Structure of the Eukaryotic Translation Initiation Factor Eif4e in Complex
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A Computational Approach for Defining a Signature of Β-Cell Golgi Stress in Diabetes Mellitus
Page 1 of 781 Diabetes A Computational Approach for Defining a Signature of β-Cell Golgi Stress in Diabetes Mellitus Robert N. Bone1,6,7, Olufunmilola Oyebamiji2, Sayali Talware2, Sharmila Selvaraj2, Preethi Krishnan3,6, Farooq Syed1,6,7, Huanmei Wu2, Carmella Evans-Molina 1,3,4,5,6,7,8* Departments of 1Pediatrics, 3Medicine, 4Anatomy, Cell Biology & Physiology, 5Biochemistry & Molecular Biology, the 6Center for Diabetes & Metabolic Diseases, and the 7Herman B. Wells Center for Pediatric Research, Indiana University School of Medicine, Indianapolis, IN 46202; 2Department of BioHealth Informatics, Indiana University-Purdue University Indianapolis, Indianapolis, IN, 46202; 8Roudebush VA Medical Center, Indianapolis, IN 46202. *Corresponding Author(s): Carmella Evans-Molina, MD, PhD ([email protected]) Indiana University School of Medicine, 635 Barnhill Drive, MS 2031A, Indianapolis, IN 46202, Telephone: (317) 274-4145, Fax (317) 274-4107 Running Title: Golgi Stress Response in Diabetes Word Count: 4358 Number of Figures: 6 Keywords: Golgi apparatus stress, Islets, β cell, Type 1 diabetes, Type 2 diabetes 1 Diabetes Publish Ahead of Print, published online August 20, 2020 Diabetes Page 2 of 781 ABSTRACT The Golgi apparatus (GA) is an important site of insulin processing and granule maturation, but whether GA organelle dysfunction and GA stress are present in the diabetic β-cell has not been tested. We utilized an informatics-based approach to develop a transcriptional signature of β-cell GA stress using existing RNA sequencing and microarray datasets generated using human islets from donors with diabetes and islets where type 1(T1D) and type 2 diabetes (T2D) had been modeled ex vivo. To narrow our results to GA-specific genes, we applied a filter set of 1,030 genes accepted as GA associated. -
Genes with 5' Terminal Oligopyrimidine Tracts Preferentially Escape Global Suppression of Translation by the SARS-Cov-2 NSP1 Protein
Downloaded from rnajournal.cshlp.org on September 28, 2021 - Published by Cold Spring Harbor Laboratory Press Genes with 5′ terminal oligopyrimidine tracts preferentially escape global suppression of translation by the SARS-CoV-2 Nsp1 protein Shilpa Raoa, Ian Hoskinsa, Tori Tonna, P. Daniela Garciaa, Hakan Ozadama, Elif Sarinay Cenika, Can Cenika,1 a Department of Molecular Biosciences, University of Texas at Austin, Austin, TX 78712, USA 1Corresponding author: [email protected] Key words: SARS-CoV-2, Nsp1, MeTAFlow, translation, ribosome profiling, RNA-Seq, 5′ TOP, Ribo-Seq, gene expression 1 Downloaded from rnajournal.cshlp.org on September 28, 2021 - Published by Cold Spring Harbor Laboratory Press Abstract Viruses rely on the host translation machinery to synthesize their own proteins. Consequently, they have evolved varied mechanisms to co-opt host translation for their survival. SARS-CoV-2 relies on a non-structural protein, Nsp1, for shutting down host translation. However, it is currently unknown how viral proteins and host factors critical for viral replication can escape a global shutdown of host translation. Here, using a novel FACS-based assay called MeTAFlow, we report a dose-dependent reduction in both nascent protein synthesis and mRNA abundance in cells expressing Nsp1. We perform RNA-Seq and matched ribosome profiling experiments to identify gene-specific changes both at the mRNA expression and translation level. We discover that a functionally-coherent subset of human genes are preferentially translated in the context of Nsp1 expression. These genes include the translation machinery components, RNA binding proteins, and others important for viral pathogenicity. Importantly, we uncovered a remarkable enrichment of 5′ terminal oligo-pyrimidine (TOP) tracts among preferentially translated genes. -
