U1278-Biologie de l’ARN, Signalisation et Cancer

Publications de l’équipe

Année de publication : 2009

Stefania Millevoi, Adrien Decorsière, Clarisse Loulergue, Jason Iacovoni, Sandra Bernat, Michael Antoniou, Stéphan Vagner (2009 Aug 1)

A physical and functional link between splicing factors promotes pre-mRNA 3′ end processing.

Nucleic acids research : 4672-83 : DOI : 10.1093/nar/gkp470 En savoir plus
Résumé

Polypyrimidine tract-binding protein (PTB) is a splicing regulator that also plays a positive role in pre-mRNA 3′ end processing when bound upstream of the polyadenylation signal (pA signal). Here, we address the mechanism of PTB stimulatory function in mRNA 3′ end formation. We identify PTB as the protein factor whose binding to the human beta-globin (HBB) 3′ UTR is abrogated by a 3′ end processing-inactivating mutation. We show that PTB promotes both in vitro 3′ end cleavage and polyadenylation and recruits directly the splicing factor hnRNP H to G-rich sequences associated with several pA signals. Increased binding of hnRNP H results in stimulation of polyadenylation through a direct interaction with poly(A) polymerase. Therefore, our results provide evidence of a concerted regulation of pA signal recognition by splicing factors bound to auxiliary polyadenylation sequence elements.

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Stephanie Gachet, Jacques Ghysdael (2009 Jun 12)

Calcineurin/NFAT signaling in lymphoid malignancies.

General physiology and biophysics : F47-54 En savoir plus
Résumé

Deregulated calcium signaling is observed at different stages of tumorigenic processes. An important signaling pathway activated in response to calcium involves the protein phosphatase calcineurin and NFAT transcriptional factors. We review here recent data that indicate an important role of the calcineurin/NFAT pathway in lymphoma/leukemogenesis and discuss the potential therapeutic implications of these findings.

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Susannah L Hewitt, Bu Yin, Yanhong Ji, Julie Chaumeil, Katarzyna Marszalek, Jeannette Tenthorey, Giorgia Salvagiotto, Natalie Steinel, Laura B Ramsey, Jacques Ghysdael, Michael A Farrar, Barry P Sleckman, David G Schatz, Meinrad Busslinger, Craig H Bassing, Jane A Skok (2009 Jun 1)

RAG-1 and ATM coordinate monoallelic recombination and nuclear positioning of immunoglobulin loci.

Nature immunology : 655-64 : DOI : 10.1038/ni.1735 En savoir plus
Résumé

Coordinated recombination of homologous antigen receptor loci is thought to be important for allelic exclusion. Here we show that homologous immunoglobulin alleles pair in a stage-specific way that mirrors the recombination patterns of these loci. The frequency of homologous immunoglobulin pairing was much lower in the absence of the RAG-1-RAG-2 recombinase and was restored in Rag1-/- developing B cells with a transgene expressing a RAG-1 active-site mutant that supported DNA binding but not cleavage. The introduction of DNA breaks on one immunoglobulin allele induced ATM-dependent repositioning of the other allele to pericentromeric heterochromatin. ATM activated by the cleaved allele acts in trans on the uncleaved allele to prevent biallelic recombination and chromosome breaks or translocations.

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Année de publication : 2008

Anne Cammas, Stephen M Lewis, Stephan Vagner, Martin Holcik (2008 Dec 1)

Post-transcriptional control of gene expression through subcellular relocalization of mRNA binding proteins.

Biochemical pharmacology : 1395-403 : DOI : 10.1016/j.bcp.2008.05.022 En savoir plus
Résumé

Eukaryotic cells have developed multiple mechanisms to respond to different physiological cues, such as cellular stress, which allow the cells to adapt themselves to their new environment. The regulation of post-transcriptional gene expression is an important component of the cellular stress response and is mediated by mRNA binding proteins (mRBPs). Recently, several studies have shown that regulated subcellular localization of mRBPs upon diverse stimuli (such as cellular stress) provides an additional level of regulation for gene expression.

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Nuno R dos Santos, Maryvonne Williame, Stéphanie Gachet, Françoise Cormier, Anne Janin, Debra Weih, Falk Weih, Jacques Ghysdael (2008 Jul 2)

RelB-dependent stromal cells promote T-cell leukemogenesis.

