RESUMEN
The MHC class I antigen presentation system enables T cell immunosurveillance of cancers and viruses. A substantial fraction of the immunopeptidome derives from rapidly degraded nascent polypeptides (DRiPs). By knocking down each of the 80 ribosomal proteins, we identified proteins that modulate peptide generation without altering source protein expression. We show that 60S ribosomal proteins L6 (RPL6) and RPL28, which are adjacent on the ribosome, play opposite roles in generating an influenza A virus-encoded peptide. Depleting RPL6 decreases ubiquitin-dependent peptide presentation, whereas depleting RPL28 increases ubiquitin-dependent and -independent peptide presentation. 40S ribosomal protein S28 (RPS28) knockdown increases total peptide supply in uninfected cells by increasing DRiP synthesis from non-canonical translation of "untranslated" regions and non-AUG start codons and sensitizes tumor cells for T cell targeting. Our findings raise the possibility of modulating immunosurveillance by pharmaceutical targeting ribosomes.
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Presentación de Antígeno , Antígenos de Histocompatibilidad Clase I/biosíntesis , Proteínas Ribosómicas/metabolismo , Subunidades Ribosómicas Grandes de Eucariotas/metabolismo , Subunidades Ribosómicas Pequeñas de Eucariotas/metabolismo , Linfocitos T/metabolismo , Animales , Línea Celular Tumoral , Técnicas de Cocultivo , Células HEK293 , Antígenos de Histocompatibilidad Clase I/inmunología , Interacciones Huésped-Patógeno , Humanos , Vigilancia Inmunológica , Virus de la Influenza A/inmunología , Virus de la Influenza A/patogenicidad , Melanoma/inmunología , Melanoma/metabolismo , Ratones Endogámicos C57BL , Ratones Transgénicos , Proteínas Ribosómicas/genética , Subunidades Ribosómicas Grandes de Eucariotas/genética , Subunidades Ribosómicas Pequeñas de Eucariotas/genética , Neoplasias Cutáneas/inmunología , Neoplasias Cutáneas/metabolismo , Linfocitos T/inmunología , Linfocitos T/virologíaRESUMEN
Regulation of mRNA translation is a crucial step in controlling gene expression in stressed cells, impacting many pathologies, including heart ischemia. In recent years, ribosome heterogeneity has emerged as a key control mechanism driving the translation of subsets of mRNAs. In this study, we investigated variations in ribosome composition in human cardiomyocytes subjected to endoplasmic reticulum stress induced by tunicamycin treatment. Our findings demonstrate that this stress inhibits global translation in cardiomyocytes while activating internal ribosome entry site (IRES)-dependent translation. Analysis of translating ribosome composition in stressed and unstressed cardiomyocytes was conducted using mass spectrometry. We observed no significant changes in ribosomal protein composition, but several mitochondrial ribosomal proteins (MRPs) were identified in cytosolic polysomes, showing drastic variations between stressed and unstressed cells. The most notable increase in polysomes of stressed cells was observed in MRPS15. Its interaction with ribosomal proteins was confirmed by proximity ligation assay (PLA) and immunoprecipitation, suggesting its intrinsic role as a ribosomal component during stress. Knock-down or overexpression experiments of MRPS15 revealed its role as an activator of IRES-dependent translation. Furthermore, polysome profiling after immunoprecipitation with anti-MRPS15 antibody revealed that the "MRPS15 ribosome" is specialized in translating mRNAs involved in the unfolded protein response.
