RESUMO
Changes in DNA methylation are among the best-documented epigenetic alterations accompanying organismal aging. However, whether and how altered DNA methylation is causally involved in aging have remained elusive. GADD45α (growth arrest and DNA damage protein 45A) and ING1 (inhibitor of growth family member 1) are adapter proteins for site-specific demethylation by TET (ten-eleven translocation) methylcytosine dioxygenases. Here we show that Gadd45a/Ing1 double-knockout mice display segmental progeria and phenocopy impaired energy homeostasis and lipodystrophy characteristic of Cebp (CCAAT/enhancer-binding protein) mutants. Correspondingly, GADD45α occupies C/EBPß/δ-dependent superenhancers and, cooperatively with ING1, promotes local DNA demethylation via long-range chromatin loops to permit C/EBPß recruitment. The results indicate that enhancer methylation can affect aging and imply that C/EBP proteins play an unexpected role in this process. Our study suggests a causal nexus between DNA demethylation, metabolism, and organismal aging.
Assuntos
Senilidade Prematura/genética , Envelhecimento/genética , Proteínas Estimuladoras de Ligação a CCAAT/genética , Proteínas Estimuladoras de Ligação a CCAAT/metabolismo , Proteínas de Ciclo Celular/metabolismo , Desmetilação do DNA , Proteína 1 Inibidora do Crescimento/metabolismo , Proteínas Nucleares/metabolismo , Animais , Proteínas de Ciclo Celular/genética , Células Cultivadas , Homeostase/genética , Proteína 1 Inibidora do Crescimento/genética , Lipodistrofia/genética , Camundongos , Camundongos Knockout , Proteínas Nucleares/genéticaRESUMO
Colorectal cancer (CRC) is the third most prevalent cancer in the world, yet the sensitivity and specificity of biomarkers for CRC diagnosis are insufficient. In the present study, we performed a protein microarray screening method to identify antibody markers for CRC. Inhibitor of growth family 1 (ING1) was identified as a candidate tumor antigen for CRC using protein microarrays (ProtoArray). Subsequent amplified luminescence proximity homogeneous assay-linked immunosorbent assay using recombinant ING1 protein showed that the serum levels of anti-ING1 antibodies were increased not only in patients with CRC but also in those with esophageal cancer (EC), gastric cancer (GC), breast cancer (BrC), and pancreatic cancer (PC) compared with those of healthy donors (HDs). Antibodies against the ING1 amino acids between 239 and 253 were present at significantly higher levels in patients with CRC than in those with EC, GC, BrC, or PC. Anti-ING1 antibody levels were significantly higher in the patients with CRC at any stages than in the HDs. Immunohistochemical staining revealed higher expression of ING1 protein in CRC cells than in the adjacent normal tissues. In luciferase reporter assays using a CRC cell line, ING1 augmented p53-mediated NOXA promoter activity but attenuated p53-stimulated Bax, p21, and PUMA promoter activities. Consequently, serum anti-ING1 antibodies can be used for sensitive and specific diagnoses of CRC.
Assuntos
Neoplasias Colorretais , Proteínas Supressoras de Tumor , Humanos , Proteína 1 Inibidora do Crescimento/metabolismo , Proteínas Supressoras de Tumor/genética , Proteínas Supressoras de Tumor/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Proteínas Nucleares/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Autoanticorpos , Neoplasias Colorretais/diagnósticoRESUMO
BACKGROUND: Enhancers are distal cis-regulatory elements required for cell-specific gene expression and cell fate determination. In cancer, enhancer variation has been proposed as a major cause of inter-patient heterogeneity-however, most predicted enhancer regions remain to be functionally tested. METHODS: We analyzed 132 epigenomic histone modification profiles of 18 primary gastric cancer (GC) samples, 18 normal gastric tissues, and 28 GC cell lines using Nano-ChIP-seq technology. We applied Capture-based Self-Transcribing Active Regulatory Region sequencing (CapSTARR-seq) to assess functional enhancer activity. An Activity-by-contact (ABC) model was employed to explore the effects of histone acetylation and CapSTARR-seq levels on enhancer-promoter interactions. RESULTS: We report a comprehensive catalog of 75,730 recurrent predicted enhancers, the majority of which are GC-associated in vivo (> 50,000) and associated with lower somatic mutation rates inferred by whole-genome sequencing. Applying CapSTARR-seq to the enhancer catalog, we observed significant correlations between CapSTARR-seq functional activity and H3K27ac/H3K4me1 levels. Super-enhancer regions exhibited increased CapSTARR-seq signals compared to regular enhancers, even when decoupled from native chromatin contexture. We show that combining histone modification and CapSTARR-seq functional enhancer data improves the prediction of enhancer-promoter interactions and pinpointing of germline single nucleotide polymorphisms (SNPs), somatic copy number alterations (SCNAs), and trans-acting TFs involved in GC expression. We identified cancer-relevant genes (ING1, ARL4C) whose expression between patients is influenced by enhancer differences in genomic copy number and germline SNPs, and HNF4α as a master trans-acting factor associated with GC enhancer heterogeneity. CONCLUSIONS: Our results indicate that combining histone modification and functional assay data may provide a more accurate metric to assess enhancer activity than either platform individually, providing insights into the relative contribution of genetic (cis) and regulatory (trans) mechanisms to GC enhancer functional heterogeneity.
