RESUMO
For healthspan and lifespan, ERK, AMPK and mTORC1 represent critical pathways and inflammation is a centrally important hallmark1-7. Here we examined whether IL-11, a pro-inflammatory cytokine of the IL-6 family, has a negative effect on age-associated disease and lifespan. As mice age, IL-11 is upregulated across cell types and tissues to regulate an ERK-AMPK-mTORC1 axis to modulate cellular, tissue- and organismal-level ageing pathologies. Deletion of Il11 or Il11ra1 protects against metabolic decline, multi-morbidity and frailty in old age. Administration of anti-IL-11 to 75-week-old mice for 25 weeks improves metabolism and muscle function, and reduces ageing biomarkers and frailty across sexes. In lifespan studies, genetic deletion of Il11 extended the lives of mice of both sexes, by 24.9% on average. Treatment with anti-IL-11 from 75 weeks of age until death extends the median lifespan of male mice by 22.5% and of female mice by 25%. Together, these results demonstrate a role for the pro-inflammatory factor IL-11 in mammalian healthspan and lifespan. We suggest that anti-IL-11 therapy, which is currently in early-stage clinical trials for fibrotic lung disease, may provide a translational opportunity to determine the effects of IL-11 inhibition on ageing pathologies in older people.
Assuntos
Envelhecimento , Interleucina-11 , Longevidade , Transdução de Sinais , Animais , Feminino , Masculino , Camundongos , Envelhecimento/efeitos dos fármacos , Envelhecimento/genética , Envelhecimento/metabolismo , Envelhecimento/patologia , Proteínas Quinases Ativadas por AMP/metabolismo , Fragilidade/genética , Fragilidade/metabolismo , Fragilidade/prevenção & controle , Inflamação/metabolismo , Inflamação/tratamento farmacológico , Interleucina-11/antagonistas & inibidores , Interleucina-11/deficiência , Interleucina-11/genética , Interleucina-11/metabolismo , Subunidade alfa de Receptor de Interleucina-11/metabolismo , Subunidade alfa de Receptor de Interleucina-11/deficiência , Longevidade/efeitos dos fármacos , Longevidade/genética , Alvo Mecanístico do Complexo 1 de Rapamicina/metabolismo , Alvo Mecanístico do Complexo 1 de Rapamicina/antagonistas & inibidores , Camundongos Endogâmicos C57BL , Transdução de Sinais/efeitos dos fármacos , Humanos , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Músculo Esquelético/efeitos dos fármacos , Músculo Esquelético/fisiologiaRESUMO
Translated small open reading frames (smORFs) can have important regulatory roles and encode microproteins, yet their genome-wide identification has been challenging. We determined the ribosome locations across six primary human cell types and five tissues and detected 7,767 smORFs with translational profiles matching those of known proteins. The human genome was found to contain highly cell-type- and tissue-specific smORFs and a subset that encodes highly conserved amino acid sequences. Changes in the translational efficiency of upstream-encoded smORFs (uORFs) and the corresponding main ORFs predominantly occur in the same direction. Integration with 456 mass-spectrometry datasets confirms the presence of 603 small peptides at the protein level in humans and provides insights into the subcellular localization of these small proteins. This study provides a comprehensive atlas of high-confidence translated smORFs derived from primary human cells and tissues in order to provide a more complete understanding of the translated human genome.
Assuntos
Regulação da Expressão Gênica , Ribossomos , Genoma Humano/genética , Humanos , Fases de Leitura Aberta/genética , Biossíntese de Proteínas , Proteínas/metabolismo , RNA/metabolismo , Ribossomos/genética , Ribossomos/metabolismoRESUMO
The three striatins (STRN, STRN3, STRN4) form the core of STRiatin-Interacting Phosphatase and Kinase (STRIPAK) complexes. These place protein phosphatase 2A (PP2A) in proximity to protein kinases thereby restraining kinase activity and regulating key cellular processes. Our aim was to establish if striatins play a significant role in cardiac remodelling associated with cardiac hypertrophy and heart failure. All striatins were expressed in control human hearts, with up-regulation of STRN and STRN3 in failing hearts. We used mice with global heterozygote gene deletion to assess the roles of STRN and STRN3 in cardiac remodelling induced by angiotensin II (AngII; 7 days). Using echocardiography, we detected no differences in baseline cardiac function or dimensions in STRN+/- or STRN3+/- male mice (8 weeks) compared with wild-type littermates. Heterozygous gene deletion did not affect cardiac function in mice treated with AngII, but the increase in left ventricle mass induced by AngII was inhibited in STRN+/- (but not STRN3+/-) mice. Histological staining indicated that cardiomyocyte hypertrophy was inhibited. To assess the role of STRN in cardiomyocytes, we converted the STRN knockout line for inducible cardiomyocyte-specific gene deletion. There was no effect of cardiomyocyte STRN knockout on cardiac function or dimensions, but the increase in left ventricle mass induced by AngII was inhibited. This resulted from inhibition of cardiomyocyte hypertrophy and cardiac fibrosis. The data indicate that cardiomyocyte striatin is required for early remodelling of the heart by AngII and identify the striatin-based STRIPAK system as a signalling paradigm in the development of pathological cardiac hypertrophy.