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Miocitos Cardíacos , Proteínas Ribosómicas , Humanos , Proteínas Ribosómicas/genética , Proteínas Ribosómicas/metabolismo , Miocitos Cardíacos/metabolismo , Ribosomas/metabolismo , Polirribosomas/metabolismo , Citosol/metabolismo , ARN Mensajero/genética , ARN Mensajero/metabolismo , Sitios Internos de Entrada al Ribosoma , Biosíntesis de ProteínasRESUMEN
The role of ribosome biogenesis in erythroid development is supported by the recognition of erythroid defects in ribosomopathies in both Diamond-Blackfan anemia and 5q- syndrome. Whether ribosome biogenesis exerts a regulatory function on normal erythroid development is still unknown. In the present study, a detailed characterization of ribosome biogenesis dynamics during human and murine erythropoiesis showed that ribosome biogenesis is abruptly interrupted by the decline in ribosomal DNA transcription and the collapse of ribosomal protein neosynthesis. Its premature arrest by the RNA Pol I inhibitor CX-5461 targeted the proliferation of immature erythroblasts. p53 was activated spontaneously or in response to CX-5461, concomitant to ribosome biogenesis arrest, and drove a transcriptional program in which genes involved in cell cycle-arrested, negative regulation of apoptosis, and DNA damage response were upregulated. RNA Pol I transcriptional stress resulted in nucleolar disruption and activation of the ATR-CHK1-p53 pathway. Our results imply that the timing of ribosome biogenesis extinction and p53 activation is crucial for erythroid development. In ribosomopathies in which ribosome availability is altered by unbalanced production of ribosomal proteins, the threshold downregulation of ribosome biogenesis could be prematurely reached and, together with pathological p53 activation, prevents a normal expansion of erythroid progenitors.
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Diferenciación Celular/fisiología , Células Eritroides/citología , Eritropoyesis/fisiología , Ribosomas/metabolismo , Proteína p53 Supresora de Tumor/metabolismo , Animales , Células Madre Hematopoyéticas , Humanos , Ratones , Biogénesis de OrganelosRESUMEN
BACKGROUND: A current critical need remains in the identification of prognostic and predictive markers in early breast cancer. It appears that a distinctive trait of cancer cells is their addiction to hyperactivation of ribosome biogenesis. Thus, ribosome biogenesis might be an innovative source of biomarkers that remains to be evaluated. METHODS: Here, fibrillarin (FBL) was used as a surrogate marker of ribosome biogenesis due to its essential role in the early steps of ribosome biogenesis and its association with poor prognosis in breast cancer when overexpressed. Using 3,275 non-metastatic primary breast tumors, we analysed FBL mRNA expression levels and protein nucleolar organisation. Usage of TCGA dataset allowed transcriptomic comparison between the different FBL expression levels-related breast tumours. RESULTS: We unexpectedly discovered that in addition to breast tumours expressing high level of FBL, about 10% of the breast tumors express low level of FBL. A correlation between low FBL mRNA level and lack of FBL detection at protein level using immunohistochemistry was observed. Interestingly, multivariate analyses revealed that these low FBL tumors displayed poor outcome compared to current clinical gold standards. Transcriptomic data revealed that FBL expression is proportionally associated with distinct amount of ribosomes, low FBL level being associated with low amount of ribosomes. Moreover, the molecular programs supported by low and high FBL expressing tumors were distinct. CONCLUSION: Altogether, we identified FBL as a powerful ribosome biogenesis-related independent marker of breast cancer outcome. Surprisingly we unveil a dual association of the ribosome biogenesis FBL factor with prognosis. These data suggest that hyper- but also hypo-activation of ribosome biogenesis are molecular traits of distinct tumors.
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Neoplasias de la Mama , Biomarcadores/metabolismo , Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Proteínas Cromosómicas no Histona , Femenino , Humanos , ARN Mensajero/genética , ARN Mensajero/metabolismo , Ribosomas/genética , Ribosomas/metabolismoRESUMEN
Ribosomal RNAs (rRNAs) are main effectors of messenger RNA (mRNA) decoding, peptide-bond formation, and ribosome dynamics during translation. Ribose 2'-O-methylation (2'-O-Me) is the most abundant rRNA chemical modification, and displays a complex pattern in rRNA. 2'-O-Me was shown to be essential for accurate and efficient protein synthesis in eukaryotic cells. However, whether rRNA 2'-O-Me is an adjustable feature of the human ribosome and a means of regulating ribosome function remains to be determined. Here we challenged rRNA 2'-O-Me globally by inhibiting the rRNA methyl-transferase fibrillarin in human cells. Using RiboMethSeq, a nonbiased quantitative mapping of 2'-O-Me, we identified a repertoire of 2'-O-Me sites subjected to variation and demonstrate that functional domains of ribosomes are targets of 2'-O-Me plasticity. Using the cricket paralysis virus internal ribosome entry site element, coupled to in vitro translation, we show that the intrinsic capability of ribosomes to translate mRNAs is modulated through a 2'-O-Me pattern and not by nonribosomal actors of the translational machinery. Our data establish rRNA 2'-O-Me plasticity as a mechanism providing functional specificity to human ribosomes.