PloS one : e2555 : DOI : 10.1371/journal.pone.0002555 En savoir plus
Résumé

The Rel/NF-kappaB transcription factors are often activated in solid or hematological malignancies. In most cases, NF-kappaB activation is found in malignant cells and results from activation of the canonical NF-kappaB pathway, leading to RelA and/or c-Rel activation. Recently, NF-kappaB activity in inflammatory cells infiltrating solid tumors has been shown to contribute to solid tumor initiation and progression. Noncanonical NF-kappaB activation, which leads to RelB activation, has also been reported in breast carcinoma, prostate cancer, and lymphoid leukemia.

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Hind Medyouf, Jacques Ghysdael (2008 Feb 1)

The calcineurin/NFAT signaling pathway: a novel therapeutic target in leukemia and solid tumors.

Cell cycle (Georgetown, Tex.) : 297-303 : DOI : 10.4161/cc.7.3.5357 En savoir plus
Résumé

The calcineurin/NFAT signaling pathway is unique to vertebrates and clear genetic evidences show that it plays critical roles in orchestrating the intricate cellular interactions that characterize vertebrate development and morphogenesis. In this setting, the transcriptional regulators of the NFAT family function as molecular integrators of specific calcium signals with other signaling pathways, including MAPkinase, WNT or NOTCH. Deregulation of calcineurin/NFAT signaling and/or abnormal expression of its components have recently been reported in solid tumors of epithelial origin, lymphoma and lymphoid leukemia. Our studies in mouse models of human T-ALL/lymphoma shows that persistent activation of calcineurin/NFAT signaling is pro-oncogenic in vivo and can be efficiently targeted by well-characterized calcineurin inhibitors. We further discuss facts and hypotheses concerning the molecular events that may act upstream and downstream of calcineurin and/or NFAT activation in different type of cancer cells.

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Cyrille Girard, Céline Verheggen, Henry Neel, Anne Cammas, Stephan Vagner, Johann Soret, Edouard Bertrand, Rémy Bordonné (2008 Jan 25)

Characterization of a short isoform of human Tgs1 hypermethylase associating with small nucleolar ribonucleoprotein core proteins and produced by limited proteolytic processing.

The Journal of biological chemistry : 2060-9 : DOI : 10.1074/jbc.M704209200 En savoir plus
Résumé

Tgs1 is the hypermethylase responsible for m(3)G cap formation of U small nuclear RNAs (U snRNAs) and small nucleolar RNAs (snoRNAs). In vertebrates, hypermethylation of snRNAs occurs in the cytoplasm, whereas this process takes place in the nucleus for snoRNAs. Accordingly, the hypermethylase is found in both compartments with a diffuse localization in the cytoplasm and a concentration in Cajal bodies in the nucleoplasm. In this study, we report that the Tgs1 hypermethylase exists as two species, a full-length cytoplasmic isoform and a shorter nuclear isoform of 65-70 kDa. The short isoform exhibits methyltransferase activity and associates with components of box C/D and H/ACA snoRNPs, pointing to a role of this isoform in hypermethylation of snoRNAs. We also show that production of the short Tgs1 isoform is inhibited by MG132, suggesting that it results from proteasomal limited processing of the full-length Tgs1 protein. Together, our results suggest that proteasome maturation constitutes a mechanism regulating Tgs1 function by generating Tgs1 species with different substrate specificities, subcellular localizations, and functions.

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Année de publication : 2007

Anne Cammas, Frédéric Pileur, Sophie Bonnal, Stephen M Lewis, Nicolas Lévêque, Martin Holcik, Stéphan Vagner (2007 Dec 1)

Cytoplasmic relocalization of heterogeneous nuclear ribonucleoprotein A1 controls translation initiation of specific mRNAs.

