Eif4e and Interactors from Unicellular Eukaryotes
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Cancer Cells Exploit Eif4e2-Directed Translation to Enhance Their Proliferation, Migration and Invasion
Cancer cells exploit eIF4E2-directed translation to enhance their proliferation, migration and invasion by Joseph F. Varga A Thesis presented to The University of Guelph In partial fulfilment of requirements for the degree of Master of Science in Molecular and Cellular Biology Guelph, Ontario, Canada © Joseph F. Varga, August, 2016 ABSTRACT Cancer cells exploit eIF4E2-directed translation to enhance their proliferation, migration and invasion Joseph F. Varga Advisor: University of Guelph, 2016 Professor James Uniacke Despite the diversity found in the genetic makeup of cancer, many cancers share the same tumor microenvironment. Hypoxia, an aspect of the tumor microenvironment, causes the suppression of the primary translational machinery. Hypoxic cells switch from using the eukaryotic initiation factor 4E (eIF4E) to using a homologue of eIF4E (eIF4E2), in order to initiate the translation of select mRNAs. This thesis investigates the role of eIF4E2-directed translation in a panel of cancer cell lines during autonomous proliferation, migration and invasion. In this thesis, we show that silencing eIF4E2 abrogates the autonomous proliferation of colon carcinoma. Silencing eIF4E2 in glioblastoma cells resulted in decreased migration and invasion. Furthermore, we link eIF4E2-directed translation of cadherin 22 with the hypoxic migration of glioblastoma. These findings answer questions regarding the biology of cancer and expand the current knowledge of genes exploited during tumor progression. This data also highlights eIF4E2 as a potential therapeutic target. Acknowledgements I would like to express my sincere gratitude to Dr. James Uniacke for taking me on as a graduate student in his laboratory and providing me with the opportunity to contribute to scientific research. -
Extensive Translation of Circular Rnas Driven by N6-Methyladenosine
Cell Research (2017) 27:626-641. ORIGINAL ARTICLE www.nature.com/cr Extensive translation of circular RNAs driven by N6-methyladenosine Yun Yang1, 2, 3, 4, *, Xiaojuan Fan2, *, Miaowei Mao4, 5, *, Xiaowei Song2, 4, Ping Wu6, 7, Yang Zhang8, Yongfeng Jin1, Yi Yang5, Ling-Ling Chen8, Yang Wang9, Catherine CL Wong6, 7, Xinshu Xiao3, Zefeng Wang2, 4 1Institute of Biochemistry, College of Life Sciences, Zhejiang University at Zijingang, Zhejiang, Hangzhou, Zhejiang 310058, Chi- na; 2CAS Key Lab for Computational Biology, CAS Center for Excellence in Molecular Cell Science, CAS-MPG Partner Institute for Computational Biology, Shanghai Institute for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031, China; 3Department of Integrative Biology and Physiology and the Molecular Biology Institute, UCLA, Los Angeles, CA 90095, USA; 4Department of Pharmacology, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA; 5Synthetic Biology and Biotechnology Laboratory, State Key Laboratory of Bioreactor Engineering, School of Pharmacy, East China University of Sci- ence and Technology, Shanghai, China; 6National Center for Protein Science, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031, China; 7Shanghai Science Research Center, Chinese Academy of Sciences, Shanghai 201204, China; 8Institute of Biochemistry and Cell Biology, Shanghai Institute for Biolog- ical Sciences, Chinese Academy of Sciences, Shanghai 200031, China; 9Institute of Cancer Stem Cell, Dalian Medical University, Dalian, Liaoning 116044, China Extensive pre-mRNA back-splicing generates numerous circular RNAs (circRNAs) in human transcriptome. However, the biological functions of these circRNAs remain largely unclear. Here we report that N6-methyladenosine (m6A), the most abundant base modification of RNA, promotes efficient initiation of protein translation from cir- cRNAs in human cells. -