EIF4E Antibody Cat
EIF4E Antibody Cat. No.: 60-600 EIF4E Antibody Formalin-fixed and paraffin-embedded human breast carcinoma reacted with EIF4E antibody (N-term), which was peroxidase- conjugated to the secondary antibody, followed by DAB staining. Specifications HOST SPECIES: Rabbit SPECIES REACTIVITY: Human Predicted species reactivity based on immunogen sequence: Zebrafish, Bovine, Mouse, HOMOLOGY: Rabbit, Rat, Xenopus This EIF4E antibody is generated from rabbits immunized with a KLH conjugated synthetic IMMUNOGEN: peptide between 32-61 amino acids from the N-terminal region of human EIF4E. TESTED APPLICATIONS: IHC-P, WB For WB starting dilution is: 1:1000 APPLICATIONS: For IHC-P starting dilution is: 1:50~100 September 27, 2021 1 https://www.prosci-inc.com/eif4e-antibody-60-600.html PREDICTED MOLECULAR 25 kDa WEIGHT: Properties This antibody is prepared by Saturated Ammonium Sulfate (SAS) precipitation followed by PURIFICATION: dialysis CLONALITY: Polyclonal ISOTYPE: Rabbit Ig CONJUGATE: Unconjugated PHYSICAL STATE: Liquid BUFFER: Supplied in PBS with 0.09% (W/V) sodium azide. CONCENTRATION: batch dependent Store at 4˚C for three months and -20˚C, stable for up to one year. As with all antibodies STORAGE CONDITIONS: care should be taken to avoid repeated freeze thaw cycles. Antibodies should not be exposed to prolonged high temperatures. Additional Info OFFICIAL SYMBOL: EIF4E Eukaryotic translation initiation factor 4E, eIF-4E, eIF4E, eIF-4F 25 kDa subunit, mRNA cap- ALTERNATE NAMES: binding protein, EIF4E, EIF4EL1, EIF4F ACCESSION NO.: P06730 PROTEIN GI NO.: 1352435 GENE ID: 1977 USER NOTE: Optimal dilutions for each application to be determined by the researcher. Background and References eIF4F is a multi-subunit complex, the composition of which varies with external and internal environmental conditions. -
Initiation Factor Eif5b Catalyzes Second GTP-Dependent Step in Eukaryotic Translation Initiation
Initiation factor eIF5B catalyzes second GTP-dependent step in eukaryotic translation initiation Joon H. Lee*†, Tatyana V. Pestova†‡§, Byung-Sik Shin*, Chune Cao*, Sang K. Choi*, and Thomas E. Dever*¶ *Laboratory of Gene Regulation and Development, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, MD 20892-2716; ‡Department of Microbiology and Immunology, State University of New York Health Science Center, Brooklyn, NY 11203; and §A. N. Belozersky Institute of Physico-Chemical Biology, Moscow State University, Moscow, Russia Edited by Harry F. Noller, University of California, Santa Cruz, CA, and approved October 31, 2002 (received for review September 19, 2002) Initiation factors IF2 in bacteria and eIF2 in eukaryotes are GTPases In addition, when nonhydrolyzable GDPNP was substituted Met that bind Met-tRNAi to the small ribosomal subunit. eIF5B, the for GTP, eIF5B catalyzed subunit joining; however, the factor eukaryotic ortholog of IF2, is a GTPase that promotes ribosomal was unable to dissociate from the 80S ribosome after subunit subunit joining. Here we show that eIF5B GTPase activity is re- joining (7). quired for protein synthesis. Mutation of the conserved Asp-759 in To dissect the function of the eIF5B G domain and test the human eIF5B GTP-binding domain to Asn converts eIF5B to an model that two GTP molecules are required in translation XTPase and introduces an XTP requirement for subunit joining and initiation, we mutated conserved residues in the eIF5B G translation initiation. Thus, in contrast to bacteria where the single domain and tested the function of the mutant proteins in GTPase IF2 is sufficient to catalyze translation initiation, eukaryotic translation initiation. -