Molecular biology of the cell : 5048-59 : DOI : 10.1091/mbc.E07-06-0603 En savoir plus
Résumé

Heterogeneous nuclear ribonucleoprotein (hnRNP) A1 is a nucleocytoplasmic shuttling protein that regulates gene expression through its action on mRNA metabolism and translation. The cytoplasmic redistribution of hnRNP A1 is a regulated process during viral infection and cellular stress. Here, we show that hnRNP A1 is an internal ribosome entry site (IRES) trans-acting factor that binds specifically to the 5′ untranslated region of both the human rhinovirus-2 and the human apoptotic peptidase activating factor 1 (apaf-1) mRNAs, thereby regulating their translation. Furthermore, the cytoplasmic redistribution of hnRNP A1 after rhinovirus infection leads to enhanced rhinovirus IRES-mediated translation, whereas the cytoplasmic relocalization of hnRNP A1 after UVC irradiation limits the UVC-triggered translational activation of the apaf-1 IRES. Therefore, this study provides a direct demonstration that IRESs behave as translational enhancer elements regulated by specific trans-acting mRNA binding proteins in given physiological conditions. Our data highlight a new way to regulate protein synthesis in eukaryotes through the subcellular relocalization of a nuclear mRNA-binding protein.

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Hind Medyouf, Hélène Alcalde, Caroline Berthier, Marie Claude Guillemin, Nuno R dos Santos, Anne Janin, Didier Decaudin, Hugues de Thé, Jacques Ghysdael (2007 May 21)

Targeting calcineurin activation as a therapeutic strategy for T-cell acute lymphoblastic leukemia.

Nature medicine : 736-41 : DOI : 10.1038/nm1588 En savoir plus
Résumé

Calcineurin is a calcium-activated serine/threonine phosphatase critical to a number of developmental processes in the cardiovascular, nervous and immune systems. In the T-cell lineage, calcineurin activation is important for pre-T-cell receptor (TCR) signaling, TCR-mediated positive selection of thymocytes into mature T cells, and many aspects of the immune response. The critical role of calcineurin in the immune response is underscored by the fact that calcineurin inhibitors, such as cyclosporin A (CsA) and FK506, are powerful immunosuppressants in wide clinical use. We observed sustained calcineurin activation in human B- and T-cell lymphomas and in all mouse models of lymphoid malignancies analyzed. In intracellular NOTCH1 (ICN1)- and TEL-JAK2-induced T-cell lymphoblastic leukemia, two mouse models relevant to human malignancies, in vivo inhibition of calcineurin activity by CsA or FK506 induced apoptosis of leukemic cells and rapid tumor clearance, and substantially prolonged mouse survival. In contrast, ectopic expression of a constitutively activated mutant of calcineurin favored leukemia progression. Moreover, CsA treatment induced apoptosis in human lymphoma and leukemia cell lines. Thus, calcineurin activation is critical for the maintenance of the leukemic phenotype in vivo, identifying this pathway as a relevant therapeutic target in lymphoid malignancies.

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Nuno R dos Santos, David S Rickman, Aurélien de Reynies, Françoise Cormier, Maryvonne Williame, Camille Blanchard, Marc-Henri Stern, Jacques Ghysdael (2007 May 1)

Pre-TCR expression cooperates with TEL-JAK2 to transform immature thymocytes and induce T-cell leukemia.

Blood : 3972-81 : DOI : 10.1182/blood-2006-09-048801 En savoir plus
Résumé

The TEL-JAK2 gene fusion, which has been identified in human leukemia, encodes a chimeric protein endowed with constitutive tyrosine kinase activity. TEL-JAK2 transgenic expression in the mouse lymphoid lineage results in fatal and rapid T-cell leukemia/lymphoma. In the present report we show that T-cell leukemic cells from EmuSRalpha-TEL-JAK2 transgenic mice present an aberrant CD8(+) differentiation phenotype, as determined by the expression of stage-specific cell surface markers and lineage-specific genes. TEL-JAK2 transforms immature CD4(-)CD8(-) double-negative thymocytes, as demonstrated by the development of T-cell leukemia with full penetrance in a Rag2-deficient genetic background. This disease is similar to the bona fide TEL-JAK2 disease as assessed by phenotypic and gene profiling analyses. Pre-TCR signaling synergizes with TEL-JAK2 to transform immature thymocytes and initiate leukemogenesis as shown by (1) the delayed leukemia onset in Rag2-, CD3epsilon- and pTalpha-deficient mice, (2) the occurrence of recurrent chromosomal alterations in pre-TCR-deficient leukemia, and (3) the correction of delayed leukemia onset in Rag2-deficient TEL-JAK2 mice by an H-Y TCRalphabeta transgene that mimics pre-TCR signaling. Although not affecting leukemia incidence and mouse survival, TCRalphabeta expression was shown to facilitate leukemic cell expansion in secondary lymphoid organs.