Assuntos
Angiotensina II , Cardiomegalia , Camundongos Knockout , Miócitos Cardíacos , Animais , Angiotensina II/farmacologia , Miócitos Cardíacos/metabolismo , Miócitos Cardíacos/patologia , Cardiomegalia/genética , Cardiomegalia/patologia , Cardiomegalia/metabolismo , Cardiomegalia/fisiopatologia , Masculino , Humanos , Proteínas Musculares/metabolismo , Proteínas Musculares/genética , Remodelação Ventricular , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Proteínas de Ligação a Calmodulina , Proteínas do Tecido NervosoRESUMO
MOTIVATION: The creation and analysis of gene regulatory networks have been the focus of bioinformatics research and underpins much of what is known about gene regulation. However, as a result of a bias in the availability of data types that are collected, the vast majority of gene regulatory network resources and tools have focused on either transcriptional regulation or protein-protein interactions. This has left other areas of regulation, for instance, translational regulation, vastly underrepresented despite them having been shown to play a critical role in both health and disease. RESULTS: In order to address this, we have developed CLIPreg, a package that integrates RNA, Ribo and CLIP- sequencing data in order to construct translational regulatory networks coordinated by RNA-binding proteins and micro-RNAs. This is the first tool of its type to be created, allowing for detailed investigation into a previously unseen layer of regulation. AVAILABILITY AND IMPLEMENTATION: CLIPreg is available at https://github.com/SGDDNB/CLIPreg. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.
Assuntos
Redes Reguladoras de Genes , MicroRNAs , RNA-Seq , Proteínas de Ligação a RNA , SoftwareRESUMO
Building a reference set of protein-coding open reading frames (ORFs) has revolutionized biological process discovery and understanding. Traditionally, gene models have been confirmed using cDNA sequencing and encoded translated regions inferred using sequence-based detection of start and stop combinations longer than 100 amino-acids to prevent false positives. This has led to small ORFs (smORFs) and their encoded proteins left un-annotated. Ribo-seq allows deciphering translated regions from untranslated irrespective of the length. In this review, we describe the power of Ribo-seq data in detection of smORFs while discussing the major challenge posed by data-quality, -depth and -sparseness in identifying the start and end of smORF translation. In particular, we outline smORF cataloguing efforts in humans and the large differences that have arisen due to variation in data, methods and assumptions. Although current versions of smORF reference sets can already be used as a powerful tool for hypothesis generation, we recommend that future editions should consider these data limitations and adopt unified processing for the community to establish a canonical catalogue of translated smORFs.
Assuntos
Proteínas , Perfil de Ribossomos , Humanos , Proteínas/genética , Fases de Leitura Aberta , Biossíntese de Proteínas , MicropeptídeosRESUMO
Fibrosis is a common pathology in cardiovascular disease. In the heart, fibrosis causes mechanical and electrical dysfunction and in the kidney, it predicts the onset of renal failure. Transforming growth factor ß1 (TGFß1) is the principal pro-fibrotic factor, but its inhibition is associated with side effects due to its pleiotropic roles. We hypothesized that downstream effectors of TGFß1 in fibroblasts could be attractive therapeutic targets and lack upstream toxicity. Here we show, using integrated imaging-genomics analyses of primary human fibroblasts, that upregulation of interleukin-11 (IL-11) is the dominant transcriptional response to TGFß1 exposure and required for its pro-fibrotic effect. IL-11 and its receptor (IL11RA) are expressed specifically in fibroblasts, in which they drive non-canonical, ERK-dependent autocrine signalling that is required for fibrogenic protein synthesis. In mice, fibroblast-specific Il11 transgene expression or Il-11 injection causes heart and kidney fibrosis and organ failure, whereas genetic deletion of Il11ra1 protects against disease. Therefore, inhibition of IL-11 prevents fibroblast activation across organs and species in response to a range of important pro-fibrotic stimuli. These results reveal a central role of IL-11 in fibrosis and we propose that inhibition of IL-11 is a potential therapeutic strategy to treat fibrotic diseases.