Early Growth Response 1 Regulates Hematopoietic Support and Proliferation in Human Primary Bone Marrow Stromal Cells
Hematopoiesis SUPPLEMENTARY APPENDIX Early growth response 1 regulates hematopoietic support and proliferation in human primary bone marrow stromal cells Hongzhe Li, 1,2 Hooi-Ching Lim, 1,2 Dimitra Zacharaki, 1,2 Xiaojie Xian, 2,3 Keane J.G. Kenswil, 4 Sandro Bräunig, 1,2 Marc H.G.P. Raaijmakers, 4 Niels-Bjarne Woods, 2,3 Jenny Hansson, 1,2 and Stefan Scheding 1,2,5 1Division of Molecular Hematology, Department of Laboratory Medicine, Lund University, Lund, Sweden; 2Lund Stem Cell Center, Depart - ment of Laboratory Medicine, Lund University, Lund, Sweden; 3Division of Molecular Medicine and Gene Therapy, Department of Labora - tory Medicine, Lund University, Lund, Sweden; 4Department of Hematology, Erasmus MC Cancer Institute, Rotterdam, the Netherlands and 5Department of Hematology, Skåne University Hospital Lund, Skåne, Sweden ©2020 Ferrata Storti Foundation. This is an open-access paper. doi:10.3324/haematol. 2019.216648 Received: January 14, 2019. Accepted: July 19, 2019. Pre-published: August 1, 2019. Correspondence: STEFAN SCHEDING - [email protected] Li et al.: Supplemental data 1. Supplemental Materials and Methods BM-MNC isolation Bone marrow mononuclear cells (BM-MNC) from BM aspiration samples were isolated by density gradient centrifugation (LSM 1077 Lymphocyte, PAA, Pasching, Austria) either with or without prior incubation with RosetteSep Human Mesenchymal Stem Cell Enrichment Cocktail (STEMCELL Technologies, Vancouver, Canada) for lineage depletion (CD3, CD14, CD19, CD38, CD66b, glycophorin A). BM-MNCs from fetal long bones and adult hip bones were isolated as reported previously 1 by gently crushing bones (femora, tibiae, fibulae, humeri, radii and ulna) in PBS+0.5% FCS subsequent passing of the cell suspension through a 40-µm filter. -
The Solution Structure of the Guanine Nucleotide Exchange Domain Of
View metadata, citation and similar papers at core.ac.uk brought to you by CORE Researchprovided Articleby Elsevier -217 Publisher Connector The solution structure of the guanine nucleotide exchange domain of human elongation factor 1b reveals a striking resemblance to that of EF-Ts from Escherichia coli Janice MJ Pérez1,2‡, Gregg Siegal2*‡, Jan Kriek1, Karl Hård2†, Jan Dijk1, Gerard W Canters2 and Wim Möller1 Background: In eukaryotic protein synthesis, the multi-subunit elongation Addresses: 1Department of Molecular Cell Biology, factor 1 (EF-1) plays an important role in ensuring the fidelity and regulating the Sylvius Laboratory, University of Leiden, Wassenaarseweg 72, NL-2333 AL Leiden, The rate of translation. EF-1α, which transports the aminoacyl tRNA to the Netherlands and 2Leiden Institute of Chemistry, β ribosome, is a member of the G-protein superfamily. EF-1 regulates the activity Gorlaeus Laboratory, University of Leiden, of EF-1α by catalyzing the exchange of GDP for GTP and thereby regenerating Einsteinweg 55, NL-2333 CC Leiden, The the active form of EF-1α. The structure of the bacterial analog of EF-1α, EF-Tu Netherlands. has been solved in complex with its GDP exchange factor, EF-Ts. These †Present address: Astra Structural Chemistry structures indicate a mechanism for GDP–GTP exchange in prokaryotes. Laboratory, S-43183 Mölndal, Sweden. Although there is good sequence conservation between EF-1α and EF-Tu, there is essentially no sequence similarity between EF-1β and EF-Ts. We ‡These two authors contributed equally to this work. wished to explore whether the prokaryotic exchange mechanism could shed any *Corresponding author. -
