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1.
Elife ; 102021 Nov 17.
Artigo em Inglês | MEDLINE | ID: mdl-34787568

RESUMO

We performed a systematic analysis of the translation rate of tumor-infiltrating lymphocytes (TILs) and the microenvironment inputs affecting it, both in humans and in mice. Measurement of puromycin incorporation, a proxy of protein synthesis, revealed an increase of translating CD4+ and CD8+ cells in tumors, compared to normal tissues. High translation levels are associated with phospho-S6 labeling downstream of mTORC1 activation, whereas low levels correlate with hypoxic areas, in agreement with data showing that T cell receptor stimulation and hypoxia act as translation stimulators and inhibitors, respectively. Additional analyses revealed the specific phenotype of translating TILs. CD8+ translating cells have enriched expression of IFN-γ and CD-39, and reduced SLAMF6, pointing to a cytotoxic phenotype. CD4+ translating cells are mostly regulatory T cells (Tregs) with enriched levels of CTLA-4 and Ki67, suggesting an expanding immunosuppressive phenotype. In conclusion, the majority of translationally active TILs is represented by cytotoxic CD8+ and suppressive CD4+ Tregs, implying that other subsets may be largely composed by inactive bystanders.

2.
Neurology ; 2021 Oct 14.
Artigo em Inglês | MEDLINE | ID: mdl-34649874

RESUMO

BACKGROUND AND OBJECTIVES: Autosomal recessive spastic ataxia of Charlevoix-Saguenay (ARSACS) is caused by mutations in SACS gene encoding sacsin, a huge multimodular protein of unknown function. More than 200 SACS mutations have been described worldwide to date. Since ARSACS presents phenotypic variability, previous empirical studies attempted to correlate the nature and position of SACS mutations with the age of onset or with disease severity, though not considering the effect of the various mutations on protein stability. In this work, we studied genotype-phenotype correlation in ARSACS at a functional level. METHODS: We analyzed a large set of skin fibroblasts derived from ARSACS patients, including both new and already published cases, carrying mutations of different type affecting diverse domains of the protein. RESULTS: We found that sacsin is almost absent in ARSACS patients, regardless of the nature of the mutation. As expected, we did not detect sacsin in patients with truncating mutations. Interestingly, we found it strikingly reduced or absent also in compound heterozygotes carrying diverse missense mutations. In this case, we excluded SACS mRNA decay, defective translation or faster post-translational degradation as possible causes of protein reduction. Conversely, our results demonstrate that nascent mutant sacsin protein undergoes cotranslational ubiquitination and degradation. DISCUSSION: Our results provide one mechanistic explanation for the lack of genotype-phenotype correlation in ARSACS. We also propose a new and unambiguous criterion for ARSACS diagnosis, that is based on the evaluation of sacsin level. Finally, we identified preemptive degradation of a mutant protein as a novel cause of a human disease.

3.
Nat Commun ; 12(1): 4878, 2021 08 12.
Artigo em Inglês | MEDLINE | ID: mdl-34385447

RESUMO

A postprandial increase of translation mediated by eukaryotic Initiation Factor 6 (eIF6) occurs in the liver. Its contribution to steatosis and disease is unknown. In this study we address whether eIF6-driven translation contributes to disease progression. eIF6 levels increase throughout the progression from Non-Alcoholic Fatty Liver Disease (NAFLD) to hepatocellular carcinoma. Reduction of eIF6 levels protects the liver from disease progression. eIF6 depletion blunts lipid accumulation, increases fatty acid oxidation (FAO) and reduces oncogenic transformation in vitro. In addition, eIF6 depletion delays the progression from NAFLD to hepatocellular carcinoma, in vivo. Mechanistically, eIF6 depletion reduces the translation of transcription factor C/EBPß, leading to a drop in biomarkers associated with NAFLD progression to hepatocellular carcinoma and preserves mitochondrial respiration due to the maintenance of an alternative mTORC1-eIF4F translational branch that increases the expression of transcription factor YY1. We provide proof-of-concept that in vitro pharmacological inhibition of eIF6 activity recapitulates the protective effects of eIF6 depletion. We hypothesize the existence of a targetable, evolutionarily conserved translation circuit optimized for lipid accumulation and tumor progression.


