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1. Biomarqueurs prédictifs de réponse aux inhibiteurs de points de contrôle immuns [Predictive biomarkers of response to immune checkpoint inhibitors.]

Author

Alexandra Lespagnol, Cédric Ménard, Romain Corre, Karin Tarte, Samuel Le Sourd, Jean Mosser, Marc Pracht, Julien Edeline, Alexandra Frelau, Microenvironment, Cell Differentiation, Immunology and Cancer (MICMAC), Université de Rennes 1 (UR1), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique ), Institut de Génétique et Développement de Rennes (IGDR), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-Centre National de la Recherche Scientifique (CNRS)-Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique ), CHU Pontchaillou [Rennes], CRLCC Eugène Marquis (CRLCC), Nutrition, Métabolismes et Cancer (NuMeCan), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), CRLCC Eugène Marquis ( CRLCC ), Microenvironnement et cancer ( MiCa ), Université de Rennes 1 ( UR1 ), Université de Rennes ( UNIV-RENNES ) -Université de Rennes ( UNIV-RENNES ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique ), Institut de Génétique et Développement de Rennes ( IGDR ), Université de Rennes ( UNIV-RENNES ) -Université de Rennes ( UNIV-RENNES ) -Centre National de la Recherche Scientifique ( CNRS ) -Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique ), Foie, métabolismes et cancer, Université de Rennes (UR)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique ), Université de Rennes (UR)-Centre National de la Recherche Scientifique (CNRS)-Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique ), Institut National de la Recherche Agronomique (INRA)-Université de Rennes (UR)-Institut National de la Santé et de la Recherche Médicale (INSERM), Service d'oncologie médicale, Molecular Engines Laboratories (MEL), Molecular Engines Laboratories, Microenvironnement et cancer (MiCa), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-Centre National de la Recherche Scientifique (CNRS), and Université de Rennes (UR)-Centre National de la Recherche Scientifique (CNRS)

Subjects
0301 basic medicine, Cancer Research, medicine.medical_treatment, Immunothérapie, [SDV.CAN]Life Sciences [q-bio]/Cancer, PD-L1 expression, Metastasis, 03 medical and health sciences, 0302 clinical medicine, Immune system, Biopsy, Medicine, Radiology, Nuclear Medicine and imaging, Neutrophil to lymphocyte ratio, ComputingMilieux_MISCELLANEOUS, Biomarqueurs, Anti-PD1, [SDV.GEN]Life Sciences [q-bio]/Genetics, biology, medicine.diagnostic_test, [ SDV ] Life Sciences [q-bio], business.industry, Hematology, General Medicine, Immunotherapy, [SDV.IMM.IMM]Life Sciences [q-bio]/Immunology/Immunotherapy, medicine.disease, Expression de PD-L1, Primary tumor, 3. Good health, 030104 developmental biology, Oncology, 030220 oncology & carcinogenesis, biology.protein, Cancer research, Biomarker (medicine), Antibody, business, PD-1 blockades, Biomarkers
Abstract

International audience; Les anti-PD-1 s’imposent dernièrement comme un traitement de référence dans de nombreux cancers métastatiques. Ils présentent l’avantage d’une efficacité significative doublée d’un profil de toxicité favorable en monothérapie. Les taux et durées de réponse sont toutefois très variables, ce qui implique, vu leur coût, d’identifier des biomarqueurs prédictifs de réponse. Dans cette revue de la littérature, nous nous intéressons aux biomarqueurs de réponse des immunothérapies anti-PD1 et anti-CTLA-4. L’expression de PD-L1 par les cellules tumorales et du micro-environnement évaluée par immunohistochimie est prédictive de réponse pour certains cancers mais reste une approche mal standardisée avec différents anticorps, différents seuils de positivité et différentes cibles (tumeur ou micro-environnement). Il s’agit aussi d’une expression dynamique au cours du temps et hétérogène donc souvent discordante entre biopsies et pièces opératoires. Les lymphocytes circulants pourraient représenter un biomarqueur prédictif notamment via la mesure du ratio polynucléaires neutrophiles sur lymphocytes (NLR), ratio simple à réaliser en pratique courante. Un taux de néoantigènes élevé serait également associé à une réponse plus importante ce qui expliquerait les taux de réponses particulièrement élevés des tumeurs avec instabilité micro-satellitaire. Des signatures génomiques pourraient également prédire la sensibilité aux anti-PD1. Par ailleurs, la présence de certaines bactéries de la flore intestinale favoriserait la réponse immunitaire antitumorale même si le mécanisme demeure mal compris. Enfin, des signatures d’expression de cytokines, médiatrices de la réponse immune, pourraient constituer des biomarqueurs pertinents. Plusieurs biomarqueurs de réponse potentiels semblent donc intéressants mais aucun n’est encore prospectivement validé à ce jour.

Published
2018

2. Genomic organization and expression of mouse Tpt1 gene☆

Author

Séverine Beaucourt, Dominique Duflaut, Alexandra Lespagnol, Adam Telerman, Giusy Fiucci, Laurent Susini, Pamela Stumptner-Cuvelette, Robert Amson, Organisation Reconstruction Travail - ORT (FRANCE), CHU Pontchaillou [Rennes], Biologie Cellulaire de l'Immunite Antitumorale, Institut National de la Santé et de la Recherche Médicale (INSERM), Molecular Engines Laboratories (MEL), Molecular Engines Laboratories, molecular engines laboratories (MEL), Laboratoire de Biologie et de Pharmacologie Appliquée (LBPA), and École normale supérieure - Cachan (ENS Cachan)-Centre National de la Recherche Scientifique (CNRS)

Subjects
Saccharomyces cerevisiae Proteins, [SDV]Life Sciences [q-bio], Pseudogene, Molecular Sequence Data, Biology, Genome, Chromosomes, Mice, 03 medical and health sciences, Translationally-controlled tumor protein, Biomarkers, Tumor, Genetics, Animals, Direct repeat, Tissue Distribution, RNA, Messenger, Promoter Regions, Genetic, 3' Untranslated Regions, Gene, Repetitive Sequences, Nucleic Acid, 030304 developmental biology, Genomic organization, 0303 health sciences, Base Sequence, Three prime untranslated region, 030302 biochemistry & molecular biology, Tumor Protein, Translationally-Controlled 1, Molecular biology, Phosphotransferases (Alcohol Group Acceptor), genomic DNA, Gene Components, 5' Untranslated Regions, Poly A, Sequence Alignment, Pseudogenes
Abstract

