Proteins, Peptides and Enzymes 2010
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Neuregulin-1 Potentiates Agrin-Induced Acetylcholine Receptor Clustering Through Muscle-Specific Kinase Phosphorylation
Research Article 1531 Neuregulin-1 potentiates agrin-induced acetylcholine receptor clustering through muscle-specific kinase phosphorylation Shyuan T. Ngo1, Rebecca N. Cole3, Nana Sunn2, William D. Phillips3,* and Peter G. Noakes1,2,*,` 1School of Biomedical Sciences, University of Queensland, St. Lucia, 4072, Queensland, Australia 2Queensland Brain Institute, University of Queensland, St. Lucia, 4072, Queensland, Australia 3Physiology and Bosch Institute, The University of Sydney, New South Wales, 2006, Australia *These authors contributed equally to this work `Author for correspondence ([email protected]) Accepted 14 November 2011 Journal of Cell Science 125, 1531–1543 ß 2012. Published by The Company of Biologists Ltd doi: 10.1242/jcs.095109 Summary At neuromuscular synapses, neural agrin (n-agrin) stabilizes embryonic postsynaptic acetylcholine receptor (AChR) clusters by signalling through the muscle-specific kinase (MuSK) complex. Live imaging of cultured myotubes showed that the formation and disassembly of primitive AChR clusters is a dynamic and reversible process favoured by n-agrin, and possibly other synaptic signals. Neuregulin-1 is a growth factor that can act through muscle ErbB receptor kinases to enhance synaptic gene transcription. Recent studies suggest that neuregulin-1–ErbB signalling can modulate n-agrin-induced AChR clustering independently of its effects on transcription. Here we report that neuregulin-1 increased the size of developing AChR clusters when injected into muscles of embryonic mice. We investigated this phenomenon using cultured myotubes, and found that in the ongoing presence of n-agrin, neuregulin-1 potentiates AChR clustering by increasing the tyrosine phosphorylation of MuSK. This potentiation could be blocked by inhibiting Shp2, a postsynaptic tyrosine phosphatase known to modulate the activity of MuSK. -
UBE2M (Mouse; Full Length), Pab
UBE2M (mouse; full length), pAb Alternate Names: Nedd8-conjugating enzyme, Ubc12, UBC-RS2, UBC12. Cat. No. 68-0025-100 Quantity: 100 µg Lot. No. 30262 Storage: -20˚C FOR RESEARCH USE ONLY NOT FOR USE IN HUMANS CERTIFICATE OF ANALYSIS Page 1 of 2 This antibody was developed and Physical Characteristics validated by the Medical Research Council Protein Phosphorylation and Quantity: 100 μg Formulation: phosphate-buffered Ubiquitylation Unit (University of saline Dundee, Dundee, UK). Concentration: to be provided on shipping Specificity:detects Ube2M at ~22 kDa Source: sheep polyclonal antibody Reactivity: mouse; other species not Background tested. Immunogen: mouse Ube2M (residues 1-183) [GST-tagged] Stability/Storage: 12 months at The enzymes of the NEDDylation pathway -20˚C; aliquot as required play a pivotal role in a number of cellular Purification:affinity-purified using processes including the indirect regula- immobilized immunogen tion and targeting of substrate proteins for proteasomal degradation. Three classes of enzymes are involved in the process of NEDDylation; the ubiquitin-like activating Research Applications and Quality Assurance enzyme APP-BP1/Uba3 (E1), the ubiquitin- Western Immunoblotting: Immunoprecipitation: like conjugating enzymes (E2s) and pro- Use 0.5 µg/ml Not tested tein ligases (E3s). UBE2M is a member of the E2 conjugating enzyme family and the gene for human UBE2M was first de- scribed by Osaka et al. (1998) and shares Dot Blotting Analysis: By dot blot assay the specific 42% sequence identity with yeast UBE2M. recognition of recombinant A trapped ubiquitin like activation complex Ube2M protein was observed has been described for the NEDD8 pathway under native and denaturing comprising, the E1 APP-BP1/Uba3, two conditions when probed with NEDD8 molecules, UBE2M and MgATP. -
Regulation of Cardiac Progenitors by Combination of Mesp1 and ETS