Mtorc1 Controls Mitochondrial Activity and Biogenesis Through 4E-BP-Dependent Translational Regulation
Cell Metabolism Article mTORC1 Controls Mitochondrial Activity and Biogenesis through 4E-BP-Dependent Translational Regulation Masahiro Morita,1,2 Simon-Pierre Gravel,1,2 Vale´ rie Che´ nard,1,2 Kristina Sikstro¨ m,3 Liang Zheng,4 Tommy Alain,1,2 Valentina Gandin,5,7 Daina Avizonis,2 Meztli Arguello,1,2 Chadi Zakaria,1,2 Shannon McLaughlan,5,7 Yann Nouet,1,2 Arnim Pause,1,2 Michael Pollak,5,6,7 Eyal Gottlieb,4 Ola Larsson,3 Julie St-Pierre,1,2,* Ivan Topisirovic,5,7,* and Nahum Sonenberg1,2,* 1Department of Biochemistry 2Goodman Cancer Research Centre McGill University, Montreal, QC H3A 1A3, Canada 3Department of Oncology-Pathology, Karolinska Institutet, Stockholm, 171 76, Sweden 4Cancer Research UK, The Beatson Institute for Cancer Research, Switchback Road, Glasgow G61 1BD, Scotland, UK 5Lady Davis Institute for Medical Research 6Cancer Prevention Center, Sir Mortimer B. Davis-Jewish General Hospital McGill University, Montreal, QC H3T 1E2, Canada 7Department of Oncology, McGill University, Montreal, QC H2W 1S6, Canada *Correspondence: [email protected] (J.S.-P.), [email protected] (I.T.), [email protected] (N.S.) http://dx.doi.org/10.1016/j.cmet.2013.10.001 SUMMARY ATP under physiological conditions in mammals and play a crit- ical role in overall energy balance (Vander Heiden et al., 2009). mRNA translation is thought to be the most energy- The mechanistic/mammalian target of rapamycin (mTOR) is a consuming process in the cell. Translation and serine/threonine kinase that has been implicated in a variety of energy metabolism are dysregulated in a variety of physiological processes and pathological states (Zoncu et al., diseases including cancer, diabetes, and heart 2011). -
Download Product Insert (PDF)
PRODUCT INFORMATION eIF4E (human recombinant) Item No. 15137 Overview and Properties Synonyms: eIF-4F 25 kDa subunit, Eukaryotic Translation Initiation Factor 4E, mRNA Cap-binding Protein Source: Recombinant protein expressed in E. coli Amino Acids: 2-217 (full length) Uniprot No.: P06730 Molecular Weight: 25.2 kDa Storage: -80°C (as supplied) Stability: ≥1 year Purity: batch specific (≥55% estimated by SDS-PAGE) Supplied in: 20 mM HEPES, pH 7.5, containing 100 mM potassium chloride, 2 mM DTT, and 10% glycerol Protein Concentration: batch specific mg/ml Image 1 2 3 4 250 kDa · · · · · · · 150 kDa · · · · · · · 100 kDa · · · · · · · 75 kDa · · · · · · · 50 kDa · · · · · · · 37 kDa · · · · · · · 25 kDa · · · · · · · 20 kDa · · · · · · · 15 kDa · · · · · · · Lane 1: MW Markers Lane 2: eIF4E (human recombinant) (4 µg) Lane 3: eIF4E (human recombinant) (2 µg) Lane 4: eIF4E (human recombinant) (1 µg) Representaõve gel image shown; actual purity may vary between each batch. WARNING CAYMAN CHEMICAL THIS PRODUCT IS FOR RESEARCH ONLY - NOT FOR HUMAN OR VETERINARY DIAGNOSTIC OR THERAPEUTIC USE. 1180 EAST ELLSWORTH RD SAFETY DATA ANN ARBOR, MI 48108 · USA This material should be considered hazardous until further information becomes available. Do not ingest, inhale, get in eyes, on skin, or on clothing. Wash thoroughly after handling. Before use, the user must review the complete Safety Data Sheet, which has been sent via email to your institution. PHONE: [800] 364-9897 WARRANTY AND LIMITATION OF REMEDY [734] 971-3335 Buyer agrees to purchase the material subject to Cayman’s Terms and Conditions. Complete Terms and Conditions including Warranty and Limitation of Liability information can be found on our website. -