Assuntos
Carcinoma Hepatocelular/genética , Neoplasias Hepáticas/genética , Hepatopatia Gordurosa não Alcoólica/genética , Fatores de Iniciação de Peptídeos/genética , Biossíntese de Proteínas/genética , Animais , Proteína beta Intensificadora de Ligação a CCAAT/genética , Proteína beta Intensificadora de Ligação a CCAAT/metabolismo , Carcinoma Hepatocelular/metabolismo , Linhagem Celular , Transformação Celular Neoplásica/efeitos dos fármacos , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/metabolismo , Clofazimina/farmacologia , Dieta Hiperlipídica/efeitos adversos , Progressão da Doença , Inativação Gênica , Humanos , Lipogênese/efeitos dos fármacos , Lipogênese/genética , Neoplasias Hepáticas/metabolismo , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Hepatopatia Gordurosa não Alcoólica/metabolismo , Obesidade/etiologia , Obesidade/genética , Obesidade/metabolismo , Fatores de Iniciação de Peptídeos/antagonistas & inibidores , Fatores de Iniciação de Peptídeos/metabolismo
4.
Cancer Res ; 80(21): 4693-4706, 2020 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-32963011

RESUMO

Multiple myeloma is a plasma cell neoplasm characterized by the production of unfolded immunoglobulins, which cause endoplasmic reticulum (ER) stress and sensitivity to proteasome inhibition. The genomic landscape of multiple myeloma is characterized by the loss of several genes rarely mutated in other cancers that may underline specific weaknesses of multiple myeloma cells. One of these is FAM46C that is lost in more than 10% of patients with multiple myeloma. We show here that FAM46C is part of a new complex containing the ER-associated protein FNDC3A, which regulates trafficking and secretion and, by impairing autophagy, exacerbates proteostatic stress. Reconstitution of FAM46C in multiple myeloma cells that had lost it induced apoptosis and ER stress. Apoptosis was preceded by an increase of intracellular aggregates, which was not linked to increased translation of IgG mRNA, but rather to impairment of autophagy. Biochemical analysis showed that FAM46C requires interaction with ER bound protein FNDC3A to reside in the cytoplasmic side of the ER. FNDC3A was lost in some multiple myeloma cell lines. Importantly, depletion of FNDC3A increased the fitness of FAM46C-expressing cells and expression of FNDC3A in cells that had lost it recapitulated the effects of FAM46C, inducing aggregates and apoptosis. FAM46C and FNDC3A formed a complex that modulates secretion routes, increasing lysosome exocytosis. The cellular landscape generated by FAM46C/FNDC3A expression predicted sensitivity to sphingosine kinase inhibition. These results suggest that multiple myeloma cells remodel their trafficking machinery to cope with ER stress. SIGNIFICANCE: This study identifies a new multiple myeloma-specific tumor suppressor complex that regulates autophagy and unconventional secretion, highlighting the sensitivity of multiple myeloma cells to the accumulation of protein aggregates.


Assuntos
Fibronectinas/metabolismo , Mieloma Múltiplo/patologia , Nucleotidiltransferases/metabolismo , Agregação Patológica de Proteínas/metabolismo , Animais , Autofagia/fisiologia , Genes Supressores de Tumor , Xenoenxertos , Humanos , Camundongos , Mieloma Múltiplo/genética , Mieloma Múltiplo/metabolismo , Nucleotidiltransferases/genética , Agregados Proteicos/fisiologia , Transporte Proteico/fisiologia , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/metabolismo
5.
Cells ; 9(1)2020 01 10.
Artigo em Inglês | MEDLINE | ID: mdl-31936702

RESUMO

Eukaryotic initiation factor 6 (eIF6) is necessary for the nucleolar biogenesis of 60S ribosomes. However, most of eIF6 resides in the cytoplasm, where it acts as an initiation factor. eIF6 is necessary for maximal protein synthesis downstream of growth factor stimulation. eIF6 is an antiassociation factor that binds 60S subunits, in turn preventing premature 40S joining and thus the formation of inactive 80S subunits. It is widely thought that eIF6 antiassociation activity is critical for its function. Here, we exploited and improved our assay for eIF6 binding to ribosomes (iRIA) in order to screen for modulators of eIF6 binding to the 60S. Three compounds, eIFsixty-1 (clofazimine), eIFsixty-4, and eIFsixty-6 were identified and characterized. All three inhibit the binding of eIF6 to the 60S in the micromolar range. eIFsixty-4 robustly inhibits cell growth, whereas eIFsixty-1 and eIFsixty-6 might have dose- and cell-specific effects. Puromycin labeling shows that eIF6ixty-4 is a strong global translational inhibitor, whereas the other two are mild modulators. Polysome profiling and RT-qPCR show that all three inhibitors reduce the specific translation of well-known eIF6 targets. In contrast, none of them affect the nucleolar localization of eIF6. These data provide proof of principle that the generation of eIF6 translational modulators is feasible.