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Biosíntesis de Proteínas , ARN Ribosómico/metabolismo , Proteínas Cromosómicas no Histona/genética , Proteínas Cromosómicas no Histona/metabolismo , Regulación de la Expresión Génica , Técnicas de Silenciamiento del Gen , Células HeLa , Humanos , MetilaciónRESUMEN
We describe here the evaluation of the cytotoxic efficacy of two platinum (II) complexes bearing an N-heterocyclic carbene (NHC) ligand, a pyridine ligand and bromide or iodide ligands on a panel of human metastatic cutaneous melanoma cell lines representing different genetic subsets including BRAF-inhibitor-resistant cell lines, namely A375, SK-MEL-28, MeWo, HMCB, A375-R, SK-MEL-5-R and 501MEL-R. Cisplatin and dacarbazine were also studied for comparison purposes. Remarkably, the iodine-labelled Pt-NHC complex strongly inhibited proliferation of all tested melanoma cells after 1-h exposure, likely due to its rapid uptake by melanoma cells. The mechanism of this inhibitory activity involves the formation of DNA double-strand breaks and apoptosis. Considering the intrinsic chemoresistance of metastatic melanoma cells of current systemic treatments, these findings are promising and could give research opportunities in the future to improve the prognosis of patients suffering from unresectable metastatic melanoma that are not eligible or that do not respond to the most effective drugs available to date, namely BRAF inhibitors and the anti-PD-1 monoclonal antibody (mAb).
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Antineoplásicos/farmacología , Resistencia a Antineoplásicos/efectos de los fármacos , Melanoma/tratamiento farmacológico , Compuestos Organoplatinos/farmacología , Neoplasias Cutáneas/tratamiento farmacológico , Antineoplásicos/química , Antineoplásicos/farmacocinética , Muerte Celular/efectos de los fármacos , Muerte Celular/fisiología , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Roturas del ADN de Doble Cadena/efectos de los fármacos , Daño del ADN , Resistencia a Antineoplásicos/genética , Ensayos de Selección de Medicamentos Antitumorales , Compuestos Heterocíclicos/química , Compuestos Heterocíclicos/farmacología , Humanos , Melanoma/patología , Metano/análogos & derivados , Metano/química , Compuestos Organoplatinos/química , Compuestos Organoplatinos/farmacocinética , Proteínas Proto-Oncogénicas B-raf/antagonistas & inhibidores , Proteínas Proto-Oncogénicas B-raf/genética , Neoplasias Cutáneas/patología , Proteína bcl-X/metabolismo , Melanoma Cutáneo MalignoRESUMEN
Downregulation of CD20, a molecular target for monoclonal antibodies (mAbs), is a clinical problem leading to decreased efficacy of anti-CD20-based therapeutic regimens. The epigenetic modulation of CD20 coding gene (MS4A1) has been proposed as a mechanism for the reduced therapeutic efficacy of anti-CD20 antibodies and confirmed with nonselective histone deacetylase inhibitors (HDACis). Because the use of pan-HDACis is associated with substantial adverse effects, the identification of particular HDAC isoforms involved in CD20 regulation seems to be of paramount importance. In this study, we demonstrate for the first time the role of HDAC6 in the regulation of CD20 levels. We show that inhibition of HDAC6 activity significantly increases CD20 levels in established B-cell tumor cell lines and primary malignant cells. Using pharmacologic and genetic approaches, we confirm that HDAC6 inhibition augments in vitro efficacy of anti-CD20 mAbs and improves survival of mice treated with rituximab. Mechanistically, we demonstrate that HDAC6 influences synthesis of CD20 protein independently of the regulation of MS4A1 transcription. We further demonstrate that translation of CD20 mRNA is significantly enhanced after HDAC6 inhibition, as shown by the increase of CD20 mRNA within the polysomal fraction, indicating a new role of HDAC6 in the posttranscriptional mechanism of CD20 regulation. Collectively, our findings suggest HDAC6 inhibition is a rational therapeutic strategy to be implemented in combination therapies with anti-CD20 monoclonal antibodies and open up novel avenues for the clinical use of HDAC6 inhibitors.