International audience; The translationally controlled tumor protein (TCTP), also known as histamine-releasing factor (HRF), is encoded by a gene (Tpt1) that is highly conserved throughout phylogeny. TCTP is implicated in cell growth, acute allergic response, and apoptosis. In the present study, seven putative Tpt1 genes with different chromosomal localizations were identified in the mouse genome. In six of them, analysis of the 5' and 3' untranslated regions revealed the presence of flanking direct repeats and residual poly(A) tails typical of pseudogenes. Only three of the seven genes can produce a protein of the expected molecular weight. We isolated the genomic DNA of these three genes to analyze their sequence, genomic organization, and in vitro promoter activity. We found that mouse Tpt1 is localized on chromosome 14 with a canonical intron-exon organization, a functional promoter, and only one transcript that is ubiquitously expressed in all tissues.

Published
2003

3. Exosome secretion, including the DNA damage-induced p53-dependent secretory pathway, is severely compromised in TSAP6/Steap3-null mice

Author

Lionel Blanc, Jean-Christophe Marine, Chantal Beekman, G Fiucci, Robert Amson, Alexandra Lespagnol, D Duflaut, Michel Vidal, Adam Telerman, Molecular Engines Laboratories (MEL), Molecular Engines Laboratories, Laboratoire de Biologie et de Pharmacologie Appliquée (LBPA), École normale supérieure - Cachan (ENS Cachan)-Centre National de la Recherche Scientifique (CNRS), Laboratory for Molecular Cancer Biology, VIB-UGent, Dynamique des interactions membranaires normales et pathologiques (DIMNP), and Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM)-Université Montpellier 2 - Sciences et Techniques (UM2)-Université Montpellier 1 (UM1)

Subjects
Reticulocytes, Transferrin receptor, Apoptosis, Cell Cycle Proteins, Biology, Exosomes, Exosome, 03 medical and health sciences, Mice, 0302 clinical medicine, Downregulation and upregulation, Translationally-controlled tumor protein, Receptors, Transferrin, Animals, Secretion, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Molecular Biology, Exosomal secretion, Secretory pathway, 030304 developmental biology, Mice, Knockout, 0303 health sciences, Membrane Proteins, Tumor Protein, Translationally-Controlled 1, Anemia, Cell Differentiation, Cell Biology, Dendritic Cells, Fibroblasts, 3. Good health, Cell biology, Secretory protein, 030220 oncology & carcinogenesis, Tumor Suppressor Protein p53, Oxidoreductases, Spleen, DNA Damage
Abstract

International audience; TSAP6 (tumor suppressor-activated pathway 6), also known as Steap3, is a direct p53 transcriptional target gene. It regulates protein secretion, for example translationally controlled tumor protein (TCTP), which is implicated in tumor reversion. In keeping with the latter, we show herein that TSAP6 is a glycosylated protein present in the trans-Golgi network, endosomal-vesicular compartment and cytoplasmic membrane. To further investigate the physiological function of TSAP6, we have generated TSAP6-deficient mice. These mice exhibit microcytic anemia with abnormal reticulocyte maturation and deficient transferrin receptor downregulation, a process known to be dependent on exosomal secretion. Moreover, we provide direct evidence that exosome production is severely compromised in TSAP6-null cells. Finally, we show that the DNA damage-induced p53-dependent nonclassical exosomal secretory pathway is abrogated in TSAP6-null cells. Given the fact that exosomes are used as cell-free vaccines against cancer and that they could be involved in the biogenesis and spread of human immunodeficiency virus, it is important to understand their regulation. The results presented here provide the first genetic demonstration that exosome formation is a tightly controlled biological process dependent of TSAP6.Cell Death and Differentiation (2008) 15, 1723-1733; doi:10.1038/cdd.2008.104; published online 11 July 2008.

Published
2008

4. TCTP protects from apoptotic cell death by antagonizing bax function

Author

J-C Martinou, Robert Amson, Didier Busso, Jean Cavarelli, Giusy Fiucci, Dominique Duflaut, Alexandra Lespagnol, Jean-Christophe Marine, Stéphanie Besse, Adam Telerman, Chantal Beekman, Dino Moras, Ar Atkinson, Laurent Susini, P Poussin, Molecular Engines Laboratories, Centre Jean Perrin [Clermont-Ferrand] (UNICANCER/CJP), UNICANCER, Molecular Engines Laboratories (MEL), CHU Pontchaillou [Rennes], Institut de génétique et biologie moléculaire et cellulaire (IGBMC), Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Louis Pasteur - Strasbourg I, Laboratoire de Biologie et de Pharmacologie Appliquée (LBPA), École normale supérieure - Cachan (ENS Cachan)-Centre National de la Recherche Scientifique (CNRS), Laboratory for Molecular Cancer Biology, VIB-UGent, Université Louis Pasteur - Strasbourg I-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Department of Cell Biology, Sciences III, and University of Geneva [Switzerland]

Subjects
[SDV]Life Sciences [q-bio], Molecular Sequence Data, Embryonic Development, [SDV.CAN]Life Sciences [q-bio]/Cancer, Apoptosis, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, Mitochondrion, Crystallography, X-Ray, Protein Structure, Secondary, Cell Line, 03 medical and health sciences, Mice, 0302 clinical medicine, Bcl-2-associated X protein, Translationally-controlled tumor protein, Biomarkers, Tumor, Animals, Humans, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Amino Acid Sequence, Inner mitochondrial membrane, Molecular Biology, PI3K/AKT/mTOR pathway, 030304 developmental biology, bcl-2-Associated X Protein, Mice, Knockout, 0303 health sciences, [SDV.GEN]Life Sciences [q-bio]/Genetics, biology, Tumor Protein, Translationally-Controlled 1, Cell Biology, Recombinant Proteins, 3. Good health, Cell biology, Mitochondria, Protein Structure, Tertiary, Elongation factor, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], 030220 oncology & carcinogenesis, Mutation, biology.protein, Translationally controlled tumour protein, Dimerization, Sequence Alignment
Abstract