Assuntos
Fatores de Iniciação de Peptídeos/metabolismo , Biossíntese de Proteínas , Subunidades Ribossômicas Maiores de Eucariotos/metabolismo , Linhagem Celular , Nucléolo Celular/efeitos dos fármacos , Nucléolo Celular/metabolismo , Sobrevivência Celular , Ensaio de Imunoadsorção Enzimática , Humanos , Iniciação Traducional da Cadeia Peptídica/efeitos dos fármacos , Polirribossomos/efeitos dos fármacos , Polirribossomos/metabolismo , Ligação Proteica/efeitos dos fármacos , Puromicina/farmacologia , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Reprodutibilidade dos Testes
6.
Front Genet ; 9: 533, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30498507

RESUMO

Ribosomes have been long considered as executors of the translational program. The fact that ribosomes can control the translation of specific mRNAs or entire cellular programs is often neglected. Ribosomopathies, inherited diseases with mutations in ribosomal factors, show tissue specific defects and cancer predisposition. Studies of ribosomopathies have paved the way to the concept that ribosomes may control translation of specific mRNAs. Studies in Drosophila and mice support the existence of heterogeneous ribosomes that differentially translate mRNAs to coordinate cellular programs. Recent studies have now shown that ribosomal activity is not only a critical regulator of growth but also of metabolism. For instance, glycolysis and mitochondrial function have been found to be affected by ribosomal availability. Also, ATP levels drop in models of ribosomopathies. We discuss findings highlighting the relevance of ribosome heterogeneity in physiological and pathological conditions, as well as the possibility that in rate-limiting situations, ribosomes may favor some translational programs. We discuss the effects of ribosome heterogeneity on cellular metabolism, tumorigenesis and aging. We speculate a scenario in which ribosomes are not only executors of a metabolic program but act as modulators.

7.
Nat Commun ; 9(1): 3921, 2018 09 20.
Artigo em Inglês | MEDLINE | ID: mdl-30237396

RESUMO

The original version of this Article contained an error in the spelling of the author Miriam Gaggianesi, which was incorrectly given as Miriam Giaggianesi. Furthermore, the affiliation details for Gabriella Gaudioso, Valentina Vaira, and Silvano Bosari incorrectly omitted 'Division of Pathology, Fondazione IRCCS Ca' Granda Ospedale Maggiore Policlinico, Milan, 20122, Italy'. Finally, the affiliation details for Alice Turdo, Miriam Gaggianesi, Aurora Chinnici and Elisa Lipari were incorrectly given as 'Dipartimento di Biotecnologie Mediche e Medicina Legale Sezione di Biochimica Medica, Facoltà di Medicina e Chirurgia, Policlinico "P.Giaccone", Università di Palermo, Palermo, 90127, Italy'. The correct affiliation is 'Department of Surgical, Oncological and Stomatological Sciences, University of Palermo, Palermo, 90127, Italy'. These errors have now been corrected in both the PDF and HTML versions of the Article.

8.
Cancer Res ; 78(20): 5741-5753, 2018 10 15.
Artigo em Inglês | MEDLINE | ID: mdl-30072395

RESUMO

The expression of miRNAs in cancer has been widely studied and has allowed the definition of oncomirs and oncosuppressors. We note that it is often underestimated that many mRNAs are expressed, but translationally silent. In spite of this, systematic identification of miRNAs in equilibrium with their target mRNAs on polysomes has not been widely exploited. To identify biologically active oncomirs, we performed a screen for miRNAs acting on the polysomes of malignant mesothelioma (MPM) cells. Only a small percentage of expressed miRNAs physically associated with polysomes. On polysomes, we identified miRNAs already characterized in MPM, as well as novel ones like miR-24-3p, which acted as a promigratory miRNA in all cancer cells tested. miR-24-3p positively regulated Rho-GTP activity, and inhibition of miR-24-3p reduced growth in MPM cells. Analysis of miR-24-3p common targets, in two mesothelioma cell lines, identified a common subset of downregulated genes. These same genes were downregulated during the progression of multiple cancer types. Among the specific targets of miR-24-3p was cingulin, a tight junction protein that inhibits Rho-GTP activity. Overexpression of miR-24-3p only partially abrogated cingulin mRNA, but completely abrogated cingulin protein, confirming its action via translational repression. We suggest that miR-24-3p is an oncomir and speculate that identification of polysome-associated miRNAs efficiently sorts out biologically active miRNAs from inactive ones.Significance: Subcellular localization of miRNAs may predict their role in cancer and identify novel oncogenic miRNAs involved in cancer progression.Graphical Abstract: http://cancerres.aacrjournals.org/content/canres/78/20/5741/F1.large.jpg Cancer Res; 78(20); 5741-53. ©2018 AACR.


Assuntos
Neoplasias Pulmonares/metabolismo , Mesotelioma/metabolismo , MicroRNAs/genética , Neoplasias/genética , Polirribossomos/metabolismo , Animais , Linhagem Celular Tumoral , Movimento Celular , Progressão da Doença , Perfilação da Expressão Gênica , Humanos , Neoplasias Pulmonares/genética , Proteínas de Membrana/metabolismo , Mesotelioma/genética , Mesotelioma Maligno , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCID , Proteínas dos Microfilamentos/metabolismo , Metástase Neoplásica , Ribossomos/metabolismo , Análise de Sequência de RNA , Cicatrização
9.
Nat Commun ; 9(1): 1024, 2018 03 09.
Artigo em Inglês | MEDLINE | ID: mdl-29523784