Composition of Herpesvirus Ribonucleoprotein Complexes †
Abstract Composition of Herpesvirus Ribonucleoprotein Complexes † Eric S. Pringle 1,2,* and Craig McCormick 1,2 1 Department of Microbiology and Immunology, Dalhousie University, Halifax, NS B3H 4R2, Canada; [email protected] 2 Beatrice Hunter Cancer Research Institute, Halifax, NS B3H 4R2, Canada * Correspondence: [email protected] † Presented at Viruses 2020—Novel Concepts in Virology, Barcelona, Spain, 5–7 February 2020. Published: 4 July 2020 Abstract: Herpesvirus genomes are decoded by host RNA polymerase enzymes, generating messenger ribonucleotides (mRNA) that are post-transcriptionally modified and exported to the cytoplasm through the combined work of host and viral factors. These viral mRNA bear 5′-m7GTP caps and poly(A) tails that should permit the assembly of canonical host eIF4F cap-binding complexes to initiate protein synthesis. However, the precise mechanisms of translation initiation remain to be investigated for Kaposi’s sarcoma-associated herpesvirus (KSHV) and other herpesviruses. During KSHV lytic replication in lymphoid cells, the activation of caspases leads to the cleavage of eIF4G and depletion of eIF4F. Translating mRNPs depleted of eIF4F retain viral mRNA, suggesting that non-eIF4F translation initiation is sufficient to support viral protein synthesis. To identify proteins required to support viral protein synthesis, we isolated and characterized actively translating messenger ribonucleoprotein (mRNP) complexes by ultracentrifugation and sucrose-gradient fractionation followed by quantitative mass spectrometry. The abundance of host translation initiation factors available to initiate viral protein synthesis were comparable between cells undergoing KSHV lytic or latent replication. The translation initiation factors eIF4E2, NCBP1, eIF4G2, and eIF3d were detected in association with actively translating mRNP complexes during KSHV lytic replication, but their depletion by RNA silencing did not affect virion production. -
EIF4G2 Rabbit Pab
Leader in Biomolecular Solutions for Life Science EIF4G2 Rabbit pAb Catalog No.: A2897 Basic Information Background Catalog No. Translation initiation is mediated by specific recognition of the cap structure by A2897 eukaryotic translation initiation factor 4F (eIF4F), which is a cap binding protein complex that consists of three subunits: eIF4A, eIF4E and eIF4G. The protein encoded by this gene Observed MW shares similarity with the C-terminal region of eIF4G that contains the binding sites for 102kDa eIF4A and eIF3; eIF4G, in addition, contains a binding site for eIF4E at the N-terminus. Unlike eIF4G, which supports cap-dependent and independent translation, this gene Calculated MW product functions as a general repressor of translation by forming translationally 98kDa/102kDa inactive complexes. In vitro and in vivo studies indicate that translation of this mRNA initiates exclusively at a non-AUG (GUG) codon. Alternatively spliced transcript variants Category encoding different isoforms of this gene have been described. Primary antibody Applications WB, IHC, IF, IP Cross-Reactivity Human, Mouse, Rat Recommended Dilutions Immunogen Information WB 1:500 - 1:1000 Gene ID Swiss Prot 1982 P78344 IHC 1:100 - 1:200 Immunogen 1:50 - 1:200 IF A synthetic peptide corresponding to a sequence within amino acids 750-850 of human EIF4G2 (NP_001409.3). IP 1:50 - 1:200 Synonyms EIF4G2;AAG1;DAP5;NAT1;P97 Contact Product Information www.abclonal.com Source Isotype Purification Rabbit IgG Affinity purification Storage Store at -20℃. Avoid freeze / thaw cycles. Buffer: PBS with 0.02% sodium azide,50% glycerol,pH7.3. Validation Data Western blot analysis of extracts of various cell lines, using EIF4G2 antibody (A2897) at 1:400 dilution. -