International audience; Translationally controlled tumor protein (TCTP) is a potential target for cancer therapy. It functions as a growth regulating protein implicated in the TSC1-TSC2 -mTOR pathway or a guanine nucleotide dissociation inhibitor for the elongation factors EF1A and EF1Bbeta. Accumulating evidence indicates that TCTP also functions as an antiapoptotic protein, through a hitherto unknown mechanism. In keeping with this, we show here that loss of tctp expression in mice leads to increased spontaneous apoptosis during embryogenesis and causes lethality between E6.5 and E9.5. To gain further mechanistic insights into this apoptotic function, we solved and refined the crystal structure of human TCTP at 2.0 A resolution. We found a structural similarity between the H2-H3 helices of TCTP and the H5-H6 helices of Bax, which have been previously implicated in regulating the mitochondrial membrane permeability during apoptosis. By site-directed mutagenesis we establish the relevance of the H2-H3 helices in TCTP's antiapoptotic function. Finally, we show that TCTP antagonizes apoptosis by inserting into the mitochondrial membrane and inhibiting Bax dimerization. Together, these data therefore further confirm the antiapoptotic role of TCTP in vivo and provide new mechanistic insights into this key function of TCTP.Cell Death and Differentiation advance online publication, 15 February 2008; doi:10.1038/cdd.2008.18.

Published
2008

5. Etudes moléculaires et fonctionnelles des gènes TSAP6 et TCTP

Author

Duflaut, Dominique, Duflaut, Dominique, molecular engines laboratories (MEL), Molecular Engines Laboratories, Université Paris Sud - Paris XI, and Adam Telerman(adam.telerman@ens-cachan.fr)

Subjects
apoptose, anémie, Steap3, apoptosis, exosomes, cristallographie, TSAP6, transferrin receptor, anemia, TCTP, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], mitochondria, Adam Telerman, Bax, mitochondrie, souris knock-out, Tpt1, crystallography, réversion tumorale, transferrine récepteur, [SDV.BBM.BC] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], knockout mice, tumor reversion
Abstract

The purpose of my PhD was a functional characterization of the TCTP and TSAP6 genes. These genes are implicated in tumor reversion and form a complex at the protein level. The Tpt1 gene encoding for the TCTP protein is the most differentially expressed one between tumor cells and revertants. It is strongly downregulated in different models of tumor reversion. The analysis of the crystal structure of TCTP reveals two helices, H2-H3, which are homologous to the H5-H6 helices of Bax. These H2-H3 helices of TCTP are implicated in the inhibition of Bax dimer formation and hence, the inhibition of apoptosis. We generated TCTP knockout mice that are lethal during early embryonic development. In parallel, we studied TSAP6,that encodes for a 6 transmembrane domains protein and is a direct transcriptional target of p53. We demonstrate that TSAP6 knockout mice have a microcytic anemia. These mice show a delay in the maturation of reticulocytes with a decrease in the secretion of the Transferrin receptor by the exosomes. More generally, our data indicate, in vivo, that TSAP6 controls the secretion of exosomes induced by the activation of p53. Finally, we show that Sertraline and Thioridazine prevent the formation of a protein complex between TSAP6 and TCTP., L'objectif de mon travail de doctorat était une caractérisation fonctionnelle des gènes TCTP et TSAP6. Ces gènes sont impliqués dans la réversion tumorale et forment un complexe protéique. Tpt1, codant pour la protéine TCTP, est le gène le plus sous-exprimé lors de la réversion tumorale. L'analyse du cristal de TCTP humain montre une forte homologie entre ses hélices H2-H3 et les hélices H5-H6 de Bax. Grâce à ses hélices, TCTP inhibe l'apoptose induite par Bax au niveau des mitochondries. En effet, nous démontrons que TCTP, entre autres par le biais de ces hélices, empêche la dimérisation de Bax. Nous avons aussi développé une lignée de souris TCTP knockout qui présentent une létalité embryonnaire précoce.En parallèle, nous avons étudié TSAP6, qui encode pour une protéine à 6 domaines transmembranaires et qui est une cible transcriptionnelle directe de la p53. Nous avons établi une lignée murine TSAP6 knockout présentant une anémie microcytaire avec une splénomégalie. Les réticulocytes issus des souris knockout présentent un retard de maturation et une anomalie de sécrétion du Récepteur à la Transferrine par les exosomes. De manière plus générale, les résultats obtenus montrent, in vivo, que TSAP6 contrôle la sécrétion des exosomes induite par activation de la P53. Nous montrons aussi que la Sertraline et la Thioridazine empêchent la formation du complexe TSAP6-TCTP.

Published
2008

6. Siah-1b is a direct transcriptional target of p53: Identification of the functional p53 responsive element in the siah-1b promoter

Author

Fiucci, Giusy, Beaucourt, Séverine, Duflaut, Dominique, Lespagnol, Alexandra, Stumptner-Cuvelette, Pamela, Géant, Anne, Buchwalter, Gilles, Tuynder, Marcel, Susini, Laurent, Lassalle, Jean-Michel, Wasylyk, Christine, Wasylyk, Bohdan, Oren, Moshe, Amson, Robert, Telerman, Adam, Geant, A., Organisation Reconstruction Travail - ORT (FRANCE), Molecular Engines Laboratories (MEL), Molecular Engines Laboratories, molecular engines laboratories (MEL), CHU Pontchaillou [Rennes], Biologie Cellulaire de l'Immunite Antitumorale, Institut National de la Santé et de la Recherche Médicale (INSERM), Institut de génétique et biologie moléculaire et cellulaire (IGBMC), Université Louis Pasteur - Strasbourg I-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Centre de Recherches sur la Cognition Animale (CRCA), Centre de Biologie Intégrative (CBI), Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS)-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS)-Institut des sciences du cerveau de Toulouse. (ISCT), Université Toulouse - Jean Jaurès (UT2J)-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-CHU Toulouse [Toulouse]-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université Toulouse - Jean Jaurès (UT2J)-CHU Toulouse [Toulouse]-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Centre National de la Recherche Scientifique (CNRS), Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université de Strasbourg (UNISTRA), Department of Molecular Cell Biology [Rehovot], Weizmann Institute of Science [Rehovot, Israël], Laboratoire de Biologie et de Pharmacologie Appliquée (LBPA), and École normale supérieure - Cachan (ENS Cachan)-Centre National de la Recherche Scientifique (CNRS)

