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Longobardi E.,Fondazione Instituto Firc Of Oncologia Molecolare Ifom | Penkov D.,Moscow State University | De Florian G.,Fondazione Instituto Firc Of Oncologia Molecolare Ifom | Blasi F.,Fondazione Instituto Firc Of Oncologia Molecolare Ifom
Developmental Dynamics | Year: 2014

TALE (three amino acids loop extension) homeodomain transcription factors are required in various steps of embryo development, in many adult physiological functions, and are involved in important pathologies. This review focuses on the PREP, MEIS, and PBX sub-families of TALE factors and aims at giving information on their biochemical properties, i.e., structure, interactors, and interaction surfaces. Members of the three sets of protein form dimers in which the common partner is PBX but they can also directly interact with other proteins forming higher-order complexes, in particular HOX. Finally, recent advances in determining the genome-wide DNA-binding sites of PREP1, MEIS1, and PBX1, and their partial correspondence with the binding sites of some HOX proteins, are reviewed. These studies have generated a few general rules that can be applied to all members of the three gene families. PREP and MEIS recognize slightly different consensus sequences: PREP prefers to bind to promoters and to have PBX as a DNA-binding partner; MEIS prefers HOX as partner, and both PREP and MEIS drive PBX to their own binding sites. This outlines the clear individuality of the PREP and MEIS proteins, the former mostly devoted to basic cellular functions, the latter more to developmental functions. © 2013 Wiley Periodicals, Inc.


Aceto N.,Friedrich Miescher Institute for Biomedical Research | Sausgruber N.,Friedrich Miescher Institute for Biomedical Research | Brinkhaus H.,Friedrich Miescher Institute for Biomedical Research | Gaidatzis D.,Friedrich Miescher Institute for Biomedical Research | And 12 more authors.
Nature Medicine | Year: 2012

New cancer therapies are likely to arise from an in-depth understanding of the signaling networks influencing tumor initiation, progression and metastasis. We show a fundamental role for Src-homology 2 domain-containing phosphatase 2 (SHP2) in these processes in human epidermal growth factor receptor 2 (HER2)-positive and triple-negative breast cancers. Knockdown of SHP2 eradicated breast tumor-initiating cells in xenograft models, and SHP2 depletion also prevented invasion in three-dimensional cultures and in a transductal invasion assay in vivo. Notably, SHP2 knockdown in established breast tumors blocked their growth and reduced metastasis. Mechanistically, SHP2 activated stemness-associated transcription factors, including v-myc myelocytomatosis viral oncogene homolog (c-Myc) and zinc finger E-box binding homeobox 1 (ZEB1), which resulted in the repression of let-7 microRNA and the expression of a set of 'SHP2 signature' genes. We found these genes to be simultaneously activated in a large subset of human primary breast tumors that are associated with invasive behavior and poor prognosis. These results provide new insights into the signaling cascades influencing tumor-initiating cells as well as a rationale for targeting SHP2 in breast cancer. © 2012 Nature America, Inc. All rights reserved.


Bermejo R.,Fondazione Instituto Firc Of Oncologia Molecolare Ifom | Capra T.,Fondazione Instituto Firc Of Oncologia Molecolare Ifom | Jossen R.,Fondazione Instituto Firc Of Oncologia Molecolare Ifom | Colosio A.,Fondazione Instituto Firc Of Oncologia Molecolare Ifom | And 12 more authors.
Cell | Year: 2011

Transcription hinders replication fork progression and stability, and the Mec1/ATR checkpoint protects fork integrity. Examining checkpoint-dependent mechanisms controlling fork stability, we find that fork reversal and dormant origin firing due to checkpoint defects are rescued in checkpoint mutants lacking THO, TREX-2, or inner-basket nucleoporins. Gene gating tethers transcribed genes to the nuclear periphery and is counteracted by checkpoint kinases through phosphorylation of nucleoporins such as Mlp1. Checkpoint mutants fail to detach transcribed genes from nuclear pores, thus generating topological impediments for incoming forks. Releasing this topological complexity by introducing a double-strand break between a fork and a transcribed unit prevents fork collapse. Mlp1 mutants mimicking constitutive checkpoint-dependent phosphorylation also alleviate checkpoint defects. We propose that the checkpoint assists fork progression and stability at transcribed genes by phosphorylating key nucleoporins and counteracting gene gating, thus neutralizing the topological tension generated at nuclear pore gated genes. © 2011 Elsevier Inc.