Assuntos
Sistema Cardiovascular/metabolismo , Sistema Cardiovascular/patologia , Fibrose/metabolismo , Fibrose/patologia , Interleucina-11/metabolismo , Animais , Comunicação Autócrina , Células Cultivadas , Feminino , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Fibroblastos/patologia , Fibrose/induzido quimicamente , Coração , Humanos , Interleucina-11/antagonistas & inibidores , Interleucina-11/genética , Subunidade alfa de Receptor de Interleucina-11/deficiência , Subunidade alfa de Receptor de Interleucina-11/genética , Rim/patologia , Masculino , Camundongos , Camundongos Knockout , Pessoa de Meia-Idade , Miocárdio/metabolismo , Miocárdio/patologia , Escores de Disfunção Orgânica , Biossíntese de Proteínas , Fator de Crescimento Transformador beta1/metabolismo , Fator de Crescimento Transformador beta1/farmacologia , Transgenes/genéticaRESUMO
The extracellular signal-regulated kinase 1/2 (ERK1/2) cascade promotes cardiomyocyte hypertrophy and is cardioprotective, with the three RAF kinases forming a node for signal integration. Our aims were to determine if BRAF is relevant for human heart failure, whether BRAF promotes cardiomyocyte hypertrophy, and if Type 1 RAF inhibitors developed for cancer (that paradoxically activate ERK1/2 at low concentrations: the 'RAF paradox') may have the same effect. BRAF was up-regulated in heart samples from patients with heart failure compared with normal controls. We assessed the effects of activated BRAF in the heart using mice with tamoxifen-activated Cre for cardiomyocyte-specific knock-in of the activating V600E mutation into the endogenous gene. We used echocardiography to measure cardiac dimensions/function. Cardiomyocyte BRAFV600E induced cardiac hypertrophy within 10â d, resulting in increased ejection fraction and fractional shortening over 6 weeks. This was associated with increased cardiomyocyte size without significant fibrosis, consistent with compensated hypertrophy. The experimental Type 1 RAF inhibitor, SB590885, and/or encorafenib (a RAF inhibitor used clinically) increased ERK1/2 phosphorylation in cardiomyocytes, and promoted hypertrophy, consistent with a 'RAF paradox' effect. Both promoted cardiac hypertrophy in mouse hearts in vivo, with increased cardiomyocyte size and no overt fibrosis. In conclusion, BRAF potentially plays an important role in human failing hearts, activation of BRAF is sufficient to induce hypertrophy, and Type 1 RAF inhibitors promote hypertrophy via the 'RAF paradox'. Cardiac hypertrophy resulting from these interventions was not associated with pathological features, suggesting that Type 1 RAF inhibitors may be useful to boost cardiomyocyte function.
Assuntos
Cardiomegalia/patologia , Sistema de Sinalização das MAP Quinases/fisiologia , Miócitos Cardíacos/patologia , Proteínas Proto-Oncogênicas B-raf/fisiologia , Animais , Carbamatos/farmacologia , Carbamatos/toxicidade , Cardiomegalia/metabolismo , Tamanho Celular/efeitos dos fármacos , Células Cultivadas , Dimerização , Técnicas de Introdução de Genes , Insuficiência Cardíaca/patologia , Humanos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Mutação de Sentido Incorreto , Miócitos Cardíacos/efeitos dos fármacos , Miócitos Cardíacos/metabolismo , Mutação Puntual , Conformação Proteica/efeitos dos fármacos , Mapeamento de Interação de Proteínas , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas c-raf/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-raf/biossíntese , Ratos , Ratos Sprague-Dawley , Sulfonamidas/farmacologia , Sulfonamidas/toxicidadeRESUMO
Interleukin 11 (IL11) is upregulated in inflammatory conditions, where it is mostly believed to have anti-inflammatory activity. However, recent studies suggest instead that IL11 promotes inflammation by activating fibroblasts. Here, we assessed whether IL11 is pro- or anti-inflammatory in fibroblasts. Primary cultures of human kidney, lung or skin fibroblasts were stimulated with IL11 that resulted in the transient phosphorylation of signal transducer and activator of transcription 3 (STAT3) and the sustained activation of extracellular signal-regulated protein kinases (ERK). RNA sequencing over a time course of IL11 stimulation revealed a robust but short-lived transcriptional response that was enriched for gene set hallmarks of inflammation and characterized by the upregulation of SERPINB2, TNFRSF18, Interleukin 33 (IL33), CCL20, IL1RL1, CXCL3/5/8, ICAM1 and IL11 itself. IL33 was the most upregulated signaling factor (38-fold, p = 9.8 × 10-5), and IL1RL1, its cognate receptor, was similarly increased (18-fold, p = 1.1 × 10-34). In proteomic studies, IL11 triggered a proinflammatory secretome with the notable upregulation of IL8, IL6, MCP1, CCL20 and CXCL1/5/6, which are important chemotaxins for neutrophils, monocytes, and lymphocytes. IL11 induced IL33 expression across fibroblast types, and the inhibition of STAT3 but not of MEK/ERK prevented this. These data establish IL11 as pro-inflammatory with specific importance for priming the IL33 alarmin response in inflammatory fibroblasts across tissues.
