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Bodega B.,CNR Institute of Molecular Genetics
Nature Structural and Molecular Biology | Year: 2017

The evolution of chromatin-based epigenetic cell memory may be driven not only by the necessity for cells to stably maintain transcription programs, but also by the need to recognize signals and allow plastic responses to environmental stimuli. The mechanistic role of the epigenome in adult postmitotic tissues, however, remains largely unknown. In vertebrates, two variants of the Polycomb repressive complex (PRC2–Ezh2 and PRC2–Ezh1) control gene silencing via methylation of histone H3 on Lys27 (H3K27me). Here we describe a reversible mechanism that involves a novel isoform of Ezh1 (Ezh1β). Ezh1β lacks the catalytic SET domain and acts in the cytoplasm of skeletal muscle cells to control nuclear PRC2–Ezh1 activity in response to atrophic oxidative stress, by regulating Eed assembly with Suz12 and Ezh1α (the canonical isoform) at their target genes. We report a novel PRC2–Ezh1 function that utilizes Ezh1β as an adaptive stress sensor in the cytoplasm, thus allowing postmitotic cells to maintain tissue integrity in response to environmental changes. © 2017 Nature Publishing Group, a division of Macmillan Publishers Limited. All Rights Reserved.


Grant
Agency: European Commission | Branch: FP7 | Program: ERC-AG | Phase: ERC-AG-LS7 | Award Amount: 2.50M | Year: 2014

Gene expression studies rely on high throughput techniques, which do not take in account conceptual limits. I will overcome this situation by exploiting two biological facts. First, RNAs that are important in tissue function are a subset of the global mass, but are always associated with the ribosomal machinery and as such should be identified. Second, gene expression is the outcome of dynamic fluctuations that with time create a unique expression pattern. We need to dynamically label cell populations that undergo stress and follow them to generate a gene expression signature. To achieve my goal, I will consider: 1. Translational stress generated by viral infection or accumulation of misfolded proteins; 2. human CD4\ T lymphocyte subsets which are key to orchestrate immune responses; 3. EIF6 model of metabolic reprogramming. 1. Activation of eIF2alpha phosphorylation by viral infection generates a translational response in which silent mRNAs containing upstream ORFs (uORF) are translated. I will exploit this observation to construct the first in vivo reporter model of translational stress. We will label genetically cells that have translational stress, to identify all the changes that a single cell undergoes after viral infection/accumulation of undegraded proteins. 2. I will selectively sequence for the first time mRNAs and ncRNAs associated with the ribosomal machinery in human cells with a defined functional status. 3. Spectacular data have shown that translation factor eIF6 regulates tumorigenesis by inducing a profound metabolic reprogramming. This observation suggests that, in vivo, translation acts upstream of transcription. We will model how a short translational input results in a complex epigenetic change. Significance: a revolution in finding biomarkers/drug targets. Generate a map of predictors of the process from stress to disease. Dscriminate biologically active sequences from background. Define how transient translation reshapes gene expression.


Aghemo A.,Am gliavacca Center For The Study Of Liver Disease | De Francesco R.,CNR Institute of Molecular Genetics
Hepatology | Year: 2013

Most direct-acting antivirals (DAAs) that are being developed as therapy against hepatitis C virus target the NS3/4A protease, the NS5A protein, and the NS5B polymerase. The latter enzyme offers different target sites: the catalytic domain for nucleos(t)ide analogues as well as a number of allosteric sites for nonnucleos(t)ide inhibitors. Two NS3/4A protease inhibitors have been approved recently, and more than 40 new NS3/4A, NS5A, or NS5B inhibitors are in development. These agents can achieve very high cure rates when combined with pegylated interferon-β and ribavirin and show promising clinical results when administered in all-oral combinations. In addition to the more canonical drug targets, new alternative viral targets for small-molecule drug development are emerging, such as p7 or NS4B and viral entry. Future research will need to define well-tolerated and cost-effective DAA combinations that provide the highest rates of viral eradication in all patients (including those with advanced liver disease), the broadest spectrum of action on viral genotypes showing minimal or no clinical resistance, and the shortest treatment duration. © 2013 American Association for the Study of Liver Diseases.