Assuntos
Elementos Facilitadores Genéticos , Epigenômica , Neoplasias Gástricas/genética , Fatores de Ribosilação do ADP/genética , Fatores de Ribosilação do ADP/metabolismo , Acetilação , Linhagem Celular Tumoral , Proliferação de Células , Cromatina , Regulação Neoplásica da Expressão Gênica , Genômica , Histonas/metabolismo , Humanos , Proteína 1 Inibidora do Crescimento/genética , Proteína 1 Inibidora do Crescimento/metabolismo , Oncogenes , Regiões Promotoras Genéticas , RNA-Seq , Transcriptoma , Sequenciamento Completo do GenomaRESUMO
Approximately 90% of cancer-related deaths can be attributed to a tumour's ability to spread. We have identified CG7379, the fly orthologue of human ING1, as a potent invasion suppressor. ING1 is a type II tumour suppressor with well-established roles in the transcriptional regulation of genes that control cell proliferation, response to DNA damage, oncogene-induced senescence and apoptosis. Recent work suggests a possible role for ING1 in cancer cell invasion and metastasis, but the molecular mechanism underlying this observation is lacking. Our results show that reduced expression of CG7379 promotes invasion in vivo in Drosophila, reduces the junctional localization of several adherens and septate junction components, and severely disrupts cell-cell junction architecture. Similarly, ING1 knockdown significantly enhances invasion in vitro and disrupts E-cadherin distribution at cell-cell junctions. A transcriptome analysis reveals that loss of ING1 affects the expression of several junctional and cytoskeletal modulators, confirming ING1 as an invasion suppressor and a key regulator of cell-cell junction integrity.
Assuntos
Neoplasias da Mama/prevenção & controle , Comunicação Celular , Proteínas de Drosophila/metabolismo , Regulação Neoplásica da Expressão Gênica , Proteína 1 Inibidora do Crescimento/metabolismo , Animais , Apoptose , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Proliferação de Células , Proteínas de Drosophila/genética , Drosophila melanogaster , Feminino , Humanos , Proteína 1 Inibidora do Crescimento/genética , Células MCF-7 , Invasividade Neoplásica , TranscriptomaRESUMO
Epigenetic regulation of activity-induced gene expression involves multiple levels of molecular interaction, including histone and DNA modifications, as well as mechanisms of DNA repair. Here we demonstrate that the genome-wide deposition of inhibitor of growth family member 1 (ING1), which is a central epigenetic regulatory protein, is dynamically regulated in response to activity in primary cortical neurons. ING1 knockdown leads to decreased expression of genes related to synaptic plasticity, including the regulatory subunit of calcineurin, Ppp3r1. In addition, ING1 binding at a site upstream of the transcription start site (TSS) of Ppp3r1 depends on yet another group of neuroepigenetic regulatory proteins, the Piwi-like family, which are also involved in DNA repair. These findings provide new insight into a novel mode of activity-induced gene expression, which involves the interaction between different epigenetic regulatory mechanisms traditionally associated with gene repression and DNA repair.