Assuntos
Interleucina-11 , Interleucina-33 , Fibroblastos/metabolismo , Humanos , Inflamação/genética , Inflamação/metabolismo , Interleucina-11/genética , Interleucina-11/metabolismo , Interleucina-33/metabolismo , ProteômicaRESUMO
Titin-truncating variants (TTNtv) are the most common genetic cause of dilated cardiomyopathy. TTNtv occur in ~1% of the general population and causes subclinical cardiac remodeling in asymptomatic carriers. In rat models with either proximal or distal TTNtv, we previously showed altered cardiac metabolism at baseline and impaired cardiac function in response to stress. However, the molecular mechanism(s) underlying these effects remains unknown. In the current study, we used rat models of TTNtv to investigate the effect of TTNtv on autophagy and mitochondrial function, which are essential for maintaining cellular metabolic homeostasis and cardiac function. In both the proximal and distal TTNtv rat models, we found increased levels of LC3B-II and p62 proteins, indicative of diminished autophagic degradation. The accumulation of autophagosomes and p62 protein in cardiomyocytes was also demonstrated by electron microscopy and immunochemistry, respectively. Impaired autophagy in the TTNtv heart was associated with increased phosphorylation of mTOR and decreased protein levels of the lysosomal protease, cathepsin B. In addition, TTNtv hearts showed mitochondrial dysfunction, as evidenced by decreased oxygen consumption rate in cardiomyocytes, increased levels of reactive oxygen species and mitochondrial protein ubiquitination. We also observed increased acetylation of mitochondrial proteins associated with decreased NAD+/NADH ratio in the TTNtv hearts. mTORC1 inhibitor, rapamycin, was able to rescue the impaired autophagy in TTNtv hearts. In summary, TTNtv leads to impaired autophagy and mitochondrial function in the heart. These changes not only provide molecular mechanisms that underlie TTNtv-associated ventricular remodeling but also offer potential targets for its intervention.
Assuntos
Autofagia/genética , Cardiomiopatia Dilatada/genética , Conectina/genética , Mitocôndrias Cardíacas/metabolismo , Miócitos Cardíacos/metabolismo , Acetilação , Animais , Autofagossomos/metabolismo , Autofagossomos/ultraestrutura , Cardiomiopatia Dilatada/metabolismo , Catepsina B/metabolismo , Células Cultivadas , Conectina/metabolismo , Masculino , Proteínas Associadas aos Microtúbulos/metabolismo , Mitocôndrias Cardíacas/patologia , Proteínas Mitocondriais/metabolismo , NAD/análogos & derivados , NAD/metabolismo , Ratos , Espécies Reativas de Oxigênio/metabolismo , Deleção de Sequência , Proteína Sequestossoma-1/metabolismo , Serina-Treonina Quinases TOR/metabolismo , UbiquitinaçãoRESUMO
OBJECTIVES: Interleukin 11 (IL11) is highly upregulated in skin and lung fibroblasts from patients with systemic sclerosis (SSc). Here we tested whether IL11 is mechanistically linked with activation of human dermal fibroblasts (HDFs) from patients with SSc or controls. METHODS: We measured serum IL11 levels in volunteers and patients with early diffuse SSc and manipulated IL11 signalling in HDFs using gain- and loss-of-function approaches that we combined with molecular and cellular phenotyping. RESULTS: In patients with SSc, serum IL11 levels are elevated as compared with healthy controls. All transforming growth factor beta (TGFß) isoforms induced IL11 secretion from HDFs, which highly express IL11 receptor α-subunit and the glycoprotein 130 (gp130) co-receptor, suggestive of an autocrine loop of IL11 activity in HDFs. IL11 stimulated ERK activation in HDFs and resulted in HDF-to-myofibroblast transformation and extracellular matrix secretion. The pro-fibrotic action of IL11 in HDFs appeared unrelated to STAT3 activity, independent of TGFß upregulation and was not associated with phosphorylation of SMAD2/3. Inhibition of IL11 signalling using either a neutralizing antibody against IL11 or siRNA against IL11RA reduced TGFß-induced HDF proliferation, matrix production and cell migration, which was phenocopied by pharmacological inhibition of ERK. CONCLUSIONS: These data reveal that autocrine IL11-dependent ERK activity alone or downstream of TGFß stimulation promotes fibrosis phenotypes in dermal fibroblasts and suggest IL11 as a potential therapeutic target in SSc.