D'Adda di Fagagna F.,Instituto Firc Of Oncologia Molecolare Ifom | D'Adda di Fagagna F.,CNR Institute of Molecular Genetics
Trends in Cell Biology | Year: 2014

Historically, the role of cellular RNA has been subordinate and ancillary to DNA. Protein-coding mRNA conveys the information content of DNA, and transfer RNAs and ribosomal RNAs allow the polymerization of amino acids into proteins. The discovery of non-protein-coding RNAs (ncRNAs) provided an additional role for RNA in finely tuning DNA expression. However, it has recently become apparent that the safeguard of DNA integrity depends on small ncRNAs acting at the site of DNA lesions to signal the presence of DNA damage in the cell, and on the genes involved in their biogenesis to achieve accurate DNA repair. I review here evidence supporting a role for small ncRNAs, termed DNA damage-response RNAs (DDRNAs) or double-strand break (DSB)-induced RNAs (diRNAs), that are generated at sites of DNA damage and control the DNA damage response (DDR). I also discuss their biogenesis, potential mechanisms of action, and their relevance in cancer. © 2013 Elsevier Ltd.


Galietta L.J.V.,CNR Institute of Molecular Genetics
Pediatric Drugs | Year: 2013

Cystic fibrosis (CF), a severe genetic disease, is caused by mutations that alter the structure and function of CFTR, a plasma membrane channel permeable to chloride and bicarbonate. Defective anion transport in CF irreversibly damages the lungs, pancreas, liver, and other organs. CF mutations cause loss of CFTR function in multiple ways. In particular, class 3 mutations such as p.Gly551Asp strongly decrease the time spent by CFTR in the open state (gating defect). Instead, class 2 mutations impair the maturation of CFTR protein and its transport from the endoplasmic reticulum to the plasma membrane (trafficking defect). The deletion of phenylalanine 508 (p.Phe508del), the most frequent mutation among CF patients (70-90 %), destabilizes the CFTR protein, thus causing both a trafficking and a gating defect. These two defects can be overcome with drug-like molecules generically called correctors and potentiators, respectively. The potentiator Kalydeco™ (also known as Ivacaftor or VX-770), developed by Vertex Pharmaceuticals, has been recently approved by the US FDA and the European Medicines Agency (EMA) for the treatment of CF patients carrying at least one CFTR allele with the p.Gly551Asp mutation (2-5 % of all patients). In contrast, the corrector VX-809, which significantly improves p.Phe508del-CFTR trafficking in vitro, is still under study in clinical trials. Because of multiple defects caused by the p.Phe508del mutation, it is probable that rescue of the mutant protein will require combined treatment with correctors having different mechanisms of action. This review evaluates the status of experimental and clinical research in pharmacotherapy for the CF basic defect. © 2013 Springer International Publishing Switzerland.


Grant
Agency: European Commission | Branch: FP7 | Program: MC-IEF | Phase: FP7-PEOPLE-2013-IEF | Award Amount: 249.24K | Year: 2015