Subjects
Transcriptional Activation, [SDV]Life Sciences [q-bio], Ubiquitin-Protein Ligases, Molecular Sequence Data, [SDV.CAN]Life Sciences [q-bio]/Cancer, Biology, In Vitro Techniques, 03 medical and health sciences, Mice, 0302 clinical medicine, Transcription (biology), Cell Line, Tumor, Consensus Sequence, Consensus sequence, Animals, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Luciferase, Northern blot, RNA, Messenger, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], Binding site, Promoter Regions, Genetic, Gene, 030304 developmental biology, 0303 health sciences, Multidisciplinary, Binding Sites, Base Sequence, Nuclear Proteins, DNA, Neoplasm, Biological Sciences, Molecular biology, Chromatin, Introns, 030220 oncology & carcinogenesis, Tumor Suppressor Protein p53, Chromatin immunoprecipitation
Abstract

Siah proteins are E3 ubiquitin ligases. They are homologues of the Drosophila seven in absentia (Sina), a protein required for the R7 photoreceptor development. We have previously found that the expression of human siah-1 and its mouse homologue siah-1b are induced by p53 during apoptosis and tumor reversion. So far, no evidence that the siah-1b gene is a direct transcriptional target of p53 has been provided. In the present study we investigate this issue. Northern blot analysis with a specific probe demonstrates an increase in siah-1b transcription on activation of endogenous and inducible exogenous p53. To explore whether this effect is directly mediated by p53 we analyzed 20 kb of chromosome X DNA, containing the siah-1b locus. A p53-binding site was identified in the siah-1b promoter, located at nucleotides -2155/-2103 relative to the translational start site. This site is composed of two half-sites, conforming to the p53-binding consensus sequence but separated by a nonclassical 33-bp spacer. In luciferase assays, p53 induces a substantial increase in siah-1b promoter activity. Gel shift and DNase-I-footprinting studies, combined with mutational analysis and chromatin immunoprecipitation, indicate that p53 effectively binds the siah-1b promoter in vitro and in vivo . Thus, the siah-1b gene is a direct transcriptional target of p53.

Published
2004

7. Siah-1 binds and regulates the function of Numb

Author

Sylvie Prieur, Marcel Tuynder, Marie-Claude Gendron, Nathalie Amzallag-Elbaz, Moshe Oren, Alain Israël, Laurent Susini, Adam Telerman, Robert Amson, Tamar Juven-Gershon, Brent Passer, Nicolas Privat, Molecular Engines Laboratories (MEL), Molecular Engines Laboratories, Weizmann Institute of Science [Rehovot, Israël], ImagoSeine, Institut Jacques Monod (IJM (UMR_7592)), Université Paris Diderot - Paris 7 (UPD7)-Centre National de la Recherche Scientifique (CNRS)-Université Paris Diderot - Paris 7 (UPD7)-Centre National de la Recherche Scientifique (CNRS), Physicochimie des Macromolecules Biologiques, Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS), The laboratory of M.O. was supported in part by the German–Israel Project Cooperation (DIP) and Yad Abraham Center for Cancer Diagnosis and Therapy., A.T. and R.A. thank D. Cohen and G. Charpak for critical suggestions and constant support. We also thank J. C. Aster (Harvard Medical School) for his helpful advice. We are grateful to P. Fontanges (Institut Fédératif de Recherche, Paris) for confocal analysis. We also thank M. Treier (European Molecular Biology Laboratory, Heidelberg) and P. Benaroch (Institut Curie, Paris) for the HA-tagged polyubiquitin construct., Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS), and Weizmann Institute of Science

Subjects
MESH: Drosophila, MESH: Apoptosis/physiology, [SDV]Life Sciences [q-bio], Regulator, Apoptosis, MESH: Oligonucleotides, Antisense/metabolism, SIAH1, 0302 clinical medicine, Drosophila Proteins, MESH: Animals, Nuclear protein, MESH: Nuclear Proteins/metabolism, 0303 health sciences, Multidisciplinary, Receptors, Notch, MESH: Juvenile Hormones/physiology, Hydrolysis, Nuclear Proteins, MESH: Nuclear Proteins/physiology, Biological Sciences, Cell biology, Juvenile Hormones, 030220 oncology & carcinogenesis, Drosophila, hormones, hormone substitutes, and hormone antagonists, MESH: Hydrolysis, Protein Binding, animal structures, MESH: Drosophila Proteins, Ubiquitin-Protein Ligases, Notch signaling pathway, Down-Regulation, [SDV.CAN]Life Sciences [q-bio]/Cancer, Cell fate determination, Biology, MESH: Two-Hybrid System Techniques, Cell Line, Gene product, 03 medical and health sciences, MESH: Down-Regulation/physiology, Two-Hybrid System Techniques, Animals, Humans, MESH: Protein Binding, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, MESH: Membrane Proteins/physiology, 030304 developmental biology, [SDV.GEN]Life Sciences [q-bio]/Genetics, MESH: Humans, fungi, Membrane Proteins, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, Oligonucleotides, Antisense, MESH: Ubiquitin-Protein Ligases, MESH: Cell Line, MESH: Tumor Suppressor Protein p53/physiology, Cytoplasm, NUMB, Tumor Suppressor Protein p53, MESH: Receptors, Notch
Abstract

The Drosophila Seven in absentia ( Sina ) gene product originally was described as a protein that controls cell fate decisions during eye development. Its mammalian homolog, Siah-1, recently was found to be involved in p53-dependent and -independent pathways of apoptosis and G 1 arrest. We report that Siah-1 interacts directly with and promotes the degradation of the cell fate regulator Numb. Siah-1-mediated Numb degradation leads to redistribution of endogenous cell-surface Notch to the cytoplasm and nucleus and to augmented Notch-regulated transcriptional activity. These data imply that through its ability to target Numb for degradation, Siah-1 can act as a key regulator of Numb-related activities, including Notch signaling.

