proteasome is associated with numerous human diseases. In this review we discuss the contributions of the proteasome to human pathology, describe mechanisms that regulate the proteolytic capacity of the proteasome, and discuss strategies to modulate proteasome function as a therapeutic approach to ameliorate diseases associated with altered UPS function. This article is part of a Special Issue entitled: Ubiquitin-Proteasome System. Guest Editors: Thomas Sommer and Dieter H. Wolf.
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Enfermedad/etiología , Salud , Complejo de la Endopetidasa Proteasomal/fisiología , Proteolisis , Animales , Activación Enzimática/fisiología , Regulación de la Expresión Génica/fisiología , Humanos , Modelos Moleculares , Procesamiento Proteico-Postraduccional/fisiologíaRESUMEN
Host defense mechanisms are multi-layered and involve constitutive as well as inducible components. The dissection of these complex processes can be greatly facilitated using a reporter gene strategy with a transparent animal. In this study, we use Caenorhabditis elegans as a model host and introduce a new pathogen-inducible fluorescent reporter involving the promoter of f57f4.4, a gene encoding a putative component of the glycocalyx. We show that this reporter construct does not respond to heavy metal or hypertonic environments, but is specifically and locally induced in the intestine upon Photorhabus luminescens and Pseudomonas aeruginosa infections. We further demonstrate that its upregulation requires live pathogens as well as elements of the nematode p38 MAP kinase and TGF-beta pathways. In addition to introducing a new tool for the study of the interactions between C. elegans and a pathogen, our results suggest a role for the glycocalyx in gut immunity.
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Proteínas de Caenorhabditis elegans/inmunología , Caenorhabditis elegans/inmunología , Infecciones por Enterobacteriaceae/inmunología , Glicocálix/inmunología , Glicoproteínas/inmunología , Infecciones por Pseudomonas/inmunología , Animales , Animales Modificados Genéticamente , Caenorhabditis elegans/genética , Caenorhabditis elegans/microbiología , Proteínas de Caenorhabditis elegans/genética , Genes Reporteros , Glicocálix/genética , Glicoproteínas/genética , Proteínas Fluorescentes Verdes/genética , Intestinos/inmunología , Sistema de Señalización de MAP Quinasas/genética , Sistema de Señalización de MAP Quinasas/inmunología , Proteínas de la Membrana , Photorhabdus/inmunología , Regiones Promotoras Genéticas , Pseudomonas aeruginosa/inmunología , Interferencia de ARN , ARN Interferente Pequeño , Factor de Crecimiento Transformador beta/inmunología , Proteínas Quinasas p38 Activadas por Mitógenos/inmunologíaRESUMEN
This manuscript was withdrawn by the author.
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Mitochondria deteriorate during the aging process, but the underlying mechanisms for the decline of this critical organelle are unknown. A new study indicates that in yeast an age-dependent reduction in vacuolar acidification leads to mitochondrial dysfunction through a surprising mechanism: loss of vacuolar neutral amino acid transport.
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Envejecimiento/fisiología , Mitocondrias/fisiologíaRESUMEN
Magmas is a growth factor responsive gene encoding an essential mitochondrial protein in mammalian cells. Pam16, the Magmas ortholog in Saccharomyces cerevisiae, is a component of the presequence translocase-associated motor. A temperature-sensitive allele (pam16-I61N) was used to query an array of non-essential gene-deletion strains for synthetic genetic interactions. The pam16-I61N mutation at ambient temperature caused synthetic lethal or sick phenotypes with genes involved in lipid metabolism, perixosome synthesis, histone deacetylation and mitochondrial protein import. The gene deletion array was also screened for suppressors of the pam16-I61N growth defect to identify compensatory pathways. Five suppressor genes were identified (SUR4, ISC1, IPT1, SKN1, and FEN1) and all are involved in sphingolipid metabolism. pam16-I61N cells cultured in glucose at non-permissive temperatures resulted in rapid growth inhibition and G1 cell cycle arrest, but cell viability was maintained. Altered mitochondria morphology, reduced peroxisome induction in glycerol/ethanol and oleate, and changes in the levels of several sphingolipids including C18 alpha-hydroxy-phytoceramide, were also observed in the temperature sensitive strain. Deletion of SUR4, the strongest suppressor, reversed the temperature sensitive fermentative growth defect, the morphological changes and the elevated levels of C18 alpha-hydroxy phytoceramide in pam16-I61N. Deletion of the other four suppressor genes had similar effects on C18 alpha-hydroxy-phytoceramide levels and restored proliferation to the pam16-I61N strain. In addition, pam16-I61N inhibited respiratory growth, likely by reducing cardiolipin, which is essential for mitochondrial function. Our results suggest that the pleiotropic effects caused by impaired Pam16/Magmas function are mediated in part by changes in lipid metabolism.