REGULATION OF CARDIAC PROGENITORS BY COMBINATION OF MESP1 AND ETS TRANSCRIPTION FACTORS A Dissertation by KUO-CHAN WENG Submitted to the Office of Graduate and Professional Studies of Texas A&M University in partial fulfillment of the requirements for the degree of DOCTOR OF PHILOSOPHY Chair of Committee, Fen Wang Co-Chair of Committee, Robert J. Schwartz Committee Members, James F. Martin Jiang Chang Head of Department, Fen Wang May 2014 Major Subject: Medical Science Copyright 2014 Kuo-Chan Weng ABSTRACT Heart disease remains the leading cause of death worldwide. By understanding the regulating networks during cardiac development we can exploit those networks to manipulate adult cells into cardiac progenitors and provide an alternative for repairing diseased hearts. Mesp1 is considered to have critical roles during cardiac development but the molecular mechanisms need to be further studied. The roles of ETS transcription factors have been primarily limited to hematopoietic differentiation and cancer progression. The ETS transcription factors are known to have proliferating roles and were hypothesized to also be involved in cardiac differentiation and may potentially be used for cell reprogramming. The first part of this study characterizes the expression pattern of Mesp1 protein in early mouse embryo from E6.5 to E9.5 and provides a full expression profile in differentiating embryoid bodies in vitro from the undifferentiated stage to Day10. Our work showed Mesp1 expresses in the posterior region of E6.5 embryo then starts migrating through the primitive streak toward anterior mesoderm and endoderm in E7.5. A Mesp1 linage tracing ES cell line was established, and it allowed us to trace the Mesp1 derived cell population. -
The HECT Domain Ubiquitin Ligase HUWE1 Targets Unassembled Soluble Proteins for Degradation
OPEN Citation: Cell Discovery (2016) 2, 16040; doi:10.1038/celldisc.2016.40 ARTICLE www.nature.com/celldisc The HECT domain ubiquitin ligase HUWE1 targets unassembled soluble proteins for degradation Yue Xu1, D Eric Anderson2, Yihong Ye1 1Laboratory of Molecular Biology, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD, USA; 2Advanced Mass Spectrometry Core Facility, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Bethesda, MD, USA In eukaryotes, many proteins function in multi-subunit complexes that require proper assembly. To maintain complex stoichiometry, cells use the endoplasmic reticulum-associated degradation system to degrade unassembled membrane subunits, but how unassembled soluble proteins are eliminated is undefined. Here we show that degradation of unassembled soluble proteins (referred to as unassembled soluble protein degradation, USPD) requires the ubiquitin selective chaperone p97, its co-factor nuclear protein localization protein 4 (Npl4), and the proteasome. At the ubiquitin ligase level, the previously identified protein quality control ligase UBR1 (ubiquitin protein ligase E3 component n-recognin 1) and the related enzymes only process a subset of unassembled soluble proteins. We identify the homologous to the E6-AP carboxyl terminus (homologous to the E6-AP carboxyl terminus) domain-containing protein HUWE1 as a ubiquitin ligase for substrates bearing unshielded, hydrophobic segments. We used a stable isotope labeling with amino acids-based proteomic approach to identify endogenous HUWE1 substrates. Interestingly, many HUWE1 substrates form multi-protein com- plexes that function in the nucleus although HUWE1 itself is cytoplasmically localized. Inhibition of nuclear entry enhances HUWE1-mediated ubiquitination and degradation, suggesting that USPD occurs primarily in the cytoplasm. -
Ubiquitination Is Not Omnipresent in Myeloid Leukemia Ramesh C