Assuntos
Regulação da Expressão Gênica , Subunidade alfa de Receptor de Interleucina-11/genética , Interleucina-11/sangue , Sistema de Sinalização das MAP Quinases/genética , RNA/genética , Escleroderma Sistêmico/sangue , Pele/patologia , Biomarcadores/sangue , Células Cultivadas , Ensaio de Imunoadsorção Enzimática , Humanos , Subunidade alfa de Receptor de Interleucina-11/biossíntese , Escleroderma Sistêmico/genética , Escleroderma Sistêmico/patologia , Transdução de SinaisRESUMO
Repetitive pulmonary injury causes fibrosis and inflammation that underlies chronic lung diseases such as idiopathic pulmonary fibrosis (IPF). Interleukin 11 (IL11) is important for pulmonary fibroblast activation but the contribution of fibroblast-specific IL11 activity to lung fibro-inflammation is not known. To address this gap in knowledge, we generated mice with loxP-flanked Il11ra1 and deleted the IL11 receptor in adult fibroblasts (CKO mice). In the bleomycin (BLM) model of lung fibrosis, CKO mice had reduced fibrosis, lesser fibroblast ERK activation, and diminished immune cell STAT3 phosphorylation. Following BLM injury, acute inflammation in CKO mice was similar to controls but chronic immune infiltrates and pro-inflammatory gene activation, including NF-kB phosphorylation, were notably reduced. Therapeutic prevention of IL11 activity with neutralizing antibodies mirrored the effects of genetic deletion of Il11ra1 in fibroblasts. These data reveal a new function for IL11 in pro-inflammatory lung fibroblasts and highlight the important contribution of the stroma to inflammation in pulmonary disease.
Assuntos
Fibroblastos/metabolismo , Inflamação/metabolismo , Subunidade alfa de Receptor de Interleucina-11/metabolismo , Interleucina-11/metabolismo , Fibrose Pulmonar/metabolismo , Animais , Bleomicina , Células Cultivadas , Doença Crônica , Fibroblastos/citologia , Fibroblastos/efeitos dos fármacos , Humanos , Inflamação/genética , Interleucina-11/farmacologia , Subunidade alfa de Receptor de Interleucina-11/genética , Camundongos Endogâmicos C57BL , Camundongos Knockout , Camundongos Transgênicos , NF-kappa B/metabolismo , Fosforilação , Fibrose Pulmonar/induzido quimicamente , Fibrose Pulmonar/genética , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genéticaRESUMO
BACKGROUND: Several genetic susceptibility loci associated with diabetic nephropathy have been documented, but no causative variants implying novel pathogenetic mechanisms have been elucidated. METHODS: We carried out whole-genome sequencing of a discovery cohort of Finnish siblings with type 1 diabetes who were discordant for the presence (case) or absence (control) of diabetic nephropathy. Controls had diabetes without complications for 15-37 years. We analyzed and annotated variants at genome, gene, and single-nucleotide variant levels. We then replicated the associated variants, genes, and regions in a replication cohort from the Finnish Diabetic Nephropathy study that included 3531 unrelated Finns with type 1 diabetes. RESULTS: We observed protein-altering variants and an enrichment of variants in regions associated with the presence or absence of diabetic nephropathy. The replication cohort confirmed variants in both regulatory and protein-coding regions. We also observed that diabetic nephropathy-associated variants, when clustered at the gene level, are enriched in a core protein-interaction network representing proteins essential for podocyte function. These genes include protein kinases (protein kinase C isoforms ε and ι) and protein tyrosine kinase 2. CONCLUSIONS: Our comprehensive analysis of a diabetic nephropathy cohort of siblings with type 1 diabetes who were discordant for kidney disease points to variants and genes that are potentially causative or protective for diabetic nephropathy. This includes variants in two isoforms of the protein kinase C family not previously linked to diabetic nephropathy, adding support to previous hypotheses that the protein kinase C family members play a role in diabetic nephropathy and might be attractive therapeutic targets.