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Stephen M Lewis, Anne Veyrier, Nicoleta Hosszu Ungureanu, Sophie Bonnal, Stéphan Vagner, Martin Holcik (2007 Apr 1)

Subcellular relocalization of a trans-acting factor regulates XIAP IRES-dependent translation.

Molecular biology of the cell : 1302-11 : DOI : 10.1091/mbc.E06-06-0515 En savoir plus
Résumé

Translation of the X-linked inhibitor of apoptosis (XIAP) proceeds by internal ribosome entry site (IRES)-mediated initiation, a process that is physiologically important because XIAP expression is essential for cell survival under conditions of compromised cap-dependent translation, such as cellular stress. The regulation of internal initiation requires the interaction of IRES trans-acting factors (ITAFs) with the IRES element. We used RNA-affinity chromatography to identify XIAP ITAFs and isolated the heterogeneous nuclear ribonucleoprotein A1 (hnRNP A1). We find that hnRNP A1 interacts with XIAP IRES RNA both in vitro and in vivo and that hnRNP A1 negatively regulates XIAP IRES activity. Moreover, XIAP IRES-dependent translation is significantly reduced when hnRNP A1 accumulates in the cytoplasm. Osmotic shock, a cellular stress that causes cytoplasmic accumulation of hnRNP A1, also leads to a decrease in XIAP levels that is abrogated by knockdown of hnRNP A1 expression. These results suggest that the subcellular localization of hnRNP A1 is an important determinant of its ability to negatively regulate XIAP IRES activity, suggesting that the subcellular distribution of ITAFs plays a critical role in regulating IRES-dependent translation. Our findings demonstrate that cytoplasmic hnRNP A1 is a negative regulator of XIAP IRES-dependent translation, indicating a novel function for the cytoplasmic form of this protein.

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Année de publication : 2006

Stefania Millevoi, Clarisse Loulergue, Sabine Dettwiler, Sarah Zeïneb Karaa, Walter Keller, Michael Antoniou, Stéphan Vagner (2006 Oct 18)

An interaction between U2AF 65 and CF I(m) links the splicing and 3′ end processing machineries.

The EMBO journal : 4854-64 : DOI : 10.1038/sj.emboj.7601331 En savoir plus
Résumé

The protein factor U2 snRNP Auxiliary Factor (U2AF) 65 is an essential component required for splicing and involved in the coupling of splicing and 3′ end processing of vertebrate pre-mRNAs. Here we have addressed the mechanisms by which U2AF 65 stimulates pre-mRNA 3′ end processing. We identify an arginine/serine-rich region of U2AF 65 that mediates an interaction with an RS-like alternating charge domain of the 59 kDa subunit of the human cleavage factor I (CF I(m)), an essential 3′ processing factor that functions at an early step in the recognition of the 3′ end processing signal. Tethered functional analysis shows that the U2AF 65/CF I(m) 59 interaction stimulates in vitro 3′ end cleavage and polyadenylation. These results therefore uncover a direct role of the U2AF 65/CF I(m) 59 interaction in the functional coordination of splicing and 3′ end processing.

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Irma G Gonzalez-Herrera, Leonel Prado-Lourenco, Frédéric Pileur, Caroline Conte, Aurélie Morin, Florence Cabon, Hervé Prats, Stephan Vagner, Francis Bayard, Sylvie Audigier, Anne-Catherine Prats (2006 Mar 1)

Testosterone regulates FGF-2 expression during testis maturation by an IRES-dependent translational mechanism.