Editorials Ubiquitination is not omnipresent in myeloid leukemia Ramesh C. Nayak1 and Jose A. Cancelas1,2 1Division of Experimental Hematology and Cancer Biology, Cincinnati Children’s Hospital Medical Center and 2Hoxworth Blood Center, University of Cincinnati Academic Health Center, Cincinnati, OH, USA E-mail: JOSE A. CANCELAS - [email protected] / [email protected] doi:10.3324/haematol.2019.224162 hronic myelogenous leukemia (CML) is a clonal tination of target proteins through their cognate E3 ubiq- biphasic hematopoietic disorder most frequently uitin ligases belonging to three different families (RING, Ccaused by the expression of the BCR-ABL fusion HERCT, RING-between-RING or RBR type E3).7 protein. The expression of BCR-ABL fusion protein with The ubiquitin conjugating enzymes including UBE2N constitutive and elevated tyrosine kinase activity is suffi- (UBC13) and UBE2C are over-expressed in a myriad of cient to induce transformation of hematopoietic stem tumors such as breast, pancreas, colon, prostate, lym- cells (HSC) and the development of CML.1 Despite the phoma, and ovarian carcinomas.8 Higher expression of introduction of tyrosine kinase inhibitors (TKI), the dis- UBE2A is associated with poor prognosis of hepatocellu- ease may progress from a manageable chronic phase to a lar cancer.9 In leukemia, bone marrow (BM) cells from clinically challenging blast crisis phase with a poor prog- pediatric acute lymphoblastic patients show higher levels nosis,2 in which myeloid or lymphoid blasts fail to differ- of UBE2Q2 -
Detailed Review Paper on Retinoid Pathway Signalling
1 1 Detailed Review Paper on Retinoid Pathway Signalling 2 December 2020 3 2 4 Foreword 5 1. Project 4.97 to develop a Detailed Review Paper (DRP) on the Retinoid System 6 was added to the Test Guidelines Programme work plan in 2015. The project was 7 originally proposed by Sweden and the European Commission later joined the project as 8 a co-lead. In 2019, the OECD Secretariat was added to coordinate input from expert 9 consultants. The initial objectives of the project were to: 10 draft a review of the biology of retinoid signalling pathway, 11 describe retinoid-mediated effects on various organ systems, 12 identify relevant retinoid in vitro and ex vivo assays that measure mechanistic 13 effects of chemicals for development, and 14 Identify in vivo endpoints that could be added to existing test guidelines to 15 identify chemical effects on retinoid pathway signalling. 16 2. This DRP is intended to expand the recommendations for the retinoid pathway 17 included in the OECD Detailed Review Paper on the State of the Science on Novel In 18 vitro and In vivo Screening and Testing Methods and Endpoints for Evaluating 19 Endocrine Disruptors (DRP No 178). The retinoid signalling pathway was one of seven 20 endocrine pathways considered to be susceptible to environmental endocrine disruption 21 and for which relevant endpoints could be measured in new or existing OECD Test 22 Guidelines for evaluating endocrine disruption. Due to the complexity of retinoid 23 signalling across multiple organ systems, this effort was foreseen as a multi-step process. -
Using the Deadly M-Conotoxins As Probes of Voltage-Gated Sodium Channels
Toxicon 44 (2004) 117–122 www.elsevier.com/locate/toxicon Mini-review Using the deadly m-conotoxins as probes of voltage-gated sodium channels Ronald A. Li*, Gordon F. Tomaselli The Johns Hopkins University School of Medicine, 720 Rutland Avenue, Ross 871, Baltimore, MD 21205, USA Accepted 23 March 2004 Available online 19 June 2004 Abstract m-Conotoxins (m-CTX) are potent Na channel inhibitory peptides isolated from the venom of the predatory marine snail Conus geographus. m-CTXs exert their biological action by physically occluding the ion-conducting pore of voltage-gated Na (Nav) channels with a 1:1 stoichiometry in an all-or-none fashion. This article reviews our current knowledge of the mechanism of m-CTX and the associated structural and functional insights into its molecular target—Nav channels. q 2004 Elsevier Ltd. All rights reserved. Keywords: Na channel; Pore; m-Conotoxin Contents 1. Well-defined primary and 3-dimensional structures of m-CTX .............................. 117 2. Molecular target of m-CTX: voltage-gated Naþ channels . ................................. 119 3. m-CTX-pore interactions are site-specific.............................................. 119 4. Docking orientation of m-CTX ..................................................... 119 5. Isoform-specificity of m-CTX block ................................................. 121 6. m-CTX versus Kþ channel pore-blocking toxins ........................................ 121 7. Conclusion.................................................................... 121 Acknowledgements -