RESUMO

Breast cancer consists of highly heterogeneous tumors, whose cell of origin and driver oncogenes are difficult to be uniquely defined. Here we report that MYC acts as tumor reprogramming factor in mammary epithelial cells by inducing an alternative epigenetic program, which triggers loss of cell identity and activation of oncogenic pathways. Overexpression of MYC induces transcriptional repression of lineage-specifying transcription factors, causing decommissioning of luminal-specific enhancers. MYC-driven dedifferentiation supports the onset of a stem cell-like state by inducing the activation of de novo enhancers, which drive the transcriptional activation of oncogenic pathways. Furthermore, we demonstrate that the MYC-driven epigenetic reprogramming favors the formation and maintenance of tumor-initiating cells endowed with metastatic capacity. This study supports the notion that MYC-driven tumor initiation relies on cell reprogramming, which is mediated by the activation of MYC-dependent oncogenic enhancers, thus establishing a therapeutic rational for treating basal-like breast cancers.


Assuntos
Neoplasias da Mama/metabolismo , Epigênese Genética , Células-Tronco Neoplásicas/metabolismo , Proteínas Proto-Oncogênicas c-myc/genética , Proteínas Proto-Oncogênicas c-myc/metabolismo , Animais , Neoplasias da Mama/genética , Neoplasias da Mama/fisiopatologia , Carcinogênese , Linhagem Celular Tumoral , Reprogramação Celular , Elementos Facilitadores Genéticos , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Camundongos , Camundongos SCID , Células-Tronco Neoplásicas/citologia
10.
Data Brief ; 14: 653-658, 2017 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-28924581

RESUMO

The data described in this article are related to "High levels of eukaryotic Initiation Factor 6 (eIF6) are required for immune system homeostasis and for steering the glycolytic flux of TCR-stimulated CD4+ T cells in both mice and humans" (Manfrini et al., in press) [1]. eIF6 is a translation initiation factor required for ribosomal biogenesis (Sanvito et al., 1999) [2] and for proper translational initiation (Gallo and Manfrini, 2015; Miluzio et al., 2016) [3], [4] whose protein abundance requires tight regulation. Here we analyze by flow cytometry the effects of eIF6 depletion on proportions of specific innate and adaptive immune system subpopulations and on thymocyte maturation in mice.

11.
Dev Comp Immunol ; 77: 69-76, 2017 12.
Artigo em Inglês | MEDLINE | ID: mdl-28743432

RESUMO

Eukaryotic Initiation Factor 6 (eIF6) is required for 60S ribosomal subunit biogenesis and efficient initiation of translation. Intriguingly, in both mice and humans, endogenous levels of eIF6 are detrimental as they act as tumor and obesity facilitators, raising the question on the evolutionary pressure that maintains high eIF6 levels. Here we show that, in mice and humans, high levels of eIF6 are required for proper immune functions. First, eIF6 heterozygous (het) mice show an increased mortality during viral infection and a reduction of peripheral blood CD4+ Effector Memory T cells. In human CD4+ T cells, eIF6 levels rapidly increase upon T-cell receptor activation and drive the glycolytic switch and the acquisition of effector functions. Importantly, in CD4+ T cells, eIF6 levels control interferon-γ (IFN-γ) secretion without affecting proliferation. In conclusion, the immune system has a high evolutionary pressure for the maintenance of a dynamic and powerful regulation of the translational machinery.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Fatores de Iniciação em Eucariotos/metabolismo , Fatores de Iniciação de Peptídeos/metabolismo , Receptores de Antígenos de Linfócitos T/metabolismo , Viroses/imunologia , Animais , Células Cultivadas , Fatores de Iniciação em Eucariotos/genética , Glicólise , Homeostase , Humanos , Sistema Imunitário , Memória Imunológica , Interferon gama/metabolismo , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fatores de Iniciação de Peptídeos/genética , Transdução de Sinais
12.
PLoS Genet ; 13(1): e1006552, 2017 01.
Artigo em Inglês | MEDLINE | ID: mdl-28056084

RESUMO

Ribosomopathies are a family of inherited disorders caused by mutations in genes necessary for ribosomal function. Shwachman-Diamond Bodian Syndrome (SDS) is an autosomal recessive disease caused, in most patients, by mutations of the SBDS gene. SBDS is a protein required for the maturation of 60S ribosomes. SDS patients present exocrine pancreatic insufficiency, neutropenia, chronic infections, and skeletal abnormalities. Later in life, patients are prone to myelodisplastic syndrome and acute myeloid leukemia (AML). It is unknown why patients develop AML and which cellular alterations are directly due to the loss of the SBDS protein. Here we derived mouse embryonic fibroblast lines from an SbdsR126T/R126T mouse model. After their immortalization, we reconstituted them by adding wild type Sbds. We then performed a comprehensive analysis of cellular functions including colony formation, translational and transcriptional RNA-seq, stress and drug sensitivity. We show that: 1. Mutant Sbds causes a reduction in cellular clonogenic capability and oncogene-induced transformation. 2. Mutant Sbds causes a marked increase in immature 60S subunits, limited impact on mRNA specific initiation of translation, but reduced global protein synthesis capability. 3. Chronic loss of SBDS activity leads to a rewiring of gene expression with reduced ribosomal capability, but increased lysosomal and catabolic activity. 4. Consistently with the gene signature, we found that SBDS loss causes a reduction in ATP and lactate levels, and increased susceptibility to DNA damage. Combining our data, we conclude that a cell-specific fragile phenotype occurs when SBDS protein drops below a threshold level, and propose a new interpretation of the disease.