Assuntos
Diabetes Mellitus Tipo 1/complicações , Nefropatias Diabéticas/genética , Sequenciamento Completo do Genoma/métodos , Adolescente , Adulto , Animais , Criança , Pré-Escolar , Diabetes Mellitus Tipo 1/genética , Feminino , Células HEK293 , Humanos , Masculino , Polimorfismo de Nucleotídeo Único , Proteína Quinase C/fisiologia , Irmãos , Adulto Jovem , Peixe-ZebraRESUMO
BACKGROUND: Fibrosis is a common pathology in many cardiac disorders and is driven by the activation of resident fibroblasts. The global posttranscriptional mechanisms underlying fibroblast-to-myofibroblast conversion in the heart have not been explored. METHODS: Genome-wide changes of RNA transcription and translation during human cardiac fibroblast activation were monitored with RNA sequencing and ribosome profiling. We then used RNA-binding protein-based analyses to identify translational regulators of fibrogenic genes. The integration with cardiac ribosome occupancy levels of 30 dilated cardiomyopathy patients demonstrates that these posttranscriptional mechanisms are also active in the diseased fibrotic human heart. RESULTS: We generated nucleotide-resolution translatome data during the transforming growth factor ß1-driven cellular transition of human cardiac fibroblasts to myofibroblasts. This identified dynamic changes of RNA transcription and translation at several time points during the fibrotic response, revealing transient and early-responder genes. Remarkably, about one-third of all changes in gene expression in activated fibroblasts are subject to translational regulation, and dynamic variation in ribosome occupancy affects protein abundance independent of RNA levels. Targets of RNA-binding proteins were strongly enriched in posttranscriptionally regulated genes, suggesting genes such as MBNL2 can act as translational activators or repressors. Ribosome occupancy in the hearts of patients with dilated cardiomyopathy suggested the same posttranscriptional regulatory network was underlying cardiac fibrosis. Key network hubs include RNA-binding proteins such as Pumilio RNA binding family member 2 (PUM2) and Quaking (QKI) that work in concert to regulate the translation of target transcripts in human diseased hearts. Furthermore, silencing of both PUM2 and QKI inhibits the transition of fibroblasts toward profibrotic myofibroblasts in response to transforming growth factor ß1. CONCLUSIONS: We reveal widespread translational effects of transforming growth factor ß1 and define novel posttranscriptional regulatory networks that control the fibroblast-to-myofibroblast transition. These networks are active in human heart disease, and silencing of hub genes limits fibroblast activation. Our findings show the central importance of translational control in fibrosis and highlight novel pathogenic mechanisms in heart failure.
Assuntos
Cardiopatias/genética , Cardiopatias/metabolismo , Miócitos Cardíacos/metabolismo , Miócitos Cardíacos/patologia , Biossíntese de Proteínas/genética , Proteínas de Ligação a RNA/genética , Células Cultivadas , Fibroblastos/metabolismo , Fibroblastos/patologia , Fibrose/genética , Fibrose/metabolismo , Fibrose/patologia , Perfilação da Expressão Gênica/métodos , Cardiopatias/patologia , Humanos , Análise de Sequência de RNA/métodos , Fator de Crescimento Transformador beta1/genética , Fator de Crescimento Transformador beta1/metabolismoRESUMO
BACKGROUND & AIMS: We studied the role of interleukin 11 (IL11) signaling in the pathogenesis of nonalcoholic steatohepatitis (NASH) using hepatic stellate cells (HSCs), hepatocytes, and mouse models of NASH. METHODS: We stimulated mouse and human fibroblasts, HSCs, or hepatocytes with IL11 and other cytokines and analyzed them by imaging, immunoblot, and functional assays and enzyme-linked immunosorbent assays. Mice were given injections of IL11. Mice with disruption of the interleukin 11 receptor subunit alpha1 gene (Il11ra1-/-) mice and Il11ra1+/+ mice were fed a high-fat methionine- and choline-deficient diet (HFMCD) or a Western diet with liquid fructose (WDF) to induce steatohepatitis; control mice were fed normal chow. db/db mice were fed with methionine- and choline-deficient diet for 12 weeks and C57BL/6 NTac were fed with HFMCD for 10 weeks or WDF for 16 weeks. Some mice were given intraperitoneal injections of anti-IL11 (X203), anti-IL11RA (X209), or a control antibody at different timepoints on the diets. Livers and blood were collected; blood samples were analyzed by biochemistry and liver tissues were analyzed by histology, RNA sequencing, immunoblots, immunohistochemistry, hydroxyproline, and mass cytometry time of flight assays. RESULTS: HSCs incubated with cytokines produced IL11, resulting in activation (phosphorylation) of ERK and expression of markers of fibrosis. Livers of mice given injections of IL11 became damaged, with increased markers of fibrosis, hepatocyte cell death and inflammation. Following the HFMCD or WDF, livers from Il11ra1-/- mice had reduced steatosis, fibrosis, expression of markers of inflammation and steatohepatitis, compared to and Il11ra1+/+ mice on the same diets. Depending on the time of administration of anti-IL11 or anti-IL11RA antibodies to wild-type mice on the HFMCD or WDF, or to db/db mice on the methionine and choline-deficient diet, the antibodies prevented, stopped, or reversed development of fibrosis and steatosis. Blood samples from Il11ra1+/+ mice fed the WDF and given injections of anti-IL11 or anti-IL11RA, as well as from Il11ra1-/- mice fed WDF, had lower serum levels of lipids and glucose than mice not injected with antibody or with disruption of Il11ra1. CONCLUSIONS: Neutralizing antibodies that block IL11 signaling reduce fibrosis, steatosis, hepatocyte death, inflammation and hyperglycemia in mice with diet-induced steatohepatitis. These antibodies also improve the cardiometabolic profile of mice and might be developed for the treatment of NASH.