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Antígenos CD20/genética , Antineoplásicos/farmacología , Inhibidores de Histona Desacetilasas/farmacología , Histona Desacetilasas/metabolismo , Leucemia Linfocítica Crónica de Células B/tratamiento farmacológico , Linfoma no Hodgkin/tratamiento farmacológico , Rituximab/farmacología , Animales , Antígenos CD20/inmunología , Linfocitos B/efectos de los fármacos , Linfocitos B/inmunología , Linfocitos B/patología , Línea Celular Tumoral , Femenino , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Histona Desacetilasa 6 , Humanos , Leucemia Linfocítica Crónica de Células B/genética , Leucemia Linfocítica Crónica de Células B/inmunología , Leucemia Linfocítica Crónica de Células B/patología , Linfoma no Hodgkin/genética , Linfoma no Hodgkin/inmunología , Linfoma no Hodgkin/patología , Ratones Endogámicos BALB C , Ratones SCID , ARN Mensajero/genética , Células Tumorales Cultivadas , Regulación hacia Arriba/efectos de los fármacosRESUMEN
G-rich oligonucleotide, AS1411, has been shown to interact with nucleolin and to inhibit cancer cell proliferation and tumor growth. This antiproliferative action is increased when AS1411 is conjugated to different types of nanoparticles. However, the molecular mechanisms are not known. In this work, we show in several cell lines that optimized AS1411-conjugated gold nanoparticles (GNS-AS1411) inhibit nucleolin expression at the RNA and protein levels. We observed an alteration of the nucleolar structure with a decrease of ribosomal RNA accumulation comparable to what is observed upon nucleolin knock down. However, the expression of genes involved in cell cycle and the cell cycle blockage by GNS-AS1411 are not regulated in the same way as that in cells where nucleolin has been knocked down. These data suggest that the anti-proliferative activity of GNS-AS1411 is not the only consequence of nucleolin targeting and down-regulation.
Asunto(s)
Puntos de Control del Ciclo Celular/efectos de los fármacos , Regulación hacia Abajo/efectos de los fármacos , Oro , Nanopartículas del Metal/química , Oligodesoxirribonucleótidos , Fosfoproteínas/biosíntesis , ARN Ribosómico/biosíntesis , Proteínas de Unión al ARN/biosíntesis , Aptámeros de Nucleótidos , Línea Celular Tumoral , Oro/química , Oro/farmacología , Humanos , Oligodesoxirribonucleótidos/química , Oligodesoxirribonucleótidos/farmacología , NucleolinaRESUMEN
Translation is one of the final steps that regulate gene expression. The ribosome is the effector of translation through to its role in mRNA decoding and protein synthesis. Many mechanisms have been extensively described accounting for translational regulation. However it emerged only recently that ribosomes themselves could contribute to this regulation. Indeed, though it is well-known that the translational efficiency of the cell is linked to ribosome abundance, studies recently demonstrated that the composition of the ribosome could alter translation of specific mRNAs. Evidences suggest that according to the status, environment, development, or pathological conditions, cells produce different populations of ribosomes which differ in their ribosomal protein and/or RNA composition. Those observations gave rise to the concept of "specialized ribosomes", which proposes that a unique ribosome composition determines the translational activity of this ribosome. The current review will present how technological advances have participated in the emergence of this concept, and to which extent the literature sustains this concept today.