Assuntos
Homeostase , Fenótipo , Proteínas/genética , Subunidades Ribossômicas Maiores de Eucariotos/genética , Trifosfato de Adenosina/metabolismo , Animais , Linhagem Celular , Transformação Celular Neoplásica , Dano ao DNA , Fibroblastos/metabolismo , Ácido Láctico/metabolismo , Camundongos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Subunidades Ribossômicas Maiores de Eucariotos/metabolismo
13.
Biochem Soc Trans ; 44(6): 1667-1673, 2016 12 15.
Artigo em Inglês | MEDLINE | ID: mdl-27913676

RESUMO

Over the past few years, there has been a growing interest in the interconnection between translation and metabolism. Important oncogenic pathways, like those elicited by c-Myc transcription factor and mTOR kinase, couple the activation of the translational machinery with glycolysis and fatty acid synthesis. Eukaryotic initiation factor 6 (eIF6) is a factor necessary for 60S ribosome maturation. eIF6 acts also as a cytoplasmic translation initiation factor, downstream of growth factor stimulation. eIF6 is up-regulated in several tumor types. Data on mice models have demonstrated that eIF6 cytoplasmic activity is rate-limiting for Myc-induced lymphomagenesis. In spite of this, eIF6 is neither transcriptionally regulated by Myc, nor post-transcriptionally regulated by mTOR. eIF6 stimulates a glycolytic and fatty acid synthesis program necessary for tumor growth. eIF6 increases the translation of transcription factors necessary for lipogenesis, such as CEBP/ß, ATF4 and CEBP/δ. Insulin stimulation leads to an increase in translation and fat synthesis blunted by eIF6 deficiency. Paradoxycally, long-term inhibition of eIF6 activity increases insulin sensitivity, suggesting that the translational activation observed upon insulin and growth factors stimulation acts as a feed-forward mechanism regulating lipid synthesis. The data on the role that eIF6 plays in cancer and in insulin sensitivity make it a tempting pharmacological target for cancers and metabolic diseases. We speculate that eIF6 inhibition will be particularly effective especially when mTOR sensitivity to rapamycin is abrogated by RAS mutations.


Assuntos
Fatores de Iniciação de Peptídeos/genética , Biossíntese de Proteínas , Serina-Treonina Quinases TOR/genética , Fatores de Transcrição/genética , Fator 4 Ativador da Transcrição/genética , Fator 4 Ativador da Transcrição/metabolismo , Animais , Proteína beta Intensificadora de Ligação a CCAAT/genética , Proteína beta Intensificadora de Ligação a CCAAT/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Humanos , Hipoglicemiantes/farmacologia , Insulina/farmacologia , Lipogênese/efeitos dos fármacos , Lipogênese/genética , Doenças Metabólicas/genética , Doenças Metabólicas/metabolismo , Camundongos , Modelos Genéticos , Neoplasias/genética , Neoplasias/metabolismo , Fatores de Iniciação de Peptídeos/metabolismo , Proteínas Proto-Oncogênicas c-myc/genética , Proteínas Proto-Oncogênicas c-myc/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Fatores de Transcrição/metabolismo
14.
Oncotarget ; 6(35): 37471-85, 2015 Nov 10.
Artigo em Inglês | MEDLINE | ID: mdl-26462016

RESUMO

eIF6 is an antiassociation factor that regulates the availability of active 80S. Its activation is driven by the RACK1/PKCß axis, in a mTORc1 independent manner. We previously described that eIF6 haploinsufficiency causes a striking survival in the Eµ-Myc mouse lymphoma model, with lifespans extended up to 18 months. Here we screen for eIF6 expression in human cancers. We show that Malignant Pleural Mesothelioma tumors (MPM) and a MPM cell line (REN cells) contain high levels of hyperphosphorylated eIF6. Enzastaurin is a PKC beta inhibitor used in clinical trials. We prove that Enzastaurin treatment decreases eIF6 phosphorylation rate, but not eIF6 protein stability. The growth of REN, in vivo, and metastasis are reduced by either Enzastaurin treatment or eIF6 shRNA. Molecular analysis reveals that eIF6 manipulation affects the metabolic status of malignant mesothelioma cells. Less glycolysis and less ATP content are evident in REN cells depleted for eIF6 or treated with Enzastaurin (Anti-Warburg effect). We propose that eIF6 is necessary for malignant mesothelioma growth, in vivo, and can be targeted by kinase inhibitors.