Published
2001

8. Parallel FISH and Immunohistochemical Studies of ALK Status in 3244 Non–Small-Cell Lung Cancers Reveal Major Discordances

Author

Audrey Gros, Julien Dagher, Laura Mesturoux, Gwendoline Soler, Dominique Dachary, Jean-Philippe Merlio, Leila Dufrenot, Dan Cristian Chiforeanu, Marc-Antoine Belaud-Rotureau, Véronique Catros, Hugues Begueret, Alexandra Lespagnol, Michele Le Calve, Vincent Jauffret, F Dugay, Florian Cabillic, Romain Corre, Service de Cytogénétique et de Biologie Cellulaire, Université de Rennes 1 (UR1), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-Hôpital Pontchaillou-CHU Pontchaillou [Rennes], Institut de Génétique et Développement de Rennes (IGDR), Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique )-Centre National de la Recherche Scientifique (CNRS)-Université de Rennes 1 (UR1), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES), Registre Multicentrique à Vocation Nationale des Mésothéliomes Pleuraux (MESONAT), CHU Caen, Normandie Université (NU)-Tumorothèque de Caen Basse-Normandie (TCBN)-Normandie Université (NU)-Tumorothèque de Caen Basse-Normandie (TCBN), Service d'anatomie et cytologie pathologiques [Rennes] = Anatomy and Cytopathology [Rennes], CHU Pontchaillou [Rennes], Service de pneumologie [Rennes] = Pneumology [Rennes], Laboratoire de Biologie et de Pharmacologie Appliquée (LBPA), École normale supérieure - Cachan (ENS Cachan)-Centre National de la Recherche Scientifique (CNRS), Molecular Engines Laboratories, Université Paris Descartes - Paris 5 (UPD5), INSERM EMI0210 (EMI0210), Institut National de la Santé et de la Recherche Médicale (INSERM), Foie, métabolismes et cancer, Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique ), Laboratoire d'Histologie et de Pathologie moléculaire des tumeurs, Université Bordeaux Segalen - Bordeaux 2-EA 2406, Université de Rennes 1 ( UR1 ), Université de Rennes ( UNIV-RENNES ) -Université de Rennes ( UNIV-RENNES ) -Hôpital Pontchaillou-CHU Pontchaillou [Rennes], Institut de Génétique et Développement de Rennes ( IGDR ), Université de Rennes ( UNIV-RENNES ) -Université de Rennes ( UNIV-RENNES ) -Centre National de la Recherche Scientifique ( CNRS ) -Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique ), Centre national référent cancers rares - mésothéliomes malins pleuraux et tumeurs péritonéales rares ( MESOPATH ), CHU Caen-Hôpital côte de nacre, Service d'anatomie et cytologie pathologiques [Rennes], Service de pneumologie, Hôpital Pontchaillou-CHU Pontchaillou [Rennes], Laboratoire de Biologie et de Pharmacologie Appliquée ( LBPA ), École normale supérieure - Cachan ( ENS Cachan ) -Centre National de la Recherche Scientifique ( CNRS ), Université Paris Descartes - Paris 5 ( UPD5 ), INSERM EMI0210 ( EMI0210 ), Institut National de la Santé et de la Recherche Médicale ( INSERM ), Université de Rennes ( UNIV-RENNES ) -Université de Rennes ( UNIV-RENNES ) -Institut National de la Santé et de la Recherche Médicale ( INSERM ) -Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique ), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-Centre National de la Recherche Scientifique (CNRS)-Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique ), Université de Rennes (UR)-Hôpital Pontchaillou-CHU Pontchaillou [Rennes], Université de Rennes (UR)-Centre National de la Recherche Scientifique (CNRS)-Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique ), and Université de Rennes (UR)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique )

Subjects
Oncology, Male, Pathology, Lung Neoplasms, Cohort Studies, Immunoenzyme Techniques, Anaplastic lymphoma kinase, 0302 clinical medicine, Carcinoma, Non-Small-Cell Lung, hemic and lymphatic diseases, Medicine, In Situ Hybridization, Fluorescence, Aged, 80 and over, Gene Rearrangement, 0303 health sciences, Middle Aged, Prognosis, Immunohistochemistry, 3. Good health, 030220 oncology & carcinogenesis, Carcinoma, Squamous Cell, Biomarker (medicine), Female, medicine.drug, Adult, Pulmonary and Respiratory Medicine, medicine.medical_specialty, medicine.drug_class, Adenocarcinoma, Fluorescent in situ hybridization, 03 medical and health sciences, Carcinoma, Adenosquamous, Young Adult, Internal medicine, Carcinoma, Biomarkers, Tumor, Humans, 030304 developmental biology, Aged, Neoplasm Staging, [SDV.GEN]Life Sciences [q-bio]/Genetics, Crizotinib, business.industry, Non–small-cell lung cancer, Receptor Protein-Tyrosine Kinases, Gene rearrangement, Biomarker, medicine.disease, ALK inhibitor, Clinical trial, business, [ SDV.GEN ] Life Sciences [q-bio]/Genetics, Non-small-cell lung cancer, Follow-Up Studies
Abstract

International audience; Introduction: Anaplastic lymphoma kinase (ALK) rearrangements occur in 1% to 7% of non-small-cell lung cancers (NSCLCs). Crizotinib, an ALK inhibitor, has been demonstrated to provide dramatic clinical benefits in ALK-positive advanced-stage NSCLC. Fluorescent in situ hybridization (FISH) has been established in clinical trials as the standard procedure method for detecting ALK rearrangements. Although the detection of ALK by immunohistochemistry (IHC) has been proposed for the screening of patients, large-scale studies are warranted to validate such a hierarchical approach. Methods: In this article, we report the largest series thus far of parallel FISH and IHC ALK testing in 3244 consecutive NSCLC cases analyzed at two independent French centers. Results: FISH-positive and/or IHC-positive results were demonstrated in 150 of 3244 cases (4.6%). An imbalanced sex ratio was detected, with women exhibiting a 2.2-fold relative risk for an alteration. Strikingly, only 80 of 150 specimens were classified as ALK positive by both techniques. The specimens with discordant FISH/IHC analyses were FISH-positive/IHC-negative (36), FISH-negative/IHC-positive (19), or FISH-noncontributive/IHC-positive (15). Thus, a single FISH or IHC analysis performed alone would have failed to detect approximately one-fourth of the ALK-positive cases with similar findings in our two centers. Conclusions: This study highlights the feasibility of systematic NSCLC testing by both FISH and IHC in routine practice. Many preanalytical factors may account for the apparent discrepancies between both methods, suggesting that hierarchical screening may underscore ALK-positive cases. This significant level of discrepancy supports the need of combined testing to optimize the detection of ALK-inhibitor-eligible patients given that some patients with discordant testing were found to respond to crizotinib.

Published
2014
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9. TCTP protects from apoptotic cell death by antagonizing bax function.