Assuntos
Córtex Cerebral/metabolismo , Epigênese Genética , Proteína 1 Inibidora do Crescimento/metabolismo , Neurônios/metabolismo , Animais , Proteínas Argonautas/metabolismo , Proteínas de Ligação ao Cálcio , Células Cultivadas , Fibroblastos/metabolismo , Regulação da Expressão Gênica , Técnicas de Silenciamento de Genes , Humanos , Proteína 1 Inibidora do Crescimento/genética , Peptídeos e Proteínas de Sinalização Intracelular/metabolismo , Camundongos Endogâmicos C57BL , Proteínas Musculares/metabolismo , RNA Mensageiro/metabolismoRESUMO
Glioblastoma is the most common and aggressive primary brain tumor in adults. New drug design and development is still a major challenge for glioma treatment. Increasing evidence has shown that nitazoxanide, an antiprotozoal drug, has a novel antitumor role in various tumors and exhibits multiple molecular functions, especially autophagic regulation. However, whether nitazoxanide-associated autophagy has an antineoplastic effect in glioma remains unclear. Here, we aimed to explore the underlying molecular mechanism of nitazoxanide in glioblastoma. Our results showed that nitazoxanide suppressed cell growth and induced cell cycle arrest in glioblastoma by upregulating ING1 expression with a favorable toxicity profile. Nitazoxanide inhibited autophagy through blockage of late-stage lysosome acidification, resulting in decreased cleavage of ING1. A combination with chloroquine or Torin1 enhanced or impaired the chemotherapeutic effect of nitazoxanide in glioblastoma cells. Taken together, these findings indicate that nitazoxanide as an autophagy inhibitor induces cell cycle arrest in glioblastoma via upregulated ING1 due to increased transcription and decreased post-translational degradation by late-stage autophagic inhibition.
Assuntos
Antiprotozoários/farmacologia , Autofagia/efeitos dos fármacos , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Glioblastoma/tratamento farmacológico , Glioblastoma/metabolismo , Proteína 1 Inibidora do Crescimento/metabolismo , Tiazóis/farmacologia , Animais , Apoptose/efeitos dos fármacos , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/metabolismo , Linhagem Celular , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Cloroquina/farmacologia , Glioma/tratamento farmacológico , Glioma/metabolismo , Células Endoteliais da Veia Umbilical Humana , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Naftiridinas/farmacologia , Nitrocompostos , Processamento de Proteína Pós-Traducional/efeitos dos fármacos , Transcrição Gênica/efeitos dos fármacos , Regulação para Cima/efeitos dos fármacosRESUMO
Deregulation of precursor mRNA splicing is associated with many illnesses and has been linked to age-related chronic diseases. Here we review recent progress documenting how defects in the machinery that performs intron removal and controls splice site selection contribute to cellular senescence and organismal aging. We discuss the functional association linking p53, IGF-1, SIRT1, and ING-1 splice variants with senescence and aging, and review a selection of splicing defects occurring in accelerated aging (progeria), vascular aging, and Alzheimer's disease. Overall, it is becoming increasingly clear that changes in the activity of splicing factors and in the production of key splice variants can impact cellular senescence and the aging phenotype.
Assuntos
Envelhecimento/genética , Processamento Alternativo , Doença de Alzheimer/genética , Progéria/genética , Precursores de RNA/genética , RNA Mensageiro/genética , Envelhecimento/metabolismo , Doença de Alzheimer/metabolismo , Doença de Alzheimer/fisiopatologia , Animais , Senescência Celular/genética , Éxons , Regulação da Expressão Gênica no Desenvolvimento , Humanos , Proteína 1 Inibidora do Crescimento/genética , Proteína 1 Inibidora do Crescimento/metabolismo , Fator de Crescimento Insulin-Like I/genética , Fator de Crescimento Insulin-Like I/metabolismo , Íntrons , Progéria/metabolismo , Progéria/fisiopatologia , Precursores de RNA/metabolismo , Fatores de Processamento de RNA/genética , Fatores de Processamento de RNA/metabolismo , RNA Mensageiro/metabolismo , Sirtuína 1/genética , Sirtuína 1/metabolismo , Proteína Supressora de Tumor p53/genética , Proteína Supressora de Tumor p53/metabolismoRESUMO