Assuntos
Biomarcadores Tumorais/metabolismo , Proliferação de Células , Fatores de Iniciação em Eucariotos/metabolismo , Mesotelioma/metabolismo , Neoplasias Pleurais/metabolismo , Trifosfato de Adenosina/metabolismo , Animais , Biomarcadores Tumorais/genética , Linhagem Celular Tumoral , Fatores de Iniciação em Eucariotos/genética , Regulação Neoplásica da Expressão Gênica , Glicólise , Humanos , Indóis/farmacologia , Mesotelioma/genética , Mesotelioma/patologia , Mesotelioma/terapia , Camundongos Endogâmicos NOD , Camundongos SCID , Fosforilação , Neoplasias Pleurais/genética , Neoplasias Pleurais/patologia , Neoplasias Pleurais/terapia , Proteína Quinase C beta/antagonistas & inibidores , Proteína Quinase C beta/metabolismo , Inibidores de Proteínas Quinases/farmacologia , Estabilidade Proteica , Interferência de RNA , Terapêutica com RNAi , Transdução de Sinais , Fatores de Tempo , Transfecção , Carga Tumoral , Regulação para Cima , Ensaios Antitumorais Modelo de Xenoenxerto
15.
Nat Commun ; 6: 8261, 2015 Sep 18.
Artigo em Inglês | MEDLINE | ID: mdl-26383020

RESUMO

Insulin regulates glycaemia, lipogenesis and increases mRNA translation. Cells with reduced eukaryotic initiation factor 6 (eIF6) do not increase translation in response to insulin. The role of insulin-regulated translation is unknown. Here we show that reduction of insulin-regulated translation in mice heterozygous for eIF6 results in normal glycaemia, but less blood cholesterol and triglycerides. eIF6 controls fatty acid synthesis and glycolysis in a cell autonomous fashion. eIF6 acts by exerting translational control of adipogenic transcription factors like C/EBPß, C/EBPδ and ATF4 that have G/C rich or uORF sequences in their 5' UTR. The outcome of the translational activation by eIF6 is a reshaping of gene expression with increased levels of lipogenic and glycolytic enzymes. Finally, eIF6 levels modulate histone acetylation and amounts of rate-limiting fatty acid synthase (Fasn) mRNA. Since obesity, type 2 diabetes, and cancer require a Fasn-driven lipogenic state, we propose that eIF6 could be a therapeutic target for these diseases.


Assuntos
Resistência à Insulina/genética , Metabolismo dos Lipídeos/genética , Fatores de Iniciação de Peptídeos/genética , Biossíntese de Proteínas/genética , RNA Mensageiro/metabolismo , Transcrição Genética/genética , Células 3T3 , Acetilação , Fator 4 Ativador da Transcrição/genética , Fator 4 Ativador da Transcrição/metabolismo , Adipócitos/metabolismo , Adipogenia/genética , Animais , Western Blotting , Proteína beta Intensificadora de Ligação a CCAAT/genética , Proteína beta Intensificadora de Ligação a CCAAT/metabolismo , Proteína delta de Ligação ao Facilitador CCAAT/genética , Proteína delta de Ligação ao Facilitador CCAAT/metabolismo , Eletroforese em Gel de Poliacrilamida , Ácido Graxo Sintases/genética , Ácido Graxo Sintases/metabolismo , Ácidos Graxos , Regulação da Expressão Gênica , Técnicas de Silenciamento de Genes , Glucose/metabolismo , Teste de Tolerância a Glucose , Glicogênio/metabolismo , Glicólise/genética , Células HEK293 , Hepatócitos/metabolismo , Código das Histonas , Humanos , Ácido Láctico/metabolismo , Lipogênese/genética , Fígado/diagnóstico por imagem , Fígado/metabolismo , Células-Tronco Mesenquimais , Camundongos , Oxirredução , Fatores de Iniciação de Peptídeos/metabolismo , Radiografia , Reação em Cadeia da Polimerase em Tempo Real , Reação em Cadeia da Polimerase Via Transcriptase Reversa
16.
Biochim Biophys Acta ; 1849(7): 830-5, 2015 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-25252159

RESUMO

Here we discuss the function of eukaryotic initiation factor 6 (eIF6; Tif6 in yeast). eIF6 binds 60S ribosomal subunits and blocks their joining to 40S. In this context, we propose that eIF6 impedes unproductive 80S formation, namely, the formation of 80S subunits without mRNA. Genetic evidence shows that eIF6 has a dual function: in yeast and mammals, nucleolar eIF6 is necessary for the biogenesis of 60S subunits. In mammals, cytoplasmic eIF6 is required for insulin and growth factor-stimulated translation. In contrast to other translation factors, eIF6 activity is not under mTOR control. The physiological significance of eIF6 impacts on cancer and on inherited Shwachman-Bodian-Diamond syndrome. eIF6 is overexpressed in specific human tumors. In a murine model of lymphomagenesis, eIF6 depletion leads to a striking increase of survival, without adverse effects. Shwachman-Bodian-Diamond syndrome is caused by loss of function of SBDS protein. In yeast, point mutations of Tif6, the yeast homolog of eIF6, rescue the quasi-lethal effect due to the loss of the SBDS homolog, Sdo1. We propose that eIF6 is a node regulator of ribosomal function and predict that prioritizing its pharmacological targeting will be of benefit in cancer and Shwachman-Bodian-Diamond syndrome. This article is part of a Special Issue entitled: Translation and Cancer.