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Eucariontes/genética , Eucariontes/metabolismo , Regulación de la Expresión Génica , Biosíntesis de Proteínas , Ribosomas/metabolismo , Animales , Células Eucariotas , Humanos , ARN Mensajero/genética , ARN Mensajero/metabolismo , ARN Ribosómico/genética , ARN Ribosómico/metabolismo , Proteínas Ribosómicas/metabolismoRESUMEN
Neuroblastoma (NB) is the most common pediatric tumor and is currently treated by several types of therapies including chemotherapies, such as bortezomib treatment. However, resistance to bortezomib is frequently observed by mechanisms that remain to be deciphered. Bortezomib treatment leads to caspase activation and aggresome formation. Using models of patients-derived NB cell lines with different levels of sensitivity to bortezomib, we show that the activated form of caspase 3 accumulates within aggresomes of NB resistant cells leading to an impairment of bortezomib-induced apoptosis and increased cell survival. Our findings unveil a new mechanism of resistance to chemotherapy based on an altered subcellular distribution of the executioner caspase 3. This mechanism could explain the resistance developed in NB patients treated with bortezomib, emphasizing the potential of drugs targeting aggresomes.
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Antineoplásicos , Neuroblastoma , Niño , Humanos , Bortezomib/farmacología , Bortezomib/uso terapéutico , Caspasa 3/farmacología , Línea Celular Tumoral , Apoptosis , Neuroblastoma/tratamiento farmacológico , Neuroblastoma/patología , Antineoplásicos/farmacología , Antineoplásicos/uso terapéuticoRESUMEN
Herpes simplex virus type 1 (HSV-1) infection induces profound nucleolar modifications at the functional and organizational levels, including nucleolar invasion by several viral proteins. One of these proteins is US11, which exhibits several different functions and displays both cytoplasmic localization and clear nucleolar localization very similar to that of the major multifunctional nucleolar protein nucleolin. To determine whether US11 interacts with nucleolin, we purified US11 protein partners by coimmunoprecipitations using a tagged protein, Flag-US11. From extracts of cells expressing Flag-US11 protein, we copurified a protein of about 100 kDa that was further identified as nucleolin. In vitro studies have demonstrated that nucleolin interacts with US11 and that the C-terminal domain of US11, which is required for US11 nucleolar accumulation, is sufficient for interaction with nucleolin. This association was confirmed in HSV-1-infected cells. We found an increase in the nucleolar accumulation of US11 in nucleolin-depleted cells, thereby revealing that nucleolin could play a role in US11 nucleocytoplasmic trafficking through one-way directional transport out of the nucleolus. Since nucleolin is required for HSV-1 nuclear egress, the interaction of US11 with nucleolin may participate in the outcome of infection.
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Fosfoproteínas/metabolismo , Proteínas de Unión al ARN/metabolismo , Proteínas Virales/metabolismo , Far-Western Blotting , Células HeLa , Humanos , Inmunoprecipitación , Unión Proteica , Transporte de Proteínas , ARN Interferente Pequeño , NucleolinaRESUMEN
Many studies highlight the potential link between the chronic degenerative Alzheimer's disease and the infection by the herpes simplex virus type-1 (HSV-1). However, the molecular mechanisms making possible this HSV-1-dependent process remain to be understood. Using neuronal cells expressing the wild type form of amyloid precursor protein (APP) infected by HSV-1, we characterized a representative cellular model of the early stage of the sporadic form of the disease and unraveled a molecular mechanism sustaining this HSV-1- Alzheimer's disease interplay. Here, we show that HSV-1 induces caspase-dependent production of the 42 amino-acid long amyloid peptide (Aß42) oligomers followed by their accumulation in neuronal cells. Aß42 oligomers and activated caspase 3 (casp3A) concentrate into intracytoplasmic structures observed in Alzheimer's disease neuronal cells called aggresomes. This casp3A accumulation in aggresomes during HSV-1 infection limits the execution of apoptosis until its term, similarly to an abortosis-like event occurring in Alzheimer's disease neuronal cells patients. Indeed, this particular HSV-1 driven cellular context, representative of early stages of the disease, sustains a failed apoptosis mechanism that could explain the chronic amplification of Aß42 production characteristic of Alzheimer's disease patients. Finally, we show that combination of flurbiprofen, a non-steroidal anti-inflammatory drug (NSAID), with caspase inhibitor reduced drastically HSV-1-induced Aß42 oligomers production. This provided mechanistic insights supporting the conclusion of clinical trials showing that NSAIDs reduced Alzheimer's disease incidence in early stage of the disease. Therefore, from our study we propose that caspase-dependent production of Aß42 oligomers together with the abortosis-like event represents a vicious circle in early Alzheimer's disease stages leading to a chronic amplification of Aß42 oligomers that contributes to the establishment of degenerative disorder like Alzheimer's disease in patients infected by HSV-1. Interestingly this process could be targeted by an association of NSAID with caspase inhibitors.