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Acetiltransferasas/genética , Regulación Fúngica de la Expresión Génica , Proteínas de Transporte de Membrana Mitocondrial/genética , Proteínas de Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/genética , Esfingolípidos/metabolismo , Acetiltransferasas/deficiencia , Puntos de Control del Ciclo Celular , Proliferación Celular , Fermentación , Eliminación de Gen , Perfilación de la Expresión Génica , Glucosa/metabolismo , Humanos , Mitocondrias/metabolismo , Proteínas de Transporte de Membrana Mitocondrial/deficiencia , Proteínas del Complejo de Importación de Proteínas Precursoras Mitocondriales , Proteínas Mitocondriales/genética , Proteínas Mitocondriales/metabolismo , Análisis de Secuencia por Matrices de Oligonucleótidos , Fenotipo , Saccharomyces cerevisiae/enzimología , Homología de Secuencia de Aminoácido , TemperaturaRESUMEN
The cellular response to ionizing radiation (IR) involves a variety of mechanisms to repair damage and maintain cell survival. We previously reported that the proteasome activator PA200 promotes long-term cell survival after IR exposure. The molecular function of PA200 is to enhance proteasome-mediated cleavage after glutamate; however, it is not known how this molecular function promotes survival after IR exposure. Here, we report that upon IR exposure, cellular demand for exogenous glutamine is increased. Cells containing PA200 are capable of surviving this IR-induced glutamine demand, whereas PA200-deficient cells show impaired long-term survival. Additional glutamine supplementation reverses the radiosensitivity of PA200-knockdown cells suggesting impaired glutamine homeostasis in these cells. Indeed, PA200-knockdown cells are unable to maintain intracellular glutamine levels. Furthermore, when extracellular glutamine is limiting, cells that contain PA200 respond by slowing growth, but PA200-knockdown cells and cells in which post-glutamyl proteasome activity is inhibited are nonresponsive and continue rapid growth. This cellular unresponsiveness to nutrient depletion is also reflected at the level of the mTOR substrate ribosomal S6 kinase (S6K). Thus, inability to restrict growth causes PA200-deficient cells to continue growing and eventually die due to lack of available glutamine. Together, these data indicate an important role for PA200 and post-glutamyl proteasome activity in maintaining glutamine homeostasis, which appears to be especially important for long-term survival of tumor cells after radiation exposure.
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Supervivencia Celular , Glutamina/metabolismo , Proteínas Nucleares/metabolismo , Radiación Ionizante , Animales , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/efectos de la radiación , Técnicas de Silenciamiento del Gen , Glutamina/farmacología , Células HeLa , Humanos , Melanoma Experimental , Ratones , Proteínas Nucleares/genética , Tolerancia a Radiación/efectos de los fármacos , Tolerancia a Radiación/genética , Transducción de SeñalRESUMEN
For optimal proteolytic function, the central core of the proteasome (core particle (CP) or 20S) has to associate with activators. We investigated the impact of the yeast activator Blm10 on proteasomal peptide and protein degradation. We found enhanced degradation of peptide substrates in the presence of Blm10 and demonstrated that Blm10 has the capacity to accelerate proteasomal turnover of the unstructured protein tau-441 in vitro. Mechanistically, proteasome activation requires the opening of a closed gate, which allows passage of unfolded proteins into the catalytic chamber. Our data indicate that gate opening by Blm10 is achieved via engagement of its C-terminal segment with the CP. Crucial for this activity is a conserved C-terminal YYX motif, with the penultimate tyrosine playing a preeminent role. Thus, Blm10 utilizes a gate opening strategy analogous to the proteasomal ATPases HbYX-dependent mechanism. Because gating incompetent Blm10 C-terminal point mutants confers a loss of function phenotype, we propose that the cellular function of Blm10 is based on CP association and activation to promote the degradation of proteasome substrates.