Assuntos
Anticorpos Neutralizantes/farmacologia , Hepatite/prevenção & controle , Subunidade alfa de Receptor de Interleucina-11/metabolismo , Interleucina-11/antagonistas & inibidores , Cirrose Hepática Experimental/prevenção & controle , Fígado/efeitos dos fármacos , Hepatopatia Gordurosa não Alcoólica/prevenção & controle , Animais , Morte Celular/efeitos dos fármacos , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Fibroblastos/patologia , Células Estreladas do Fígado/efeitos dos fármacos , Células Estreladas do Fígado/metabolismo , Células Estreladas do Fígado/patologia , Hepatite/genética , Hepatite/metabolismo , Hepatite/patologia , Hepatócitos/efeitos dos fármacos , Hepatócitos/metabolismo , Hepatócitos/patologia , Humanos , Mediadores da Inflamação/metabolismo , Interleucina-11/metabolismo , Subunidade alfa de Receptor de Interleucina-11/deficiência , Subunidade alfa de Receptor de Interleucina-11/genética , Fígado/metabolismo , Fígado/patologia , Cirrose Hepática Experimental/genética , Cirrose Hepática Experimental/metabolismo , Cirrose Hepática Experimental/patologia , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout , Hepatopatia Gordurosa não Alcoólica/genética , Hepatopatia Gordurosa não Alcoólica/metabolismo , Hepatopatia Gordurosa não Alcoólica/patologia , Transdução de Sinais/efeitos dos fármacos , Células THP-1RESUMO
SUMMARY: To enable direct comparison of ancestry background in different studies, we developed LASER to estimate individual ancestry by placing either sezquenced or genotyped samples in a common ancestry space, regardless of the sequencing strategy or genotyping array used to characterize each sample. Here we describe the LASER server to facilitate application of the method to a wide range of genetic studies. The server provides genetic ancestry estimation for different geographic regions and user-friendly interactive visualization of the results. AVAILABILITY AND IMPLEMENTATION: The LASER server is freely accessible at http://laser.sph.umich.edu/. CONTACT: dtaliun@umich.edu or wangcl@gis.a-star.edu.sg. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.
Assuntos
Variação Genética , Filogeografia/métodos , Grupos Populacionais/genética , Análise de Sequência de DNA/métodos , Software , HumanosRESUMO
BACKGROUND: Untranslated regions (UTRs) are important mediators of post-transcriptional regulation. The length of UTRs and the composition of regulatory elements within them are known to vary substantially across genes, but little is known about the reasons for this variation in humans. Here, we set out to determine whether this variation, specifically in 5'UTRs, correlates with gene dosage sensitivity. RESULTS: We investigate 5'UTR length, the number of alternative transcription start sites, the potential for alternative splicing, the number and type of upstream open reading frames (uORFs) and the propensity of 5'UTRs to form secondary structures. We explore how these elements vary by gene tolerance to loss-of-function (LoF; using the LOEUF metric), and in genes where changes in dosage are known to cause disease. We show that LOEUF correlates with 5'UTR length and complexity. Genes that are most intolerant to LoF have longer 5'UTRs, greater TSS diversity, and more upstream regulatory elements than their LoF tolerant counterparts. We show that these differences are evident in disease gene-sets, but not in recessive developmental disorder genes where LoF of a single allele is tolerated. CONCLUSIONS: Our results confirm the importance of post-transcriptional regulation through 5'UTRs in tight regulation of mRNA and protein levels, particularly for genes where changes in dosage are deleterious and lead to disease. Finally, to support gene-based investigation we release a web-based browser tool, VuTR, that supports exploration of the composition of individual 5'UTRs and the impact of genetic variation within them.
Assuntos
Regiões 5' não Traduzidas , Fases de Leitura Aberta , Biossíntese de Proteínas , Humanos , Dosagem de Genes , Regulação da Expressão Gênica , Sítio de Iniciação de Transcrição , Processamento Alternativo , Conformação de Ácido NucleicoRESUMO
Our ability to understand and control stem cell biology is being augmented by developments on two fronts, our ability to collect more data describing cell state and our capability to comprehend these data using deep learning models. Here we consider the impact deep learning will have in the future of stem cell research. We explore the importance of generating data suitable for these methods, the requirement for close collaboration between experimental and computational researchers, and the challenges we face to do this fairly and effectively. Achieving this will ensure that the resulting deep learning models are biologically meaningful and computationally tractable.