Phosphoinositides (PIs) are an important group of phospholipid messengers that regulate diverse cellular functions. A combination of mono, bis and tris-phosphorylation generates seven different PIs present in various subcellular compartments and importantly found within the nucleus. Nuclear PI levels change in response to diverse stimuli such as differentiation, growth factor receptor activation, growth responses in vivo, cell cycle progression, DNA damage, and cellular stressors. Chromatin forms a highly ordered repressive structure that limits access to the DNA sequence. It is widely held that disruption of chromatin compaction, which can be induced by histone tail modification is a rate-determining step in processes such as RNA transcription, DNA replication and DNA repair. Histones modification is reversible and its homeostasis is maintained by antagonistic enzymes that modify (writers) and those that remove the modification (erasers), while proteins that interact with the modifications (readers) act to interpret them. Differential states of histone modification appear to exhibit distinct distribution patterns in mammalian chromatin and appear to define different transcriptional landscapes. We have shown that many proteins that act as writers, readers and erasers of histone marks such as PHD finger containing proteins are receptors for nuclear PIs, suggesting a role for nuclear PI in chromatin modulation. In this study I will develop and use novel technologies and proprietary specific small molecular weight inhibitors of PI-kinases, to define the topographical environment of nuclear PI-signalling in relation to different chromatin landscapes and the genome. I will then determine if and how specific modulation of nuclear PIs in different chromatin landscapes functionally regulates transcriptional output. I expect to uncover a novel regulatory layer of chromatin modulation that depends on changes in nuclear PIs induced by extra and intra cellular cues.


Grant
Agency: European Commission | Branch: FP7 | Program: ERC-CG | Phase: ERC-CG-2013-LS7 | Award Amount: 2.00M | Year: 2014

Although tumor tissues can be infiltrated by T cells specific for tumor antigens, the effector functions of these lymphocytes are generally suppressed by CD4\ regulatory T cells (Tregs). Since tumor infiltrating Tregs can display function heterogeneity, depending on both the tumor type and the inflammatory milieu, only inhibition of the right Treg activity should result in the unleash of an effective anti-tumor T cell responses. Experimental plan: To identify the Tregs that truly inhibit anti-tumor T cells, we will profile by RNA-Seq the transcriptome of Tregs infiltrating both tumor and healthy tissues. In particular, we will focus on LncRNAs and the gene networks they modulate, since they have recently emerged as relevant epigenetic regulators of cell differentiation and identity. We will exploit this new knowledge to create a panel of regulatory transcripts, which will be assessed at single cell level on tumor infiltrating Tregs, so to determine the association of specific transcripts with different Treg populations. Since downregulation of specific lncRNAs might be an efficient way to inhibit the unwanted Tregs at tumor sites, we aim at targeting lncRNAs uniquely expressed in these Tregs and propose to develop AsiCs, chimeric molecules composed by an aptamer, single stranded oligonucleotides that bind to cell surface markers, and a siRNA, short RNAs downregulating specific lncRNAs. Deliverables and conclusions: this proposal will provide new knowledge on tumor infiltrating Tregs possibly allowing definition of molecular signatures of Tregs with either positive or negative effects on antitumor T cell responses. Moreover, we will develop new molecules that specifically target lncRNAs of interest and that will help identifying new antitumor therapeutic targets. In conclusion, the possibility to modulate Tregs effector functions may not only offer new anti-tumor therapy but more in general may be relevant to any immunomodulatory therapeutic strategies.


A method for diagnosing or prognosticating hepatocellular carcinoma, also in the early stages, or for assessing the risk of developing hepatocellular carcinoma, or for monitoring the effectiveness of an anti-tumour therapy against hepatocellular carcinoma by measuring the expression level of at least one miRNA gene product in a peripheral blood sample or in a biological fluid sample. Said method comprises measuring, in an isolated sample of peripheral blood or biological fluid, the expression level of at least one miRNA gene product, and comparing said measured expression level with a reference level. Such method can also be used to diagnose or assess the risk of developing liver cirrhosis in patients affected by chronic hepatitis, or to prognosticate the evolution of cirrhosis in patients affected by cirrhosis, or to monitor the effectiveness of a pharmacological therapy against liver cirrhosis.


Patent
CNR Institute of Molecular Genetics | Date: 2015-01-28

The present invention relates to miR-122 and miR-885-5p and their use in diagnostic and/or prognostic methods of liver diseases as well as modulators of miR-885-Sp for the treatment and/or prevention of a liver disease.


Patent
CNR Institute of Molecular Genetics | Date: 2013-04-10

The present invention relates to a biomarker of immunity response for use in monitoring the acquired immunity of an immunized subject, to an in vitro method and a kit for monitoring the acquired immunity of an immunized subject.

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