Robust Heat Shock Induces Eif2α-Phosphorylation
2078 Research Article Robust heat shock induces eIF2α-phosphorylation- independent assembly of stress granules containing eIF3 and 40S ribosomal subunits in budding yeast, Saccharomyces cerevisiae Tomás Grousl1, Pavel Ivanov1,*, Ivana Frydlová1, Pavla Vasicová1, Filip Janda1, Jana Vojtová1, Katerina Malínská1, Ivana Malcová1, Lenka Nováková1, Dana Janosková1, Leos Valásek2 and Jirí Hasek1,‡ 1Laboratory of Cell Reproduction, Institute of Microbiology of the AS CR, v.v.i., Prague, Czech Republic 2Laboratory of Regulation of Gene Expression, Institute of Microbiology of the AS CR, v.v.i., Prague, Czech Republic *Present address: A.N. Belozersky Institute of Physico-Chemical Biology MSU, Moscow, Russia ‡Author for correspondence (e-mail: [email protected]) Accepted 18 March 2009 Journal of Cell Science 122, 2078-2088 Published by The Company of Biologists 2009 doi:10.1242/jcs.045104 Summary Environmental stresses inducing translation arrest are markers also colocalized with eIF3a. Microscopic analyses of accompanied by the deposition of translational components into the edc3Δlsm4ΔC mutant demonstrated that different stress granules (SGs) serving as mRNA triage sites. It has scaffolding proteins are required to induce SGs upon robust recently been reported that, in Saccharomyces cerevisiae, heat shock as opposed to glucose deprivation. Even though formation of SGs occurs as a result of a prolonged glucose eIF2α became phosphorylated under these stress conditions, the starvation. However, these SGs did not contain eIF3, one of decrease in polysomes and formation of SGs occurred hallmarks of mammalian SGs. We have analyzed the effect of independently of phosphorylation of eIF2α. We conclude that robust heat shock on distribution of eIF3a/Tif32p/Rpg1p and under specific stress conditions, such as robust heat shock, yeast showed that it results in the formation of eIF3a accumulations SGs do contain eIF3 and 40S ribosomes and utilize alternative containing other eIF3 subunits, known yeast SG components routes for their assembly. -
Early Alterations of RNA Metabolism and Splicing from Adult Corticospinal Neurons In
bioRxiv preprint doi: https://doi.org/10.1101/667733; this version posted June 12, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. 1 Early alterations of RNA metabolism and splicing from adult corticospinal neurons in 2 an ALS mouse model 3 4 Christine Marques1,2, Mathieu Fischer1,3, Céline Keime4, Thibaut Burg1, Aurore Brunet1, 5 Jelena Scekic-Zahirovic1 & Caroline Rouaux1* 6 7 8 9 1Inserm UMR_S 1118, Mécanismes centraux et périphériques de la neurodégénérescence, 10 Faculté de Médecine, Université de Strasbourg, Strasbourg, France. 11 2Current address: Department of Neurobiology, Harvard Medical School, Boston, MA, USA; 12 Department of Neurology, Massachusetts General Hospital, Boston, MA, USA. 13 3Current address: Department of Paediatrics, John Radcliffe Hospital, University of Oxford, 14 Oxford, UK. 15 4Inserm UMR_S 1258, CRNS UMR_S 7104, Université de Strasbourg, IGBMC, Strasbourg, 16 France. 17 18 *Correspondence should be addressed to: C.R. ([email protected]) 1 bioRxiv preprint doi: https://doi.org/10.1101/667733; this version posted June 12, 2019. The copyright holder for this preprint (which was not certified by peer review) is the author/funder. All rights reserved. No reuse allowed without permission. Abstract Amyotrophic lateral sclerosis (ALS) is a devastating neurodegenerative disease clinically defined as the combined degeneration of corticospinal and corticobulbar neurons (CSN), and bulbar and spinal motor neurons (MN). A growing body of evidence points to the motor cortex, where CSN are located, as the potential initiation site of ALS. However, little is known about the spatiotemporal dynamics of CSN degeneration and the molecular pathways involved. -