Assuntos
Fatores de Iniciação em Eucariotos/metabolismo , Linfoma/metabolismo , Proteínas de Neoplasias/metabolismo , Fatores de Iniciação de Peptídeos/metabolismo , Biossíntese de Proteínas , Subunidades Ribossômicas Maiores de Eucariotos/metabolismo , Animais , Doenças da Medula Óssea/genética , Doenças da Medula Óssea/metabolismo , Doenças da Medula Óssea/patologia , Fatores de Iniciação em Eucariotos/genética , Insuficiência Pancreática Exócrina/genética , Insuficiência Pancreática Exócrina/metabolismo , Insuficiência Pancreática Exócrina/patologia , Humanos , Peptídeos e Proteínas de Sinalização Intracelular/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Lipomatose/genética , Lipomatose/metabolismo , Lipomatose/patologia , Linfoma/genética , Linfoma/patologia , Camundongos , Proteínas de Neoplasias/genética , Neoplasias Experimentais/genética , Neoplasias Experimentais/metabolismo , Neoplasias Experimentais/patologia , Fatores de Iniciação de Peptídeos/genética , Subunidades Ribossômicas Maiores de Eucariotos/genética , Síndrome de Shwachman-Diamond , Serina-Treonina Quinases TOR/genética , Serina-Treonina Quinases TOR/metabolismo , Transativadores
17.
Cell Cycle ; 10(20): 3441-6, 2011 Oct 15.
Artigo em Inglês | MEDLINE | ID: mdl-22031223

RESUMO

Ribosome biogenesis and translation can be simplified as the processes of generating ribosomes and their use for decoding mRNA into a protein. Ribosome biogenesis has been efficiently studied in unicellular organisms like the budding yeast, allowing us a deep and basic knowledge of this process in growing cells. Translation has been modeled in vitro and in unicellular organisms. These studies have given us an important insight into the mechanisms and evolutionarily conserved aspects of ribosome biology. However, we advocate the need of the direct study of these processes in multicellular organisms. Analysis of ribosome biogenesis and translation in vivo in Metazoa and mammalian models is emerging and unveils the unexpected consequences of perturbed ribosome biogenesis and translation. Here, we will describe how one factor, eIF6, plays a crucial role both in the generation of the large ribosomal subunit and its availability for translation. From there, we will make specific conclusions on the physiological relevance of eIF6 in 80S formation, cell cycle progression and disease, raising the point that the control of gene expression may occur at the unexpected level of the large ribosomal subunit. In the future, the modulation of eIF6 binding to the 60S may be pharmacologically exploited to reduce the growth of cancer cells or ameliorate the phenotype of SDS syndrome.


Assuntos
Doenças da Medula Óssea/metabolismo , Ciclo Celular/fisiologia , Transformação Celular Neoplásica/metabolismo , Fatores de Iniciação em Eucariotos/metabolismo , Insuficiência Pancreática Exócrina/metabolismo , Regulação da Expressão Gênica/fisiologia , Lipomatose/metabolismo , Biossíntese de Proteínas/fisiologia , Subunidades Ribossômicas Maiores/fisiologia , Animais , Humanos , Modelos Biológicos , Subunidades Ribossômicas Maiores/metabolismo , Síndrome de Shwachman-Diamond , Transdução de Sinais/fisiologia
18.
Cancer Cell ; 19(6): 765-75, 2011 Jun 14.
Artigo em Inglês | MEDLINE | ID: mdl-21665150

RESUMO

Eukaryotic Initiation Factor 6 (eIF6) controls translation by regulating 80S subunit formation. eIF6 is overexpressed in tumors. Here, we demonstrate that eIF6 inactivation delays tumorigenesis and reduces tumor growth in vivo. eIF6(+/-) mice resist to Myc-induced lymphomagenesis and have prolonged tumor-free survival and reduced tumor growth. eIF6(+/-) mice are also protected by p53 loss. Myc-driven lymphomas contain PKCßII and phosphorylated eIF6; eIF6 is phosphorylated by tumor-derived PKCßII, but not by the eIF4F activator mTORC1. Mutation of PKCßII phosphosite of eIF6 reduces tumor growth. Thus, eIF6 is a rate-limiting controller of initiation of translation, able to affect tumorigenesis and tumor growth. Modulation of eIF6 activity, independent from eIF4F complex, may lead to a therapeutical avenue in tumor therapy.


