RESUMEN
The nucleolus is the site of ribosome biogenesis, one of the most resource-intensive processes in eukaryotic cells. Accordingly, nucleolar morphology and activity are highly responsive to growth signaling and nucleolar insults which are collectively included in the actively evolving concept of nucleolar stress. Importantly, nucleolar alterations are a prominent feature of multiple human pathologies, including cancer and neurodegeneration, as well as being associated with aging. The past decades have seen numerous attempts to isolate compounds targeting different facets of nucleolar activity. We provide an overview of therapeutic opportunities for targeting nucleoli in different pathologies and currently available therapies.
Asunto(s)
Neoplasias , Ribosomas , Humanos , Nucléolo Celular/patología , Neoplasias/tratamiento farmacológico , Neoplasias/patología , EnvejecimientoRESUMEN
Whole-exome sequencing of autism spectrum disorder (ASD) probands and unaffected family members has identified many genes harboring de novo variants suspected to play a causal role in the disorder. Of these, chromodomain helicase DNA-binding protein 8 (CHD8) is the most recurrently mutated. Despite the prevalence of CHD8 mutations, we have little insight into how CHD8 loss affects genome organization or the functional consequences of these molecular alterations in neurons. Here, we engineered two isogenic human embryonic stem cell lines with CHD8 loss-of-function mutations and characterized differences in differentiated human cortical neurons. We identified hundreds of genes with altered expression, including many involved in neural development and excitatory synaptic transmission. Field recordings and single-cell electrophysiology revealed a 3-fold decrease in firing rates and synaptic activity in CHD8+/- neurons, as well as a similar firing-rate deficit in primary cortical neurons from Chd8+/- mice. These alterations in neuron and synapse function can be reversed by CHD8 overexpression. Moreover, CHD8+/- neurons displayed a large increase in open chromatin across the genome, where the greatest change in compaction was near autism susceptibility candidate 2 (AUTS2), which encodes a transcriptional regulator implicated in ASD. Genes with changes in chromatin accessibility and expression in CHD8+/- neurons have significant overlap with genes mutated in probands for ASD, intellectual disability, and schizophrenia but not with genes mutated in healthy controls or other disease cohorts. Overall, this study characterizes key molecular alterations in genome structure and expression in CHD8+/- neurons and links these changes to impaired neuronal and synaptic function.
Asunto(s)
Trastorno del Espectro Autista , Trastorno Autístico , Humanos , Animales , Ratones , Trastorno Autístico/genética , Trastorno del Espectro Autista/genética , Cromatina/genética , Proteínas de Unión al ADN/genética , Proteínas de Unión al ADN/metabolismo , Expresión Génica , Factores de Transcripción/genéticaRESUMEN
Several viruses hijack various forms of endocytosis in order to infect host cells. Here, we report the discovery of a molecule with antiviral properties that we named virapinib, which limits viral entry by macropinocytosis. The identification of virapinib derives from a chemical screen using high-throughput microscopy, where we identified chemical entities capable of preventing infection with a pseudotype virus expressing the spike (S) protein from SARS-CoV-2. Subsequent experiments confirmed the capacity of virapinib to inhibit infection by SARS-CoV-2, as well as by additional viruses, such as mpox virus and TBEV. Mechanistic analyses revealed that the compound inhibited macropinocytosis, limiting this entry route for the viruses. Importantly, virapinib has no significant toxicity to host cells. In summary, we present the discovery of a molecule that inhibits macropinocytosis, thereby limiting the infectivity of viruses that use this entry route such as SARS-CoV2.
RESUMEN
We here conducted an image-based chemical screen to evaluate how medically approved drugs, as well as drugs that are currently under development, influence overall translation levels. None of the compounds up-regulated translation, which could be due to the screen being performed in cancer cells grown in full media where translation is already present at very high levels. Regarding translation down-regulators, and consistent with current knowledge, inhibitors of the mechanistic target of rapamycin (mTOR) signaling pathway were the most represented class. In addition, we identified that inhibitors of sphingosine kinases (SPHKs) also reduce mRNA translation levels independently of mTOR. Mechanistically, this is explained by an effect of the compounds on the membranes of the endoplasmic reticulum (ER), which activates the integrated stress response (ISR) and contributes to the toxicity of SPHK inhibitors. Surprisingly, the toxicity and activation of the ISR triggered by 2 independent SPHK inhibitors, SKI-II and ABC294640, the latter in clinical trials, are also observed in cells lacking SPHK1 and SPHK2. In summary, our study provides a useful resource on the effects of medically used drugs on translation, identified compounds capable of reducing translation independently of mTOR and has revealed that the cytotoxic properties of SPHK inhibitors being developed as anticancer agents are independent of SPHKs.