Inhibitor of growth 2 (ING2) is involved in chromatin remodeling and it has previously been suggested that ING2 may regulate gene expression. The authors previously identified matrix metalloproteinase 13 (MMP13) as a target gene of ING2 in colorectal cancer. The aim of the present study was to identify novel genes regulated by ING2 and histone deacetylase 1 (HDAC1) and to clarify the biological significance of the ING2 structure. The present study generated the point mutant constructs of ING2 and deletion constructs consisting of partial ING2 to investigate the effect on gene expression and verify the interaction with HDAC1, mSin3A and sap30. A microarray was performed to find novel ING2/HDAC1 target genes using cell cooverexpression of ING2 and HDAC1. Plasminogen activator inhibitor1 (PAI1) was upregulated with overexpression of ING1b and ING2. The mutation of the PHD domain at 218 significantly attenuated the MMP13 and PAI1 expression, whereas the mutation at 224 resulted in increased expression. Furthermore, the expression levels were slightly reduced by the mutation of the Cterminal. The lack of the PHD domain and the Cterminal in ING2 resulted in a decreased ability to induce gene expression. The Cterminal with PHD domain, which lacked the Nterminal, maintained the transactive function for regulating the target genes. In addition to MMP13 and PAI1, eight genes [heat shock protein family A member 1A (HSPA1A), MIR73 host gene, chorionic somatomammotropin hormone 1, growth arrest and DNA damage inducible b, dehydrogenase/reductase 2, galectin 1, myosin light chain 1, and VGF nerve growth factor inducible] were demonstrated to be associated with ING2/HDAC1. The present study demonstrated that ING2/HDAC1 regulated PAI1 and HSPA1A expression and the PHD domain and the Cterminal of ING2, which are binding sites of HDAC1 and mSin3A, are essential regions for the regulation of gene expression.
Assuntos
Proteínas de Choque Térmico HSP70/metabolismo , Histona Desacetilase 1/metabolismo , Proteínas de Homeodomínio/metabolismo , Inibidor 1 de Ativador de Plasminogênio/metabolismo , Receptores Citoplasmáticos e Nucleares/metabolismo , Proteínas Repressoras/metabolismo , Proteínas Supressoras de Tumor/metabolismo , Células HEK293 , Proteínas de Choque Térmico HSP70/genética , Histona Desacetilase 1/genética , Histona Desacetilases/genética , Histona Desacetilases/metabolismo , Proteínas de Homeodomínio/química , Proteínas de Homeodomínio/genética , Humanos , Proteína 1 Inibidora do Crescimento/química , Proteína 1 Inibidora do Crescimento/genética , Proteína 1 Inibidora do Crescimento/metabolismo , Metaloproteinase 13 da Matriz/genética , Metaloproteinase 13 da Matriz/metabolismo , Metaloproteinase 2 da Matriz/metabolismo , Dedos de Zinco PHD , Plasmídeos/genética , Plasmídeos/metabolismo , Inibidor 1 de Ativador de Plasminogênio/genética , Mutação Puntual , Análise Serial de Proteínas , Receptores Citoplasmáticos e Nucleares/química , Receptores Citoplasmáticos e Nucleares/genética , Proteínas Repressoras/genética , Complexo Correpressor Histona Desacetilase e Sin3 , Proteínas Supressoras de Tumor/química , Proteínas Supressoras de Tumor/genética , Regulação para CimaRESUMO
We have studied the effect of inhibition of IRE1 (inositol requiring enzyme 1), which is a central mediator of endoplasmic reticulum stress and a controller of cell proliferation and tumor growth, on hypoxic regulation of the expression of different proliferation related genes in U87 glioma cells. It was shown that hypoxia leads to up-regulation of the expression of IL13RA2, CD24, ING1, ING2, ENDOG, and POLG genes and to down-regulation of KRT18, TRAPPC3, TSFM, and MTIF2 genes at the mRNA level in control glioma cells. Changes for ING1 and CD24 genes were more significant. At the same time, inhibition of IRE1 modifies the effect of hypoxia on the expression of all studied genes. In particular, it increases sensitivity to hypoxia of the expression of IL13RA2, TRAPPC3, ENDOG, and PLOG genes and suppresses the effect of hypoxia on the expression of ING1 gene. Additionally, it eliminates hypoxic regulation of KRT18, CD24, ING2, TSFM, and MTIF2 genes expressions and introduces sensitivity to hypoxia of the expression of BET1 gene in glioma cells. The present study demonstrates that hypoxia, which often contributes to tumor growth, affects the expression of almost all studied genes. Additionally, inhibition of IRE1 can both enhance and suppress the hypoxic regulation of these gene expressions in a gene specific manner and thus possibly contributes to slower glioma growth, but several aspects of this regulation must be further clarified.