` Probing the Epigenome Andrea Huston1, Cheryl H Arrowsmith1,2
` Probing the Epigenome Andrea Huston1, Cheryl H Arrowsmith1,2, Stefan Knapp3,4,*, Matthieu Schapira1,5,* 1. Structural Genomics Consortium, University of Toronto, Toronto, ON M5G 1L7, Canada 2. Princess Margaret Cancer Centre and Department of Medical Biophysics, University of Toronto , Toronto, ON M5G 1L7, Canada 3. Nuffield Department of Clinical Medicine, Target Discovery Institute, and Structural Genomic Consortium, University of Oxford, Headington, Oxford OX3 7DQ, United Kingdom 4. Institute for Pharmaceutical Chemistry, Johann Wolfgang Goethe University, D-60438 Frankfurt am Main, Germany 5. Department of Pharmacology and Toxicology, University of Toronto, Toronto, ON M5S 1A8, Canada * Correspondence: [email protected], [email protected] Epigenetic chemical probes are having a strong impact on biological discovery and target validation. Systematic coverage of emerging epigenetic target classes with these potent, selective, cell-active chemical tools will profoundly influence our understanding of the human biology and pathology of chromatin-templated mechanisms. ` Chemical probes are research-enablers Advances in genomics and proteomics methodologies in recent years have made it possible to associate thousands of genes and proteins with specific diseases, biological processes, molecular networks and pathways. However, data from these large scale initiatives alone has not translated widely into new studies on these disease-associated proteins, and the biomedical research community still tends to focus on proteins that were already known before the sequencing of the human genome1. The human kinome for instance, a target class of direct relevance to cancer and other disease areas, is a telling example: based on the number of research articles indexed in pubmed in 2011, 75% of the research activity focused on only 10% of the 518 human kinases – largely the same kinases that were the focus of research before sequencing of the human genome - while 60% of the kinome, some 300 enzymes, was virtually ignored by the community2. -
Table S1 the Four Gene Sets Derived from Gene Expression Profiles of Escs and Differentiated Cells
Table S1 The four gene sets derived from gene expression profiles of ESCs and differentiated cells Uniform High Uniform Low ES Up ES Down EntrezID GeneSymbol EntrezID GeneSymbol EntrezID GeneSymbol EntrezID GeneSymbol 269261 Rpl12 11354 Abpa 68239 Krt42 15132 Hbb-bh1 67891 Rpl4 11537 Cfd 26380 Esrrb 15126 Hba-x 55949 Eef1b2 11698 Ambn 73703 Dppa2 15111 Hand2 18148 Npm1 11730 Ang3 67374 Jam2 65255 Asb4 67427 Rps20 11731 Ang2 22702 Zfp42 17292 Mesp1 15481 Hspa8 11807 Apoa2 58865 Tdh 19737 Rgs5 100041686 LOC100041686 11814 Apoc3 26388 Ifi202b 225518 Prdm6 11983 Atpif1 11945 Atp4b 11614 Nr0b1 20378 Frzb 19241 Tmsb4x 12007 Azgp1 76815 Calcoco2 12767 Cxcr4 20116 Rps8 12044 Bcl2a1a 219132 D14Ertd668e 103889 Hoxb2 20103 Rps5 12047 Bcl2a1d 381411 Gm1967 17701 Msx1 14694 Gnb2l1 12049 Bcl2l10 20899 Stra8 23796 Aplnr 19941 Rpl26 12096 Bglap1 78625 1700061G19Rik 12627 Cfc1 12070 Ngfrap1 12097 Bglap2 21816 Tgm1 12622 Cer1 19989 Rpl7 12267 C3ar1 67405 Nts 21385 Tbx2 19896 Rpl10a 12279 C9 435337 EG435337 56720 Tdo2 20044 Rps14 12391 Cav3 545913 Zscan4d 16869 Lhx1 19175 Psmb6 12409 Cbr2 244448 Triml1 22253 Unc5c 22627 Ywhae 12477 Ctla4 69134 2200001I15Rik 14174 Fgf3 19951 Rpl32 12523 Cd84 66065 Hsd17b14 16542 Kdr 66152 1110020P15Rik 12524 Cd86 81879 Tcfcp2l1 15122 Hba-a1 66489 Rpl35 12640 Cga 17907 Mylpf 15414 Hoxb6 15519 Hsp90aa1 12642 Ch25h 26424 Nr5a2 210530 Leprel1 66483 Rpl36al 12655 Chi3l3 83560 Tex14 12338 Capn6 27370 Rps26 12796 Camp 17450 Morc1 20671 Sox17 66576 Uqcrh 12869 Cox8b 79455 Pdcl2 20613 Snai1 22154 Tubb5 12959 Cryba4 231821 Centa1 17897 -