Author

Susini L, Besse S, Duflaut D, Lespagnol A, Beekman C, Fiucci G, Atkinson AR, Busso D, Poussin P, Marine JC, Martinou JC, Cavarelli J, Moras D, Amson R, and Telerman A

Subjects
Amino Acid Sequence, Animals, Biomarkers, Tumor genetics, Cell Line, Crystallography, X-Ray, Dimerization, Embryonic Development, Humans, Mice, Mice, Knockout, Molecular Sequence Data, Mutation, Protein Structure, Secondary, Protein Structure, Tertiary, Recombinant Proteins chemistry, Recombinant Proteins metabolism, Sequence Alignment, Tumor Protein, Translationally-Controlled 1, bcl-2-Associated X Protein chemistry, Apoptosis, Biomarkers, Tumor chemistry, Biomarkers, Tumor metabolism, Mitochondria metabolism, bcl-2-Associated X Protein metabolism
Abstract

Translationally controlled tumor protein (TCTP) is a potential target for cancer therapy. It functions as a growth regulating protein implicated in the TSC1-TSC2 -mTOR pathway or a guanine nucleotide dissociation inhibitor for the elongation factors EF1A and EF1Bbeta. Accumulating evidence indicates that TCTP also functions as an antiapoptotic protein, through a hitherto unknown mechanism. In keeping with this, we show here that loss of tctp expression in mice leads to increased spontaneous apoptosis during embryogenesis and causes lethality between E6.5 and E9.5. To gain further mechanistic insights into this apoptotic function, we solved and refined the crystal structure of human TCTP at 2.0 A resolution. We found a structural similarity between the H2-H3 helices of TCTP and the H5-H6 helices of Bax, which have been previously implicated in regulating the mitochondrial membrane permeability during apoptosis. By site-directed mutagenesis we establish the relevance of the H2-H3 helices in TCTP's antiapoptotic function. Finally, we show that TCTP antagonizes apoptosis by inserting into the mitochondrial membrane and inhibiting Bax dimerization. Together, these data therefore further confirm the antiapoptotic role of TCTP in vivo and provide new mechanistic insights into this key function of TCTP.

Published
2008
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10. TSAP6 facilitates the secretion of translationally controlled tumor protein/histamine-releasing factor via a nonclassical pathway.

Author

Amzallag N, Passer BJ, Allanic D, Segura E, Théry C, Goud B, Amson R, and Telerman A

Subjects
Biomarkers, Tumor, Cell Cycle Proteins, Cell Line, Cycloheximide pharmacology, Humans, Neoplasm Proteins analysis, Nuclear Proteins analysis, Oncogene Proteins analysis, Oxidoreductases, Protein Transport, Secretory Vesicles chemistry, Tumor Protein, Translationally-Controlled 1, Neoplasm Proteins metabolism, Nuclear Proteins metabolism, Oncogene Proteins physiology
Abstract

Translationally controlled tumor protein (TCTP) is cytoplasmic and structurally related to guanine-nucleotide free chaperones. TCTP (also called histamine-releasing factor) has been described previously as a secreted protein that participates in inflammatory responses by promoting the release of histamine. How TCTP is eventually exported out of the cell to promote such activities is unknown. Here we show that TCTP secretion was insensitive to either brefeldin A or monensin, suggesting that it proceeds via an endoplasmic reticulum/Golgi-independent or nonclassical pathway. Moreover, our analyses also suggest that secreted TCTP originates from pre-existing pools. TSAP6, a p53-inducible 5-6 transmembrane protein, was found to interact with TCTP in a yeast two-hybrid hunt. GST pull down assays confirmed their direct interaction, and immunofluorescence analysis revealed their partial co-distribution to vesicular-like structures at the plasma membrane and around the nucleus. Functionally, the overexpression of TSAP6 consistently leads to enhanced secretion of both endogenously and exogenously expressed TCTP. Finally, we found TCTP in preparations of small secreted vesicles called exosomes, which have been suggested as a possible pathway for nonclassical secretion. Overexpression of TSAP6 also increased TCTP levels in exosome preparations. Altogether, these data identify a novel role for TSAP6 in the export of TCTP and indicate that this multipass membrane protein could have a general role in the regulation of vesicular trafficking and secretion.

Published
2004
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11. Translationally controlled tumor protein is a target of tumor reversion.

Author

Tuynder M, Fiucci G, Prieur S, Lespagnol A, Géant A, Beaucourt S, Duflaut D, Besse S, Susini L, Cavarelli J, Moras D, Amson R, and Telerman A

Subjects
Animals, Base Sequence, Cell Line, Transformed, Cell Line, Tumor, DNA Primers, Humans, Mice, NIH 3T3 Cells, Phenotype, Tumor Protein, Translationally-Controlled 1, Neoplasm Proteins genetics, Neoplasms pathology, Protein Biosynthesis
Abstract

By analyzing the gene expression profile between tumor cells and revertant counterparts that have a suppressed malignant phenotype, we previously reported a significant down-regulation of translationally controlled tumor protein (TCTP) in the revertants. In the present study, we derived, by using the H1 parvovirus as a selective agent, revertants from three major solid cancers: colon, lung, and melanoma cell lines. These cells have a strongly suppressed malignant phenotype both in vitro and in vivo. The level of TCTP is decreased in most of the revertants. To verify whether inhibition of TCTP expression induces changes in the malignant phenotype, in the classical, well established model of "flat reversion," v-src-transformed NIH3T3 cells were transfected with antisense TCTP. By inhibiting the expression of TCTP, the number of revertant cells was raised to 30%, instead of the reported rate for spontaneous flat revertants of 10(-6). Because TCTP encodes for a histamine-releasing factor, we tested the hypothesis that inhibitors of the histaminic pathway could be effective against tumor cells. We show that some antihistaminic compounds (hydroxyzine and promethazine) and other pharmacological compounds with a related structure (including thioridazine and sertraline) kill tumor cells and significantly decrease the level of TCTP. All together, these data suggest that, with tumor reversion used as a working model, TCTP was identified as a target and drugs were selected that decrease its expression and kill tumor cells.

Published
2004
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12. Siah-1b is a direct transcriptional target of p53: identification of the functional p53 responsive element in the siah-1b promoter.