Kumar A.,Fondazione Instituto Firc Of Oncologia Molecolare Ifom | Mazzanti M.,University of Milan | Mistrik M.,Palacky University | Kosar M.,Danish Cancer Society | And 13 more authors.
Cell | Year: 2014

ATR controls chromosome integrity and chromatin dynamics. We have previously shown that yeast Mec1/ATR promotes chromatin detachment from the nuclear envelope to counteract aberrant topological transitions during DNA replication. Here, we provide evidence that ATR activity at the nuclear envelope responds to mechanical stress. Human ATR associates with the nuclear envelope during S phase and prophase, and both osmotic stress and mechanical stretching relocalize ATR to nuclear membranes throughout the cell cycle. The ATR-mediated mechanical response occurs within the range of physiological forces, is reversible, and is independent of DNA damage signaling. ATR-defective cells exhibit aberrant chromatin condensation and nuclear envelope breakdown. We propose that mechanical forces derived from chromosome dynamics and torsional stress on nuclear membranes activate ATR to modulate nuclear envelope plasticity and chromatin association to the nuclear envelope, thus enabling cells to cope with the mechanical strain imposed by these molecular processes. © 2014 The Authors.


Shubassi G.,Fondazione Instituto Firc Of Oncologia Molecolare Ifom | Robert T.,Fondazione Instituto Firc Of Oncologia Molecolare Ifom | Robert T.,French National Center for Scientific Research | Vanoli F.,Fondazione Instituto Firc Of Oncologia Molecolare Ifom | And 5 more authors.
Cancer Research | Year: 2012

Histone deacetylase (HDAC) inhibitors are clinically relevant because they are used as anticancer drugs. Recent evidence sheds light on an intriguing connection among the DNA damage response (DDR), protein acetylation, and autophagy. HDAC inhibitors have been shown to counteract key steps in the cellular response to double-strand break formation by affecting checkpoint activation, homologous recombination-mediated repair of DNA lesions, and stability of crucial enzymes involved in resection of DNA ends. The degradation of the resection factors depends on autophagy, which plays a detrimental role when cells are in a hyperacetylated state and experience treatment with radiomimetic anticancer drugs. Future work will be required to further investigate the mechanisms underlying the link between acetylation, autophagy, and the DDR, as well as the significance of mTORC1 inhibitors, which are potent inducers of autophagy that are now used in cancer treatment. ©2012 AACR.


Tordonato C.,Italian National Cancer Institute | Di Fiore P.P.,Italian National Cancer Institute | Di Fiore P.P.,Fondazione Instituto Firc Of Oncologia Molecolare Ifom | Di Fiore P.P.,University of Milan | Nicassio F.,Italian Institute of Technology
Frontiers in Genetics | Year: 2015

The outlook on stem cell biology is shifting from a rigid hierarchical to a more flexible model in which the identity and the behavior of adult stem cells (SCs), far from being fixed, are determined by the dynamic integration of cell autonomous and non-autonomous mechanisms. Within this framework, the recent discovery of thousands of non-coding RNAs (ncRNAs) with regulatory function is redefining the landscape of transcriptome regulation, highlighting the interplay of epigenetic, transcriptional and post-transcriptional mechanisms in the specification of cell fate and in the regulation of developmental processes. Furthermore, the expression of ncRNAs is often tissue- or even cell type-specific, emphasizing their involvement in defining space, time and developmental stages in gene regulation. Such a role of ncRNAs has been investigated in embryonic and induced pluripotent stem cells, and in numerous types of adult SCs and progenitors, including those of the breast, which will be the topic of this review. We will focus on ncRNAs with an important role in breast cancer, in particular in mammary cancer stem cells and progenitors, and highlight the ncRNA-based circuitries whose subversion alters a number of the epigenetic, transcriptional and post-transcriptional events that control "stemness" in the physiological setting. © 2015 Tordonato, Di_fiore and Nicassio.


Callari M.,Fondazione IRCCS Instituto Nazionale dei Tumori | Cappelletti V.,Fondazione IRCCS Instituto Nazionale dei Tumori | De Cecco L.,Fondazione IRCCS Instituto Nazionale dei Tumori | De Cecco L.,Fondazione Instituto Firc Of Oncologia Molecolare Ifom | And 7 more authors.
Breast Cancer Research and Treatment | Year: 2011

Male breast cancer (MBC) is a poorly characterized disease because of its rarity. Clinical management is based on results obtained from randomized trials conducted in women notwithstanding data in the literature suggesting relevant gender-associated differences in terms of biological and clinical behavior. However, a genome-wide characterization of MBC on a transcriptional level is lacking. In this study, gene expression profiles of 37 estrogen receptor positive (ER+) MBC specimens were compared to that of 53 ER+ Female Breast Cancer (FBC) samples similar for clinical and patho-biological features. Almost 1000 genes were found differentially expressed (FDR < 1%) between female and male patients and biological interpretation highlighted a gender-associated modulation of key biological processes ranging from energy metabolism to regulation of translation and matrix remodeling as well as immune system recruitment. Moreover, an analysis of genes correlated to steroid receptors and ERBB2 suggested a prominent role for the androgen receptor in MBC with a minor relevance for progesterone receptor and ERBB2, although, similarly to FBC, a genomic amplification could be observed. Our findings support the idea that breast cancer is a quite different disease in male and female patients and the underlying gender-related biological differences are likely to have clinical implications connected with different susceptibility to treatment. © 2010 Springer Science+Business Media, LLC.