Asunto(s)
Fosfotransferasas (Aceptor de Grupo Alcohol)/antagonistas & inhibidores , Biosíntesis de Proteínas/fisiología , Animales , Línea Celular , Diseño de Fármacos , Inhibidores Enzimáticos/farmacología , Ensayos Analíticos de Alto Rendimiento/métodos , Humanos , Procesamiento de Imagen Asistido por Computador/métodos , Lisofosfolípidos/metabolismo , Espectrometría de Masas/métodos , Estructura Molecular , Fosfotransferasas (Aceptor de Grupo Alcohol)/metabolismo , Biosíntesis de Proteínas/efectos de los fármacos , ARN Mensajero/genética , ARN Mensajero/metabolismo , Transducción de Señal/efectos de los fármacos , Bibliotecas de Moléculas Pequeñas , Esfingosina/metabolismoRESUMEN
Translation elongation factor eIF5A binds to ribosomes to promote peptide bonds between problematic amino acids for the reaction like prolines. eIF5A is highly conserved and essential in eukaryotes, which usually contain two similar but differentially expressed paralogue genes. The human eIF5A-1 isoform is abundant and implicated in some cancer types; the eIF5A-2 isoform is absent in most cells but becomes overexpressed in many metastatic cancers. Several reports have connected eIF5A and mitochondria because it co-purifies with the organelle or its inhibition reduces respiration and mitochondrial enzyme levels. However, the mechanisms of eIF5A mitochondrial function, and whether eIF5A expression is regulated by the mitochondrial metabolism, are unknown. We analysed the expression of yeast eIF5A isoforms Tif51A and Tif51B under several metabolic conditions and in mutants. The depletion of Tif51A, but not Tif51B, compromised yeast growth under respiration and reduced oxygen consumption. Tif51A expression followed dual positive regulation: by high glucose through TORC1 signalling, like other translation factors, to promote growth and by low glucose or non-fermentative carbon sources through Snf1 and heme-dependent transcription factor Hap1 to promote respiration. Upon iron depletion, Tif51A was down-regulated and Tif51B up-regulated. Both were Hap1-dependent. Our results demonstrate eIF5A expression regulation by cellular metabolic status.
Asunto(s)
Nutrientes , Factores de Iniciación de Péptidos/metabolismo , Proteínas de Unión al ARN/metabolismo , Proteínas de Saccharomyces cerevisiae/metabolismo , Saccharomyces cerevisiae/metabolismo , Transducción de Señal , Aerobiosis/efectos de los fármacos , Carbono/farmacología , Ciclo del Ácido Cítrico/efectos de los fármacos , Regulación hacia Abajo/efectos de los fármacos , Regulación hacia Abajo/genética , Fermentación/efectos de los fármacos , Regulación Fúngica de la Expresión Génica/efectos de los fármacos , Glucosa/metabolismo , Hemo/metabolismo , Hierro/metabolismo , Deficiencias de Hierro , Lisina/análogos & derivados , Lisina/metabolismo , Diana Mecanicista del Complejo 1 de la Rapamicina/metabolismo , Análisis de Flujos Metabólicos , Modelos Biológicos , Isoformas de Proteínas/metabolismo , Saccharomyces cerevisiae/efectos de los fármacos , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/crecimiento & desarrollo , Transducción de Señal/efectos de los fármacos , Regulación hacia Arriba/efectos de los fármacos , Regulación hacia Arriba/genética , Factor 5A Eucariótico de Iniciación de TraducciónRESUMEN
The expansion of GGGGCC repeats within the first intron of C9ORF72 constitutes the most common cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Through repeat-associated non-ATG translation, these expansions are translated into dipeptide repeats (DPRs), some of which accumulate at nucleoli and lead to cell death. We here performed a chemical screen to identify compounds reducing the toxicity of ALS-related poly(PR) peptides. Our screening identified sodium phenylbutyrate, currently in clinical trials, and BET Bromodomain inhibitors as modifiers of poly(PR) toxicity in cell lines and developing zebrafish embryos. Mechanistically, we show that BET Bromodomain inhibitors rescue the nucleolar stress induced by poly(PR) or actinomycin D, alleviating the effects of the DPR in nucleolus-related functions such as mRNA splicing or translation. Our work suggests that BET Bromodomain inhibitors might have beneficial effects in diseases linked to nucleolar stress such as ALS/FTD.
Asunto(s)
Apoptosis/efectos de los fármacos , Proteína C9orf72/química , Péptidos/toxicidad , Esclerosis Amiotrófica Lateral/metabolismo , Esclerosis Amiotrófica Lateral/patología , Animales , Proteína C9orf72/genética , Proteína C9orf72/metabolismo , Línea Celular Tumoral , Nucléolo Celular/efectos de los fármacos , Nucléolo Celular/metabolismo , Proteínas Cromosómicas no Histona/metabolismo , Expansión de las Repeticiones de ADN , Dactinomicina/toxicidad , Embrión no Mamífero/efectos de los fármacos , Embrión no Mamífero/fisiología , Demencia Frontotemporal/metabolismo , Demencia Frontotemporal/patología , Inhibidores de Histona Desacetilasas/farmacología , Humanos , Péptidos/síntesis química , Proteínas/antagonistas & inhibidores , Proteínas/metabolismo , Pez Cebra/crecimiento & desarrolloRESUMEN
Bacteria are needed for a vast range of biotechnological processes, which they carry out either as pure cultures or in association with other bacteria and/or fungi. The potential of bacteria as biofactories is hampered, though, by their limited mobility in solid or semisolid media such as agricultural or domestic waste. This work represents an attempt toward overcoming this limitation by associating bacterial biotechnological properties with the transport ability of the nematode Caenorhabditis elegans. We report here biofilm formation on C. elegans by engineered Escherichia coli expressing a Xhenorhabdus nematophila adhesion operon and induction of nematode social feeding behavior (clumping) through an E. coli-mediated iRNA blocking on the expression of FLP-21, a neuropeptide involved in worm solitary behavior.