WO 2019/079361 Al 25 April 2019 (25.04.2019) W 1P O PCT
(12) INTERNATIONAL APPLICATION PUBLISHED UNDER THE PATENT COOPERATION TREATY (PCT) (19) World Intellectual Property Organization I International Bureau (10) International Publication Number (43) International Publication Date WO 2019/079361 Al 25 April 2019 (25.04.2019) W 1P O PCT (51) International Patent Classification: CA, CH, CL, CN, CO, CR, CU, CZ, DE, DJ, DK, DM, DO, C12Q 1/68 (2018.01) A61P 31/18 (2006.01) DZ, EC, EE, EG, ES, FI, GB, GD, GE, GH, GM, GT, HN, C12Q 1/70 (2006.01) HR, HU, ID, IL, IN, IR, IS, JO, JP, KE, KG, KH, KN, KP, KR, KW, KZ, LA, LC, LK, LR, LS, LU, LY, MA, MD, ME, (21) International Application Number: MG, MK, MN, MW, MX, MY, MZ, NA, NG, NI, NO, NZ, PCT/US2018/056167 OM, PA, PE, PG, PH, PL, PT, QA, RO, RS, RU, RW, SA, (22) International Filing Date: SC, SD, SE, SG, SK, SL, SM, ST, SV, SY, TH, TJ, TM, TN, 16 October 2018 (16. 10.2018) TR, TT, TZ, UA, UG, US, UZ, VC, VN, ZA, ZM, ZW. (25) Filing Language: English (84) Designated States (unless otherwise indicated, for every kind of regional protection available): ARIPO (BW, GH, (26) Publication Language: English GM, KE, LR, LS, MW, MZ, NA, RW, SD, SL, ST, SZ, TZ, (30) Priority Data: UG, ZM, ZW), Eurasian (AM, AZ, BY, KG, KZ, RU, TJ, 62/573,025 16 October 2017 (16. 10.2017) US TM), European (AL, AT, BE, BG, CH, CY, CZ, DE, DK, EE, ES, FI, FR, GB, GR, HR, HU, ΓΕ , IS, IT, LT, LU, LV, (71) Applicant: MASSACHUSETTS INSTITUTE OF MC, MK, MT, NL, NO, PL, PT, RO, RS, SE, SI, SK, SM, TECHNOLOGY [US/US]; 77 Massachusetts Avenue, TR), OAPI (BF, BJ, CF, CG, CI, CM, GA, GN, GQ, GW, Cambridge, Massachusetts 02139 (US). -
Gene Expression Profiling Analysis Contributes to Understanding the Association Between Non-Syndromic Cleft Lip and Palate, and Cancer
2110 MOLECULAR MEDICINE REPORTS 13: 2110-2116, 2016 Gene expression profiling analysis contributes to understanding the association between non-syndromic cleft lip and palate, and cancer HONGYI WANG, TAO QIU, JIE SHI, JIULONG LIANG, YANG WANG, LIANGLIANG QUAN, YU ZHANG, QIAN ZHANG and KAI TAO Department of Plastic Surgery, General Hospital of Shenyang Military Area Command, PLA, Shenyang, Liaoning 110016, P.R. China Received March 10, 2015; Accepted December 18, 2015 DOI: 10.3892/mmr.2016.4802 Abstract. The present study aimed to investigate the for NSCL/P were implicated predominantly in the TGF-β molecular mechanisms underlying non-syndromic cleft lip, signaling pathway, the cell cycle and in viral carcinogenesis. with or without cleft palate (NSCL/P), and the association The TP53, CDK1, SMAD3, PIK3R1 and CASP3 genes were between this disease and cancer. The GSE42589 data set found to be associated, not only with NSCL/P, but also with was downloaded from the Gene Expression Omnibus data- cancer. These results may contribute to a better understanding base, and contained seven dental pulp stem cell samples of the molecular mechanisms of NSCL/P. from children with NSCL/P in the exfoliation period, and six controls. Differentially expressed genes (DEGs) were Introduction screened using the RankProd method, and their potential functions were revealed by pathway enrichment analysis and Non-syndromic cleft lip, with or without cleft palate (NSCL/P) construction of a pathway interaction network. Subsequently, is one of the most common types of congenital defect and cancer genes were obtained from six cancer databases, and affects 3.4-22.9/10,000 individuals worldwide (1). -