Disome-Seq Reveals Widespread Ribosome Collisions That Recruit Co-Translational Chaperones
SUPPLEMENTARY INFORMATION FOR Disome-seq reveals widespread ribosome collisions that recruit co-translational chaperones T. Zhao, Y.-M. Chen, Y. Li, J. Wang, S. Chen, N. Gao, and W. Qian Supplementary information includes Supplementary Figures S1-11 and Supplementary Tables S1-5. Supplementary Figures Figure S1. Disomes persisted after RNase digestion. Samples containing an equal amount of ribosome-bound mRNA (5000 A260 unit) were treated with 100 U, 250 U, and 500 U RNase I, respectively. As the concentration of RNase I increased, the abundance of monosome reduced, and that of free ribonucleoprotein (RNP) increased, suggesting the disruption of ribosomes by the excessive RNase I digestion. However, the disome persisted – the mRNA fragment in- between was resistant to the RNase digestion likely due to the steric effect. Figure S2. Correlations between libraries of disome-seq, monosome-seq, and mRNA-seq. (A) The top three charts are the scatter plots of the number of mapped reads in each gene between biological replicates. The bottom chart shows the correlation of reads in each gene between disome-seq and monosome-seq, in the unit of reads per million. The average of two replicates is shown. All libraries were obtained from 3-AT treated yeast cells. Each dot represents a gene. The P-values were given by Pearson’s correlation. (B) Same to (A), except yeast cells were cultured in the rich medium. Figure S3. The size distribution of monosome and disome footprints. (A-B) We mapped the monosome (A) and disome (B) footprints obtained from 3-AT treated yeast cells to the yeast genome. -
Viral and Cellular Mrna-Specific Activators Harness PABP and Eif4g to Promote Translation Initiation Downstream of Cap Binding
Viral and cellular mRNA-specific activators harness PABP and eIF4G to promote translation initiation downstream of cap binding Richard W. P. Smitha,b,c,1, Ross C. Andersona,b,2, Osmany Larraldec,3, Joel W. S. Smithb, Barbara Gorgonia,b,4, William A. Richardsona,b, Poonam Malikc,d,5, Sheila V. Grahamc, and Nicola K. Graya,b,1 aMedical Research Council Centre for Reproductive Health, Queen’s Medical Research Institute, University of Edinburgh, Edinburgh EH16 4TJ, United Kingdom; bMedical Research Council Human Genetics Unit, University of Edinburgh, Western General Hospital, Edinburgh EH4 2XU, United Kingdom; cMedical Research Council-University of Glasgow Centre for Virus Research, Garscube Campus, Glasgow G61 1QH, United Kingdom; and dWellcome Trust Centre for Cell Biology and Institute of Cell Biology, University of Edinburgh, Edinburgh EH9 3BF, United Kingdom Edited by Nahum Sonenberg, McGill University, Montreal, QC, Canada, and approved April 28, 2017 (received for review July 4, 2016) Regulation of mRNA translation is a major control point for gene ribosomal subunit to form an 80S ribosome. Like the cap, the poly expression and is critical for life. Of central importance is the (A) tail serves as a primary determinant of translational efficiency (3) complex between cap-bound eukaryotic initiation factor 4E via the action of poly(A)-binding protein (PABP). Analysis of (eIF4E), eIF4G, and poly(A) tail-binding protein (PABP) that circu- mRNA-ribosomal subunit association has shown that PABP pro- larizes mRNAs, promoting translation and stability. This complex is motes small subunit recruitment, an activity attributed to its ability to often targeted to regulate overall translation rates, and also by stimulate cap binding by eIF4E (4). -