Gonzalez-Huici V.,Fondazione Instituto Firc Of Oncologia Molecolare Ifom | Gonzalez-Huici V.,University of Dundee | Szakal B.,Fondazione Instituto Firc Of Oncologia Molecolare Ifom | Urulangodi M.,Fondazione Instituto Firc Of Oncologia Molecolare Ifom | And 9 more authors.
EMBO Journal | Year: 2014

DNA replication is sensitive to damage in the template. To bypass lesions and complete replication, cells activate recombination mediated (error-free) and translesion synthesis-mediated (errorprone) DNA damage tolerance pathways. Crucial for error-free DNA damage tolerance is template switching, which depends on the formation and resolution of damage-bypass intermediates consisting of sister chromatid junctions. Here we show that a chromatin architectural pathway involving the high mobility group box protein Hmo1 channels replication- associated lesions into the errorfree DNA damage tolerance pathway mediated by Rad5 and PCNA polyubiquitylation, while preventing mutagenic bypass and toxic recombination. In the process of template switching, Hmo1 also promotes sister chromatid junction formation predominantly during replication. Its C-terminal tail, implicated in chromatin bending, facilitates the formation of catenations/hemicatenations and mediates the roles of Hmo1 in DNA damage tolerance pathway choice and sister chromatid junction formation. Together, the results suggest that replication-associated topological changes involving the molecular DNA bender, Hmo1, set the stage for dedicated repair reactions that limit errors during replication and impact on genome stability. © 2014 The Authors.


Valente C.,National Research Council Italy | Valente C.,Telethon Institute of Genetics and Medicine | Turacchio G.,National Research Council Italy | Mariggio S.,National Research Council Italy | And 14 more authors.
Nature Cell Biology | Year: 2012

Large pleiomorphic carriers leave the Golgi complex for the plasma membrane by en bloc extrusion of specialized tubular domains, which then undergo fission. Several components of the underlying molecular machinery have been identified, including those involved in the budding/initiation of tubular carrier precursors (for example, the phosphoinositide kinase PI(4)KIIIβ 2, the GTPase ARF, and FAPP2), and in the fission of these precursors (for example, PKD, CtBP1-S/BARS). However, how these proteins interact to bring about carrier formation is poorly understood. Here, we describe a protein complex that mediates carrier formation and contains budding and fission molecules, as well as other molecules, such as the adaptor protein 14-3-3γ3. Specifically, we show that 14-3-3γ 3 dimers bridge CtBP1-S/BARS with PI(4)KIIIβ 2, and that the resulting complex is stabilized by phosphorylation by PKD and PAK. Disrupting the association of these proteins inhibits the fission of elongating carrier precursors, indicating that this complex couples the carrier budding and fission processes. © 2012 Macmillan Publishers Limited. All rights reserved.


Wollscheid H.-P.,Fondazione Instituto Firc Of Oncologia Molecolare Ifom
Nature Structural and Molecular Biology | Year: 2016

Myosin VI functions in endocytosis and cell motility. Alternative splicing of myosin VI mRNA generates two distinct isoform types, myosin VIshort and myosin VIlong, which differ in the C-terminal region. Their physiological and pathological roles remain unknown. Here we identified an isoform-specific regulatory helix, named the α2-linker, that defines specific conformations and hence determines the target selectivity of human myosin VI. The presence of the α2-linker structurally defines a new clathrin-binding domain that is unique to myosin VIlong and masks the known RRL interaction motif. This finding is relevant to ovarian cancer, in which alternative myosin VI splicing is aberrantly regulated, and exon skipping dictates cell addiction to myosin VIshort in tumor-cell migration. The RRL interactor optineurin contributes to this process by selectively binding myosin VIshort. Thus, the α2-linker acts like a molecular switch that assigns myosin VI to distinct endocytic (myosin VIlong) or migratory (myosin VIshort) functional roles. © 2016 Nature Publishing Group, a division of Macmillan Publishers Limited. All Rights Reserved.

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