FASEB journal : official publication of the Federation of American Societies for Experimental Biology : 476-8 : DOI : 10.1096/fj.04-3314fje En savoir plus
Résumé

Spermatogenesis is a complex process involving cell proliferation, differentiation, and apoptosis. Fibroblast growth factor 2 (FGF-2) is involved in testicular function, but its role in spermatogenesis has not been fully documented. The control of FGF-2 expression particularly occurs at the translational level, by an internal ribosome entry site (IRES)-dependent mechanism driving the use of alternative initiation codons. To study IRES activity regulation in vivo, we have developed transgenic mice expressing a bicistronic construct coding for two luciferase genes. Here, we show that the FGF-2 IRES is age-dependently activated in mouse testis, whereas EMCV and c-myc IRESs are not. Real-time PCR confirms that this regulation is translational. By using immunohistological techniques, we demonstrate that FGF-2 IRES stimulation occurs in adult, but not in immature, type-A spermatogonias. This is correlated with activation of endogenous FGF-2 expression in spermatogonia; whereas FGF-2 mRNA transcription is known to decrease in adult testis. Interestingly, the FGF-2 IRES activation is triggered by testosterone and is partially inhibited by siRNA directed against the androgen receptor. Two-dimensional analysis of proteins bound to the FGF-2 mRNA 5’UTR after UV cross-linking reveals that testosterone treatment correlates with the binding of several proteins. These data suggest a paracrine loop where IRES-dependent FGF-2 expression, stimulated by Sertoli cells in response to testosterone produced by Leydig cells, would in turn activate Leydig function and testosterone production. In addition, nuclear FGF-2 isoforms could be involved in an intracrine function of FGF-2 in the start of spermatogenesis, mitosis, or meiosis initiation. This report demonstrates that mRNA translation regulation by an IRES-dependent mechanism participates in a physiological process.

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Année de publication : 2005

Sophie Bonnal, Frédéric Pileur, Cécile Orsini, Fabienne Parker, Françoise Pujol, Anne-Catherine Prats, Stéphan Vagner (2005 Feb 11)

Heterogeneous nuclear ribonucleoprotein A1 is a novel internal ribosome entry site trans-acting factor that modulates alternative initiation of translation of the fibroblast growth factor 2 mRNA.

The Journal of biological chemistry : 4144-53 : DOI : 10.1074/jbc.M411492200 En savoir plus
Résumé

Alternative initiation of translation of the human fibroblast growth factor 2 (FGF-2) mRNA at five in-frame CUG or AUG translation initiation codons requires various RNA cis-acting elements, including an internal ribosome entry site (IRES). Here we describe the purification of a trans-acting factor controlling FGF-2 mRNA translation achieved by several biochemical purification approaches. We have identified the heterogeneous nuclear ribonucleoprotein A1 (hnRNP A1) as a factor that binds to the FGF-2 5′-leader RNA and that also complements defective FGF-2 translation in vitro in rabbit reticulocyte lysate. Recombinant hnRNP A1 stimulates in vitro translation at the four IRES-dependent initiation codons but has no effect on the cap-dependent initiation codon. Consistent with a role of hnRNP A1 in the control of alternative initiation of translation, short interfering RNA-mediated knock down of hnRNP A1 specifically inhibits translation at the four IRES-dependent initiation codons. Furthermore, hnRNP A1 binds to the FGF-2 IRES, implicating this interaction in the control of alternative initiation of translation.

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Année de publication : 2004

Christel Boutonnet, Olivier Boijoux, Sandra Bernat, Abdelhakkim Kharrat, Gilles Favre, Jean-Charles Faye, Stéphan Vagner (2004 Jul 1)

Pharmacological-based translational induction of transgene expression in mammalian cells.

EMBO reports : 721-7 : DOI : 10.1038/sj.embor.7400170 En savoir plus
Résumé

In the quest for the development of pharmacological switches that control gene expression, no system has been reported that regulates at the translational level. To permit small-molecule control of transgene translation, we have constructed a farnesyl transferase inhibitor-responsive translation initiation factor. This artificial protein is a three-component chimaera consisting of the ribosome recruitment core of the eIF4G1 eukaryotic translation initiation factor, the RNA-binding domain of the R17 bacteriophage coat protein and the plasma membrane localization CAAX motif of farnesylated H-Ras. This membrane-delocalized translation factor is inactive unless liberated in the cytosol. Farnesyl transferase inhibitor FTI-277 prevents the membrane association of the CAAX motif and thus increases the cytoplasmic levels of the eIF4G fusion protein, which is then capable of inducing translation of the second cistron of a bicistronic messenger RNA containing an R17-binding site in its intercistronic space. Such direct translational control by farnesyl transferase inhibitors provides a system for fast, graded and reversible regulation of transgene expression.

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