The Female Post-Mating Response Requires Genes Expressed in the Secondary Cells of the Male Accessory Gland in Drosophila Melanogaster
| INVESTIGATION The Female Post-Mating Response Requires Genes Expressed in the Secondary Cells of the Male Accessory Gland in Drosophila melanogaster Jessica L. Sitnik,*,1 Dragan Gligorov,†,1 Robert K. Maeda,† François Karch,†,2 and Mariana F. Wolfner*,2 *Department of Molecular Biology and Genetics, Cornell University, Ithaca, New York 14853, and †Department of Genetics and Evolution, University of Geneva, 1211 Geneva, Switzerland ORCID ID: 0000-0003-2701-9505 (M.F.W.) ABSTRACT Seminal proteins from the Drosophila male accessory gland induce post-mating responses (PMR) in females. The PMR comprise behavioral and physiological changes that include increased egg laying, decreased receptivity to courting males, and changes in the storage and use of sperm. Many of these changes are induced by a “sex peptide” (SP) and are maintained by SP’s binding to, and slow release from, sperm. The accessory gland contains two secretory cell types with distinct morphological and developmental characteristics. Products of these “main” and “secondary” cells work interdependently to induce and maintain the PMR. To identify individual genes needed for the morphology and function of secondary cells, we studied iab-6cocu males, whose secondary cells have abnormal morphology and fail to provide products to maintain the PMR. By RNA-seq, we identified 77 genes that are downregulated by a factor of .53 in iab-6cocu males. By functional assays and microscopy, we tested 20 candidate genes and found that at least 9 are required for normal storage and release of SP in mated females. Knockdown of each of these 9 genes consequently leads to a reduction in egg laying and an increase in receptivity over time, confirming a role for the secondary cells in maintaining the long-term PMR. -
Translation Initiation Factor Modifications and the Regulation of Protein Synthesis in Apoptotic Cells
Cell Death and Differentiation (2000) 7, 603 ± 615 ã 2000 Macmillan Publishers Ltd All rights reserved 1350-9047/00 $15.00 www.nature.com/cdd Translation initiation factor modifications and the regulation of protein synthesis in apoptotic cells ,1 1 1 2 MJ Clemens* , M Bushell , IW Jeffrey , VM Pain and Introduction SJ Morley2 Apoptosis is now recognized to be an important physiological 1 Department of Biochemistry and Immunology, Cellular and Molecular process by which cell and tissue growth, differentiation and Sciences Group, St George's Hospital Medical School, Cranmer Terrace, programmes of development are regulated. The molecular London SW17 ORE, UK mechanisms of apoptosis have been the subject of intense 2 Biochemistry Group, School of Biological Sciences, University of Sussex, research in recent years (for reviews see1±5). Cell death is Brighton BN1 9QG, UK induced following the stimulation of specific cell surface * Corresponding author: MJ Clemens, Department of Biochemistry and Immunology, Cellular and Molecular Sciences Group, St George's Hospital receptors such as the CD95 (Apo-1/Fas) antigen or the 6 Medical School, Cranmer Terrace, London SW17 ORE, UK. Tel: +44 20 8725 tumour necrosis factor-a (TNFa) receptor-1 (TNFR-1). It can 5770; Fax: +44 20 8725 2992; E-mail: [email protected] also result from intracellular events such as DNA damage or from a lack of specific growth factors. The relative importance Received 6.12.99; revised 25.1.00; accepted 20.3.00 of these different influences varies between cell types. The Edited by M Piacentini apoptotic process can be divided into a commitment phase and an execution phase.