Apoptotic Genes As Potential Markers of Metastatic Phenotype in Human Osteosarcoma Cell Lines
17-31 10/12/07 14:53 Page 17 INTERNATIONAL JOURNAL OF ONCOLOGY 32: 17-31, 2008 17 Apoptotic genes as potential markers of metastatic phenotype in human osteosarcoma cell lines CINZIA ZUCCHINI1, ANNA ROCCHI2, MARIA CRISTINA MANARA2, PAOLA DE SANCTIS1, CRISTINA CAPANNI3, MICHELE BIANCHINI1, PAOLO CARINCI1, KATIA SCOTLANDI2 and LUISA VALVASSORI1 1Dipartimento di Istologia, Embriologia e Biologia Applicata, Università di Bologna, Via Belmeloro 8, 40126 Bologna; 2Laboratorio di Ricerca Oncologica, Istituti Ortopedici Rizzoli; 3IGM-CNR, Unit of Bologna, c/o Istituti Ortopedici Rizzoli, Via di Barbiano 1/10, 40136 Bologna, Italy Received May 29, 2007; Accepted July 19, 2007 Abstract. Metastasis is the most frequent cause of death among malignant primitive bone tumor, usually developing in children patients with osteosarcoma. We have previously demonstrated and adolescents, with a high tendency to metastasize (2). in independent experiments that the forced expression of Metastases in osteosarcoma patients spread through peripheral L/B/K ALP and CD99 in U-2 OS osteosarcoma cell lines blood very early and colonize primarily the lung, and later markedly reduces the metastatic ability of these cancer cells. other skeleton districts (3). Since disseminated hidden micro- This behavior makes these cell lines a useful model to assess metastases are present in 80-90% of OS patients at the time the intersection of multiple and independent gene expression of diagnosis, the identification of markers of invasiveness signatures concerning the biological problem of dissemination. and metastasis forms a target of paramount importance in With the aim to characterize a common transcriptional profile planning the treatment of osteosarcoma lesions and enhancing reflecting the essential features of metastatic behavior, we the prognosis. -
GPRC5A Suppresses Protein Synthesis at the Endoplasmic Reticulum to Prevent Radiation-Induced Lung Tumorigenesis
ARTICLE Received 8 Mar 2016 | Accepted 28 Apr 2016 | Published 8 Jun 2016 DOI: 10.1038/ncomms11795 OPEN GPRC5A suppresses protein synthesis at the endoplasmic reticulum to prevent radiation-induced lung tumorigenesis Jian Wang1, Alton B. Farris2, Kaiming Xu1, Ping Wang1, Xiangming Zhang1, Duc M. Duong3, Hong Yi4, Hui-Kuo Shu1, Shi-Yong Sun5 & Ya Wang1 GPRC5A functions as a lung tumour suppressor to prevent spontaneous and environmentally induced lung carcinogenesis; however, the underlying mechanism remains unclear. Here we reveal that GPRC5A at the endoplasmic reticulum (ER) membrane suppresses synthesis of the secreted or membrane-bound proteins including a number of oncogenes, the most important one being Egfr. The ER-located GPRC5A disturbs the assembly of the eIF4F-mediated translation initiation complex on the mRNA cap through directly binding to the eIF4F complex with its two middle extracellular loops. Particularly, suppression of EGFR by GPRC5A contributes significantly to preventing ionizing radiation (IR)-induced lung tumorigenesis. Thus, GPRC5A deletion enhances IR-promoted EGFR expression through an increased translation rate, thereby significantly increasing lung tumour incidence in Gprc5a À / À mice. Our findings indicate that under-expressed GPRC5A during lung tumor- igenesis enhances any transcriptional stimulation through an active translational status, which can be used to control oncogene expression and potentially the resulting related disease. 1 Department of Radiation Oncology, Emory University School of Medicine and the Winship Cancer Institute, Emory University, Atlanta, Georgia 30322, USA. 2 Department of Pathology, Emory University School of Medicine and the Winship Cancer Institute, Emory University, Atlanta, Georgia 30322, USA. 3 Emory Integrated Proteomics Core and Biochemistry Department, Atlanta, Georgia 30322, USA. -