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Enfermedad de Alzheimer , Herpesvirus Humano 1 , Humanos , Enfermedad de Alzheimer/metabolismo , Herpesvirus Humano 1/metabolismo , Neuronas/metabolismo , Antiinflamatorios no Esteroideos , Caspasas/metabolismo , Péptidos beta-Amiloides/metabolismo , Fragmentos de Péptidos/metabolismoRESUMEN
In the adult mammalian central nervous system (CNS), axons fail to regenerate spontaneously after injury because of a combination of extrinsic and intrinsic factors. Despite recent advances targeting the intrinsic regenerative properties of adult neurons, the molecular mechanisms underlying axon regeneration are not fully understood. Here, we uncover a regulatory mechanism that controls the expression of key proteins involved in regeneration at the translational level. Our results show that mRNA-specific translation is critical for promoting axon regeneration. Indeed, we demonstrate that specific ribosome-interacting proteins, such as the protein Huntingtin (HTT), selectively control the translation of a specific subset of mRNAs. Moreover, modulating the expression of these translationally regulated mRNAs is crucial for promoting axon regeneration. Altogether, our findings highlight that selective translation through the customization of the translational complex is a key mechanism of axon regeneration with major implications in the development of therapeutic strategies for CNS repair.
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Axones , Regeneración Nerviosa , Animales , Axones/metabolismo , Regeneración Nerviosa/genética , Sistema Nervioso Central/metabolismo , Neuronas/metabolismo , ARN Mensajero/metabolismo , Mamíferos/metabolismoRESUMEN
Embryonic stem cell (ESC) fate decisions are regulated by a complex circuitry that coordinates gene expression at multiple levels from chromatin to mRNA processing. Recently, ribosome biogenesis and translation have emerged as key pathways that efficiently control stem cell homeostasis, yet the underlying molecular mechanisms remain largely unknown. Here, we identified RSL24D1 as highly expressed in both mouse and human pluripotent stem cells. RSL24D1 is associated with nuclear pre-ribosomes and is required for the biogenesis of 60S subunits in mouse ESCs. Interestingly, RSL24D1 depletion significantly impairs global translation, particularly of key pluripotency factors and of components from the Polycomb Repressive Complex 2 (PRC2). While having a moderate impact on differentiation, RSL24D1 depletion significantly alters ESC self-renewal and lineage commitment choices. Altogether, these results demonstrate that RSL24D1-dependant ribosome biogenesis is both required to sustain the expression of pluripotent transcriptional programs and to silence PRC2-regulated developmental programs, which concertedly dictate ESC homeostasis.
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Células Madre Embrionarias , Células Madre Pluripotentes , Humanos , Animales , Ratones , Células Madre Embrionarias/metabolismo , Diferenciación Celular/genética , Complejo Represivo Polycomb 2/metabolismoRESUMEN
Resistance of advanced hormone-dependent endometrial carcinoma to endocrine therapy remains a worldwide clinical issue. We recently reported that the combination of Vistusertib (V, mTOR inhibitor) and Anastrozole (A, aromatase inhibitor) improves the progression-free rate compared to Anastrozole alone. However, a better patient selection based on biomarkers would improve patient outcome. We evaluate for the first time the usage of ribosome biogenesis (RiBi) factors as a source of innovative markers. Using 47 FFPE tumours (A n = 18; V + A n = 29), 32 blood samples (A n = 13; V + A n = 19) and 30 samples of total RNAs (A n = 12; V + A n = 18) from the VICTORIA clinical trial, we observed an association between RiBi-associated markers and drug activity or prediction of treatment response. NOP10 and NHP2 mRNA levels were significantly higher in non-responders compared to responders in the Vistusertib + Anastrozole arm (P = 0.0194 and P = 0.0002 respectively; i.e. 8 weeks progression-free survival as endpoint). This study provides RiBi-based markers relevant for a better selection of patients with advanced endometrial carcinoma by predicting the response of endocrine therapy combined with mTOR inhibitor.