Induction of Myasthenia by Immunization Against Muscle- Specific Kinase
Induction of myasthenia by immunization against muscle- specific kinase Kazuhiro Shigemoto, … , Norifumi Ueda,, Seiji Matsuda J Clin Invest. 2006;116(4):1016-1024. https://doi.org/10.1172/JCI21545. Research Article Autoimmunity Muscle-specific kinase (MuSK) is critical for the synaptic clustering of nicotinic acetylcholine receptors (AChRs) and plays multiple roles in the organization and maintenance of neuromuscular junctions (NMJs). MuSK is activated by agrin, which is released from motoneurons, and induces AChR clustering at the postsynaptic membrane. Although autoantibodies against the ectodomain of MuSK have been found in a proportion of patients with generalized myasthenia gravis (MG), it is unclear whether MuSK autoantibodies are the causative agent of generalized MG. In the present study, rabbits immunized with MuSK ectodomain protein manifested MG-like muscle weakness with a reduction of AChR clustering at the NMJs. The autoantibodies activated MuSK and blocked AChR clustering induced by agrin or by mediators that do not activate MuSK. Thus MuSK autoantibodies rigorously inhibit AChR clustering mediated by multiple pathways, an outcome that broadens our general comprehension of the pathogenesis of MG. Find the latest version: https://jci.me/21545/pdf Research article Induction of myasthenia by immunization against muscle-specific kinase Kazuhiro Shigemoto,1,2 Sachiho Kubo,2 Naoki Maruyama,2 Naohito Hato,3 Hiroyuki Yamada,3 Chen Jie,4 Naoto Kobayashi,4 Katsumi Mominoki,5 Yasuhito Abe,6 Norifumi Ueda,6 and Seiji Matsuda4 1Department of Preventive Medicine, Ehime University School of Medicine, Ehime, Japan. 2Department of Molecular Pathology, Tokyo Metropolitan Institute for Gerontology, Tokyo, Japan. 3Department of Otolaryngology and 4Department of Integrated Basic Medical Science, Ehime University School of Medicine, Ehime, Japan. -
C-Myc Regulates Cell Proliferation During Lens Development
c-myc Regulates Cell Proliferation during Lens Development Gabriel R. Cavalheiro., Gabriel E. Matos-Rodrigues., Anielle L. Gomes, Paulo M. G. Rodrigues, Rodrigo A. P. Martins* Programa de Biologia Celular e do Desenvolvimento, Instituto de Cieˆncias Biome´dicas, Universidade Federal do Rio de Janeiro, Rio de Janeiro, Rio de Janeiro, Brazil Abstract Myc protooncogenes play important roles in the regulation of cell proliferation, growth, differentiation and survival during development. In various developing organs, c-myc has been shown to control the expression of cell cycle regulators and its misregulated expression is detected in many human tumors. Here, we show that c-myc gene (Myc) is highly expressed in developing mouse lens. Targeted deletion of c-myc gene from head surface ectoderm dramatically impaired ocular organogenesis, resulting in severe microphtalmia, defective anterior segment development, formation of a lens stalk and/or aphakia. In particular, lenses lacking c-myc presented thinner epithelial cell layer and growth impairment that was detectable soon after its inactivation. Defective development of c-myc-null lens was not caused by increased cell death of lens progenitor cells. Instead, c-myc loss reduced cell proliferation, what was associated with an ectopic expression of Prox1 and p27Kip1 proteins within epithelial cells. Interestingly, a sharp decrease in the expression of the forkhead box transcription factor Foxe3 was also observed following c-myc inactivation. These data represent the first description of the physiological roles played by a Myc family member in mouse lens development. Our findings support the conclusion that c- myc regulates the proliferation of lens epithelial cells in vivo and may, directly or indirectly, modulate the expression of classical cell cycle regulators in developing mouse lens. -
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.