Relevance of Translation Initiation in Diffuse Glioma Biology and Its
cells Review Relevance of Translation Initiation in Diffuse Glioma Biology and its Therapeutic Potential Digregorio Marina 1, Lombard Arnaud 1,2, Lumapat Paul Noel 1, Scholtes Felix 1,2, Rogister Bernard 1,3 and Coppieters Natacha 1,* 1 Laboratory of Nervous System Disorders and Therapy, GIGA-Neurosciences Research Centre, University of Liège, 4000 Liège, Belgium; [email protected] (D.M.); [email protected] (L.A.); [email protected] (L.P.N.); [email protected] (S.F.); [email protected] (R.B.) 2 Department of Neurosurgery, CHU of Liège, 4000 Liège, Belgium 3 Department of Neurology, CHU of Liège, 4000 Liège, Belgium * Correspondence: [email protected] Received: 18 October 2019; Accepted: 26 November 2019; Published: 29 November 2019 Abstract: Cancer cells are continually exposed to environmental stressors forcing them to adapt their protein production to survive. The translational machinery can be recruited by malignant cells to synthesize proteins required to promote their survival, even in times of high physiological and pathological stress. This phenomenon has been described in several cancers including in gliomas. Abnormal regulation of translation has encouraged the development of new therapeutics targeting the protein synthesis pathway. This approach could be meaningful for glioma given the fact that the median survival following diagnosis of the highest grade of glioma remains short despite current therapy. The identification of new targets for the development of novel therapeutics is therefore needed in order to improve this devastating overall survival rate. This review discusses current literature on translation in gliomas with a focus on the initiation step covering both the cap-dependent and cap-independent modes of initiation. -
Proteomic Analysis of Mtor Inhibition-Mediated Phosphorylation
Protein Cell DOI 10.1007/s13238-016-0279-0 Protein & Cell LETTER Proteomic analysis of mTOR inhibition- mediated phosphorylation changes in ribosomal proteins and eukaryotic translation initiation factors Dear Editor, analysis based on the SILAC (stable isotope labeling by amino acids in cell culture) method was carried out to analyze affinity The mammalian target of rapamycin (mTOR), as a critical enriched phosphoproteins from the untreated and rapamycin- Cell energy sensor and cell-growth regulator, controls protein treated 293T cells. The experimental workflow was displayed & 12 14 synthesis, autophagy and many important cellular processes in Fig. 1A. Briefly, cells grown in light medium ( C6 N2-Lysine 12 0 0 through forming functional distinct complexes, mTORC1 and and C6-Arginine, K R ) were treated with 200 nmol/L rapa- 13 15 mTORC2. mTORC1 that is sensitive to rapamycin, regulates mycin for 2 h, while cells grown in heavy medium ( C6 N2- 13 8 6 cell growth and protein synthesis, while mTORC2 that is Lysine and C6-Arginine, K R ) were untreated. Sucrose insensitive to rapamycin, regulates cellular metabolism and cushion centrifugation was used to isolate ribosomes. Pro- Protein the cytoskeletal organization (Gingras et al., 2001; Hay and teins extracted from the whole cell lysate or the isolated ribo- Sonenberg, 2004). Translation initiation is the rate-limiting some fraction of the untreated and rapamycin-treated cells step in protein synthesis, which proceeds through a multi- were mixed and trypsin digested. Then phosphopeptides step process that can be divided into three major steps. First, were enriched with TiO2 beads and analyzed by nano-LC-MS/ eukaryotic translation initiation factor 2 (eIF2) binds with GTP MS.