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Neoplasias de la Mama , Neoplasias Endometriales , Humanos , Femenino , Anastrozol/uso terapéutico , Nitrilos/uso terapéutico , Triazoles/uso terapéutico , Inhibidores de la Aromatasa/uso terapéutico , Biomarcadores , Serina-Treonina Quinasas TOR , Neoplasias Endometriales/tratamiento farmacológico , Neoplasias Endometriales/genética , Ribosomas , Neoplasias de la Mama/tratamiento farmacológico , Antineoplásicos Hormonales/uso terapéuticoRESUMEN
Nutrient availability is a key determinant of tumor cell behavior. While nutrient-rich conditions favor proliferation and tumor growth, scarcity, and particularly glutamine starvation, promotes cell dedifferentiation and chemoresistance. Here, linking ribosome biogenesis plasticity with tumor cell fate, we uncover that the amino acid sensor general control non-derepressible 2 (GCN2; also known as eIF-2-alpha kinase 4) represses the expression of the precursor of ribosomal RNA (rRNA), 47S, under metabolic stress. We show that blockade of GCN2 triggers cell death by an irremediable nucleolar stress and subsequent TP53-mediated apoptosis in patient-derived models of colon adenocarcinoma (COAD). In nutrient-rich conditions, a cell-autonomous GCN2 activity supports cell proliferation by stimulating 47S rRNA transcription, independently of the canonical integrated stress response (ISR) axis. Impairment of GCN2 activity prevents nuclear translocation of methionyl-tRNA synthetase (MetRS), resulting in nucleolar stress, mTORC1 inhibition and, ultimately, autophagy induction. Inhibition of the GCN2-MetRS axis drastically improves the cytotoxicity of RNA polymerase I (RNA pol I) inhibitors, including the first-line chemotherapy oxaliplatin, on patient-derived COAD tumoroids. Our data thus reveal that GCN2 differentially controls ribosome biogenesis according to the nutritional context. Furthermore, pharmacological co-inhibition of the two GCN2 branches and RNA pol I activity may represent a valuable strategy for elimination of proliferative and metabolically stressed COAD cells.
RESUMEN
BACKGROUND: High-grade adult-type diffuse gliomas (HGGs) constitute a heterogeneous group of aggressive tumors that are mostly incurable. Recent advances highlighting the contribution of ribosomes to cancer development have offered new clinical perspectives. Here, we uncovered that isocitrate dehydrogenase (IDH)wt and IDHmut HGGs display distinct alterations of ribosome biology, in terms of rRNA epitranscriptomics and ribosome biogenesis, which could constitute novel hallmarks that can be exploited for the management of these pathologies. METHODS: We analyzed (1) the ribosomal RNA 2'O-ribose methylation (rRNA 2'Ome) using RiboMethSeq and in-house developed bioinformatics tools (https://github.com/RibosomeCRCL/ribomethseq-nfandrRMSAnalyzer) on 3 independent cohorts compiling 71 HGGs (IDHwt n = 30, IDHmut n = 41) and 9 non-neoplastic samples, (2) the expression of ribosome biogenesis factors using medium throughput RT-qPCR as a readout of ribosome biogenesis, and (3) the sensitivity of 5 HGG cell lines to RNA Pol I inhibitors (CX5461, BMH-21). RESULTS: Unsupervised analysis demonstrated that HGGs could be distinguished based on their rRNA 2'Ome epitranscriptomic profile, with IDHwt glioblastomas displaying the most significant alterations of rRNA 2'Ome at specific sites. In contrast, IDHmut HGGs are largely characterized by an overexpression of ribosome biogenesis factors compared to non-neoplastic tissues or IDHwt glioblastomas. Finally, IDHmut HGG-derived spheroids display higher cytotoxicity to CX5461 than IDHwt glioblastoma, while all HGG spheroids display a similar cytotoxicity to BMH-21. CONCLUSIONS: In HGGs, IDH mutational status is associated with specific alterations of the ribosome biology and with distinct sensitivities to RNA Pol I inhibitors.