Assuntos
Aprendizado Profundo , Pesquisa com Células-TroncoRESUMO
The kidney has large regenerative capacity, but this is compromised when kidney damage is excessive and renal tubular epithelial cells (TECs) undergo SNAI1-driven growth arrest. Here we investigate the role of IL11 in TECs, kidney injury and renal repair. IL11 stimulation of TECs induces ERK- and p90RSK-mediated GSK3ß inactivation, SNAI1 upregulation and pro-inflammatory gene expression. Mice with acute kidney injury upregulate IL11 in TECs leading to SNAI1 expression and kidney dysfunction, which is not seen in Il11 deleted mice or in mice administered a neutralizing IL11 antibody in either preemptive or treatment modes. In acute kidney injury, anti-TGFß reduces renal fibrosis but exacerbates inflammation and tubule damage whereas anti-IL11 reduces all pathologies. Mice with TEC-specific deletion of Il11ra1 have reduced pathogenic signaling and are protected from renal injury-induced inflammation, fibrosis, and failure. In a model of chronic kidney disease, anti-IL11 therapy promotes TEC proliferation and parenchymal regeneration, reverses fibroinflammation and restores renal mass and function. These data highlight IL11-induced mesenchymal transition of injured TECs as an important renal pathology and suggest IL11 as a therapeutic target for restoring stalled endogenous regeneration in the diseased kidney.
Assuntos
Injúria Renal Aguda , Anticorpos Neutralizantes , Interleucina-11 , Túbulos Renais , Nefrite , Regeneração , Insuficiência Renal Crônica , Animais , Camundongos , Injúria Renal Aguda/terapia , Fibrose , Subunidade alfa de Receptor de Interleucina-11/genética , Túbulos Renais/fisiologia , Nefrite/terapia , Interleucina-11/antagonistas & inibidores , Interleucina-11/fisiologia , Deleção de Genes , Anticorpos Neutralizantes/uso terapêutico , Insuficiência Renal Crônica/terapia , Modelos Animais de DoençasRESUMO
Loss of function (LOF) in IL11RA infers IL11 signaling as important for fertility, fibrosis, inflammation and incompletely penetrant craniosynostosis. The impact of LOF in IL11 has not been characterized. We generated IL11 knockout (Il11-/-) mice that are born in expected ratios and have normal hematological profiles. Lung fibroblasts from Il11-/- mice are resistant to pro-fibrotic stimulation with TGFß1. Following bleomycin-induced lung injury, Il11-/- mice are protected from pulmonary fibrosis and exhibit lesser ERK, STAT3 and NF-kB activation, reduced Il1b, Timp1, Ccl2 and diminished IL6 expression, both at baseline and after injury: placing Il11 activity upstream of IL6 in this model. Il11-/- female mice are infertile. Unlike Il11ra1-/- mice, Il11-/- mice do not have craniosynostosis, have normal long bone mass and reduced body weights. These data further establish the role of IL11 signaling in lung fibrosis while suggesting that bone development abnormalities can be associated with mutation of IL11RA but not IL11, which may have implications for therapeutic targeting of IL11 signaling.
Assuntos
Craniossinostoses/complicações , Fertilidade , Inflamação/complicações , Inflamação/patologia , Subunidade alfa de Receptor de Interleucina-11/metabolismo , Interleucina-11/metabolismo , Pulmão/patologia , Animais , Bleomicina , Diferenciação Celular , Craniossinostoses/sangue , Feminino , Fibronectinas/metabolismo , Humanos , Infertilidade Feminina/sangue , Infertilidade Feminina/patologia , Inflamação/sangue , Metabolômica , Camundongos Knockout , Miofibroblastos/patologia , NF-kappa B/metabolismo , Fosforilação , Fibrose Pulmonar/sangue , Fibrose Pulmonar/complicações , Fibrose Pulmonar/patologia , Fator de Transcrição STAT3/metabolismo , Proteína Smad2RESUMO
Mito-SEPs are small open reading frame-encoded peptides that localize to the mitochondria to regulate metabolism. Motivated by an intriguing negative association between mito-SEPs and inflammation, here we screen for mito-SEPs that modify inflammatory outcomes and report a mito-SEP named "Modulator of cytochrome C oxidase during Inflammation" (MOCCI) that is upregulated during inflammation and infection to promote host-protective resolution. MOCCI, a paralog of the NDUFA4 subunit of cytochrome C oxidase (Complex IV), replaces NDUFA4 in Complex IV during inflammation to lower mitochondrial membrane potential and reduce ROS production, leading to cyto-protection and dampened immune response. The MOCCI transcript also generates miR-147b, which targets the NDUFA4 mRNA with similar immune dampening effects as MOCCI, but simultaneously enhances RIG-I/MDA-5-mediated viral immunity. Our work uncovers a dual-component pleiotropic regulation of host inflammation and immunity by MOCCI (C15ORF48) for safeguarding the host during infection and inflammation.