Assuntos
Transformação Celular Neoplásica/metabolismo , Linfoma/etiologia , Fatores de Iniciação de Peptídeos/fisiologia , Animais , Apoptose , Ciclo Celular , Citoplasma/metabolismo , Progressão da Doença , Genes myc , Linfoma/patologia , Camundongos , Camundongos Endogâmicos C57BL , Biossíntese de Proteínas , Proteína Quinase C/fisiologia , Proteína Quinase C beta , Proteína Supressora de Tumor p53/fisiologia
19.
EMBO Rep ; 10(5): 459-65, 2009 May.
Artigo em Inglês | MEDLINE | ID: mdl-19373251

RESUMO

Eukaryotic ribosome biogenesis and translation are linked processes that limit the rate of cell growth. Although ribosome biogenesis and translation are mainly controlled by distinct factors, eukaryotic initiation factor 6 (eIF6) has been found to regulate both processes. eIF6 is a necessary protein with a unique anti-association activity, which prevents the interaction of 40S ribosomal subunits with 60S subunits through its binding to 60S ribosomes. In the nucleolus, eIF6 is a component of the pre-ribosomal particles and is required for the biogenesis of 60S subunits, whereas in the cytoplasm it mediates translation downstream from growth factors. The translational activity of eIF6 could be due to its anti-association properties, which are regulated by post-translational modifications; whether this anti-association activity is required for the biogenesis and nuclear export of ribosomes is unknown. eIF6 is necessary for tissue-specific growth and oncogene-driven transformation, and could be a new rate-limiting step for the initiation of translation.


Assuntos
Fatores de Iniciação de Peptídeos/fisiologia , Biossíntese de Proteínas/fisiologia , Subunidades Ribossômicas Maiores de Eucariotos/metabolismo , Animais , Humanos , Modelos Biológicos , Fatores de Iniciação de Peptídeos/química , Fatores de Iniciação de Peptídeos/metabolismo , Biossíntese de Proteínas/genética
20.
Nature ; 455(7213): 684-8, 2008 Oct 02.
Artigo em Inglês | MEDLINE | ID: mdl-18784653

RESUMO

Cell growth and proliferation require coordinated ribosomal biogenesis and translation. Eukaryotic initiation factors (eIFs) control translation at the rate-limiting step of initiation. So far, only two eIFs connect extracellular stimuli to global translation rates: eIF4E acts in the eIF4F complex and regulates binding of capped messenger RNA to 40S subunits, downstream of growth factors, and eIF2 controls loading of the ternary complex on the 40S subunit and is inhibited on stress stimuli. No eIFs have been found to link extracellular stimuli to the activity of the large 60S ribosomal subunit. eIF6 binds 60S ribosomes precluding ribosome joining in vitro. However, studies in yeasts showed that eIF6 is required for ribosome biogenesis rather than translation. Here we show that mammalian eIF6 is required for efficient initiation of translation, in vivo. eIF6 null embryos are lethal at preimplantation. Heterozygous mice have 50% reduction of eIF6 levels in all tissues, and show reduced mass of hepatic and adipose tissues due to a lower number of cells and to impaired G1/S cell cycle progression. eIF6(+/-) cells retain sufficient nucleolar eIF6 and normal ribosome biogenesis. The liver of eIF6(+/-) mice displays an increase of 80S in polysomal profiles, indicating a defect in initiation of translation. Consistently, isolated hepatocytes have impaired insulin-stimulated translation. Heterozygous mouse embryonic fibroblasts recapitulate the organism phenotype and have normal ribosome biogenesis, reduced insulin-stimulated translation, and delayed G1/S phase progression. Furthermore, eIF6(+/-) cells are resistant to oncogene-induced transformation. Thus, eIF6 is the first eIF associated with the large 60S subunit that regulates translation in response to extracellular signals.


Assuntos
Transformação Celular Neoplásica , Iniciação Traducional da Cadeia Peptídica , Fatores de Iniciação de Peptídeos/metabolismo , Fase S , Tecido Adiposo/citologia , Animais , Peso Corporal , Divisão Celular/efeitos dos fármacos , Nucléolo Celular/metabolismo , Células Cultivadas , Citoplasma/metabolismo , Fibroblastos , Fase G1/efeitos dos fármacos , Heterozigoto , Insulina/farmacologia , Fígado/citologia , Fígado/crescimento & desenvolvimento , Camundongos , Células NIH 3T3 , Oncogenes/genética , Iniciação Traducional da Cadeia Peptídica/efeitos dos fármacos , Fatores de Iniciação de Peptídeos/deficiência , Fatores de Iniciação de Peptídeos/genética , Ribossomos/química , Ribossomos/metabolismo , Fase S/efeitos dos fármacos
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