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Neoplasias Encefálicas , Glioblastoma , Glioma , Adulto , Humanos , Isocitrato Deshidrogenasa/genética , Isocitrato Deshidrogenasa/metabolismo , Neoplasias Encefálicas/patología , Glioblastoma/genética , Glioblastoma/metabolismo , ARN Ribosómico/genética , ARN Ribosómico/metabolismo , Glioma/patología , Metilación , Ribosomas/genética , Ribosomas/metabolismo , Ribosomas/patología , MutaciónRESUMEN
The ribosome is the central effector of protein synthesis, and its synthesis is intimately coordinated with that of proteins. At present, the most documented way to modulate ribosome biogenesis involves control of rDNA transcription by RNA polymerase I (RNA Pol I). Here we show that after infection of human cells with herpes simplex virus type 1 (HSV-1) the rate of ribosome biogenesis is modulated independently of RNA Pol I activity by a dramatic change in the rRNA maturation pathway. This process permits control of the ribosome biogenesis rate, giving the possibility of escaping ribosomal stress and eventually allowing assembly of specialized kinds of ribosomes.
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Herpes Simple/metabolismo , Herpesvirus Humano 1/fisiología , ARN Polimerasa I/metabolismo , Ribosomas/metabolismo , Secuencia de Bases , Activación Enzimática , Células HeLa , Herpes Simple/genética , Humanos , Modelos Biológicos , ARN Polimerasa I/fisiología , Precursores del ARN/metabolismo , Procesamiento Postranscripcional del ARN/fisiología , Transducción de Señal/fisiologíaRESUMEN
Bone is a frequent site of metastasis. Bone metastasis is associated with a short-term prognosis in cancer patients, and current treatments aim to slow its growth, but are rarely curative. Thus, revealing molecular mechanisms that explain why metastatic cells are attracted to the bone micro-environment, and how they successfully settle in the bone marrow-taking advantage over bone resident cells-and grow into macro-metastasis, is essential to propose new therapeutic approaches. MicroRNAs and snoRNAs are two classes of small non-coding RNAs that post-transcriptionally regulate gene expression. Recently, microRNAs and snoRNAs have been pointed out as important players in bone metastasis by (i) preparing the pre-metastatic niche, directly and indirectly affecting the activities of osteoclasts and osteoblasts, (ii) promoting metastatic properties within cancer cells, and (iii) acting as mediators within cells to support cancer cell growth in bone. This review aims to highlight the importance of microRNAs and snoRNAs in metastasis, specifically in bone, and how their roles can be linked together. We then discuss how microRNAs and snoRNAs are secreted by cancer cells and be found as extracellular vesicle cargo. Finally, we provide evidence of how microRNAs and snoRNAs can be potential therapeutic targets, at least in pre-clinical settings, and how their detection in liquid biopsies can be a useful diagnostic and/or prognostic biomarker to predict the risk of relapse in cancer patients.
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Growing evidence exposes translation and its translational machinery as key players in establishing and maintaining physiological and pathological biological processes. Examining translation may not only provide new biological insight but also identify novel innovative therapeutic targets in several fields of biology, including that of epithelial-to-mesenchymal transition (EMT). EMT is currently considered as a dynamic and reversible transdifferentiation process sustaining the transition from an epithelial to mesenchymal phenotype, known to be mainly driven by transcriptional reprogramming. However, it seems that the characterization of EMT plasticity is challenging, relying exclusively on transcriptomic and epigenetic approaches. Indeed, heterogeneity in EMT programs was reported to depend on the biological context. Here, by reviewing the involvement of translational control, translational machinery and ribosome biogenesis characterizing the different types of EMT, from embryonic and adult physiological to pathological contexts, we discuss the added value of integrating translational control and its machinery to depict the heterogeneity and dynamics of EMT programs.