Author

Fiucci G, Beaucourt S, Duflaut D, Lespagnol A, Stumptner-Cuvelette P, Géant A, Buchwalter G, Tuynder M, Susini L, Lassalle JM, Wasylyk C, Wasylyk B, Oren M, Amson R, and Telerman A

Subjects
Animals, Base Sequence, Binding Sites genetics, Cell Line, Tumor, Chromatin genetics, Chromatin metabolism, Consensus Sequence, DNA, Neoplasm genetics, DNA, Neoplasm metabolism, In Vitro Techniques, Introns, Mice, Molecular Sequence Data, RNA, Messenger biosynthesis, RNA, Messenger genetics, Transcriptional Activation, Ubiquitin-Protein Ligases, Nuclear Proteins genetics, Promoter Regions, Genetic, Tumor Suppressor Protein p53 metabolism
Abstract

Siah proteins are E3 ubiquitin ligases. They are homologues of the Drosophila seven in absentia (Sina), a protein required for the R7 photoreceptor development. We have previously found that the expression of human siah-1 and its mouse homologue siah-1b are induced by p53 during apoptosis and tumor reversion. So far, no evidence that the siah-1b gene is a direct transcriptional target of p53 has been provided. In the present study we investigate this issue. Northern blot analysis with a specific probe demonstrates an increase in siah-1b transcription on activation of endogenous and inducible exogenous p53. To explore whether this effect is directly mediated by p53 we analyzed 20 kb of chromosome X DNA, containing the siah-1b locus. A p53-binding site was identified in the siah-1b promoter, located at nucleotides -2155/-2103 relative to the translational start site. This site is composed of two half-sites, conforming to the p53-binding consensus sequence but separated by a nonclassical 33-bp spacer. In luciferase assays, p53 induces a substantial increase in siah-1b promoter activity. Gel shift and DNase-I-footprinting studies, combined with mutational analysis and chromatin immunoprecipitation, indicate that p53 effectively binds the siah-1b promoter in vitro and in vivo. Thus, the siah-1b gene is a direct transcriptional target of p53.

Published
2004
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13. Translationally controlled tumor protein acts as a guanine nucleotide dissociation inhibitor on the translation elongation factor eEF1A.

Author

Cans C, Passer BJ, Shalak V, Nancy-Portebois V, Crible V, Amzallag N, Allanic D, Tufino R, Argentini M, Moras D, Fiucci G, Goud B, Mirande M, Amson R, and Telerman A

Subjects
Biomarkers, Tumor genetics, Drug Stability, Guanine Nucleotide Dissociation Inhibitors genetics, Guanosine Diphosphate metabolism, Humans, In Vitro Techniques, Kinetics, Peptide Elongation Factor 1 genetics, Protein Biosynthesis, Recombinant Fusion Proteins genetics, Recombinant Fusion Proteins metabolism, Tumor Protein, Translationally-Controlled 1, Two-Hybrid System Techniques, Biomarkers, Tumor metabolism, Guanine Nucleotide Dissociation Inhibitors metabolism, Guanine Nucleotides metabolism, Peptide Elongation Factor 1 metabolism
Abstract

Recently, we demonstrated that the expression levels of the translationally controlled tumor protein (TCTP) were strongly down-regulated at the mRNA and protein levels during tumor reversion/suppression and by the activation of p53 and Siah-1. To better characterize the function of TCTP, a yeast two-hybrid hunt was performed. Subsequent analysis identified the translation elongation factor, eEF1A, and its guanine nucleotide exchange factor, eEF1Bbeta, as TCTP-interacting partners. In vitro and in vivo studies confirmed that TCTP bound specifically eEF1Bbeta and eEF1A. Additionally, MS analysis also identified eEF1A as a TCTP interactor. Because eEF1A is a GTPase, we investigated the role of TCTP on the nucleotide exchange reaction of eEF1A. Our results show that TCTP preferentially stabilized the GDP form of eEF1A, and, furthermore, impaired the GDP exchange reaction promoted by eEF1Bbeta. These data suggest that TCTP has guanine nucleotide dissociation inhibitor activity, and, moreover, implicate TCTP in the elongation step of protein synthesis.

Published
2003
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14. Genomic organization and expression of mouse Tpt1 gene.

Author

Fiucci G, Lespagnol A, Stumptner-Cuvelette P, Beaucourt S, Duflaut D, Susini L, Amson R, and Telerman A

Subjects
3' Untranslated Regions, 5' Untranslated Regions, Animals, Base Sequence, Chromosomes, Molecular Sequence Data, Phosphotransferases (Alcohol Group Acceptor) metabolism, Poly A, Promoter Regions, Genetic, Pseudogenes, RNA, Messenger, Repetitive Sequences, Nucleic Acid, Saccharomyces cerevisiae Proteins metabolism, Sequence Alignment, Tissue Distribution, Tumor Protein, Translationally-Controlled 1, Biomarkers, Tumor genetics, Gene Components, Genome, Mice genetics, Phosphotransferases (Alcohol Group Acceptor) genetics, Saccharomyces cerevisiae Proteins genetics
Abstract

The translationally controlled tumor protein (TCTP), also known as histamine-releasing factor (HRF), is encoded by a gene (Tpt1) that is highly conserved throughout phylogeny. TCTP is implicated in cell growth, acute allergic response, and apoptosis. In the present study, seven putative Tpt1 genes with different chromosomal localizations were identified in the mouse genome. In six of them, analysis of the 5' and 3' untranslated regions revealed the presence of flanking direct repeats and residual poly(A) tails typical of pseudogenes. Only three of the seven genes can produce a protein of the expected molecular weight. We isolated the genomic DNA of these three genes to analyze their sequence, genomic organization, and in vitro promoter activity. We found that mouse Tpt1 is localized on chromosome 14 with a canonical intron-exon organization, a functional promoter, and only one transcript that is ubiquitously expressed in all tissues.

Published
2003
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15. The p53-inducible TSAP6 gene product regulates apoptosis and the cell cycle and interacts with Nix and the Myt1 kinase.

Author

Passer BJ, Nancy-Portebois V, Amzallag N, Prieur S, Cans C, Roborel de Climens A, Fiucci G, Bouvard V, Tuynder M, Susini L, Morchoisne S, Crible V, Lespagnol A, Dausset J, Oren M, Amson R, and Telerman A

Subjects
Amino Acid Sequence, Animals, Blotting, Northern, Blotting, Western, Cell Cycle, Cell Cycle Proteins, Cloning, Molecular, DNA, Complementary metabolism, Flow Cytometry, G1 Phase, G2 Phase, Glutathione Transferase metabolism, HeLa Cells, Humans, Luciferases metabolism, Mice, Mitosis, Molecular Sequence Data, Oligonucleotides, Antisense pharmacology, Oxidoreductases, Phosphorylation, Precipitin Tests, Promoter Regions, Genetic, Protein Binding, Protein Biosynthesis, Protein Structure, Tertiary, Proto-Oncogene Proteins c-bcl-2 metabolism, RNA, Messenger metabolism, RNA, Small Interfering metabolism, Recombinant Fusion Proteins metabolism, S Phase, Sequence Homology, Amino Acid, Time Factors, Transcription, Genetic, Tumor Cells, Cultured, Two-Hybrid System Techniques, Apoptosis, Membrane Proteins genetics, Membrane Proteins metabolism, Membrane Proteins physiology, Oncogene Proteins, Protein Serine-Threonine Kinases metabolism, Protein-Tyrosine Kinases metabolism, Proto-Oncogene Proteins, Tumor Suppressor Protein p53 metabolism, Tumor Suppressor Proteins
Abstract

The p53 tumor suppressor protein plays a crucial role in tumorigenesis by controlling cell-cycle progression and apoptosis. We have previously described a transcript designated tumor suppressor activated pathway-6 (TSAP6) that is up-regulated in the p53-inducible cell line, LTR6. Cloning of the murine and human full-length TSAP6 cDNA revealed that it encodes a 488-aa protein with five to six transmembrane domains. This gene is the murine and human homologue of the recently published rat pHyde. Antibodies raised against murine and human TSAP6 recognize a 50- to 55-kDa band induced by p53. Analysis of the TSAP6 promoter identified a functional p53-responsive element. Functional studies demonstrated that TSAP6 antisense cDNA diminished levels of the 50- to 55-kDa protein and decreased significantly the levels of p53-induced apoptosis. Furthermore, TSAP6 small interfering RNA inhibited apoptosis in TSAP6-overexpressing cells. Yeast two-hybrid analysis followed by GST/in vitro-transcribed/translated pull-down assays and in vivo coimmunoprecipitations revealed that TSAP6 associated with Nix, a proapoptotic Bcl-2-related protein and the Myt1 kinase, a negative regulator of the G(2)/M transition. Moreover, TSAP6 enhanced the susceptibility of cells to apoptosis and cooperated with Nix to exacerbate this effect. Cell-cycle studies indicated that TSAP6 could augment Myt1 activity. Overall, these data suggest that TSAP6 may act downstream to p53 to interface apoptosis and cell-cycle progression.

Published
2003
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16. Biological models and genes of tumor reversion: cellular reprogramming through tpt1/TCTP and SIAH-1.

Author

Tuynder M, Susini L, Prieur S, Besse S, Fiucci G, Amson R, and Telerman A

Subjects
Basement Membrane pathology, Breast Neoplasms, Female, Humans, K562 Cells, Models, Genetic, Recombinant Proteins metabolism, Transfection, Tumor Cells, Cultured, Tumor Protein, Translationally-Controlled 1, U937 Cells, Ubiquitin-Protein Ligases, Biomarkers, Tumor genetics, DNA, Antisense genetics, DNA, Neoplasm genetics, Gene Expression Regulation, Neoplastic, Neoplasms genetics, Nuclear Proteins genetics, RNA, Small Interfering genetics
Abstract

Tumor reversion is the process by which some cancer cells lose their malignant phenotype. This study was aimed at defining some of the molecular and phenotypic properties of this process. Biological models of tumor reversion were isolated from human leukemia and breast cancer cell lines by using the H-1 parvovirus as a selective agent. Differential gene expression analysis was performed between the parental malignant cells and their revertants or alternatively between these parental cells and their SIAH-1 transfectant counterparts. These SIAH-1 transfectants have a suppressed malignant phenotype and were used as a control for a viral-free system. Two hundred sixty-three genes were found to be either activated or inhibited during the reversion process, as confirmed by Northern blot analysis or quantitative PCR. Of these, 32% were differentially expressed in all systems, irrespective of whether parvovirus-selected, SIAH-1 overexpressing, or p53 mutant or wild-type cell lines were used, suggesting the existence of a universal mechanism underlying tumor reversion. Translationally Controlled Tumor Protein (tpt1/TCTP) has the strongest differential expression, down-regulated in the reversion of U937- and SIAH-1-overexpressing cells. Inhibition of TCTP expression by anti-sense cDNA or small interfering RNA molecules results in suppression of the malignant phenotype and in cellular reorganization, similar to the effect of SIAH-1. Hence, tumor reversion can be defined at the molecular level, not just as the reversal of malignant transformation, but as a biological process in its own right involving a cellular reprogramming mechanism, overriding genetic changes in cancer, by triggering an alternative pathway leading to suppression of tumorigenicity.

Published
2002
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17. Siah-1 binds and regulates the function of Numb.

Author

Susini L, Passer BJ, Amzallag-Elbaz N, Juven-Gershon T, Prieur S, Privat N, Tuynder M, Gendron MC, Israël A, Amson R, Oren M, and Telerman A

Subjects
Animals, Apoptosis physiology, Cell Line, Down-Regulation physiology, Drosophila, Drosophila Proteins, Humans, Hydrolysis, Membrane Proteins physiology, Nuclear Proteins physiology, Oligonucleotides, Antisense metabolism, Protein Binding, Receptors, Notch, Tumor Suppressor Protein p53 physiology, Two-Hybrid System Techniques, Ubiquitin-Protein Ligases, Juvenile Hormones physiology, Nuclear Proteins metabolism
Abstract

The Drosophila Seven in absentia (Sina) gene product originally was described as a protein that controls cell fate decisions during eye development. Its mammalian homolog, Siah-1, recently was found to be involved in p53-dependent and -independent pathways of apoptosis and G(1) arrest. We report that Siah-1 interacts directly with and promotes the degradation of the cell fate regulator Numb. Siah-1-mediated Numb degradation leads to redistribution of endogenous cell-surface Notch to the cytoplasm and nucleus and to augmented Notch-regulated transcriptional activity. These data imply that through its ability to target Numb for degradation, Siah-1 can act as a key regulator of Numb-related activities, including Notch signaling.

Published
2001
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