DOCSLIB.ORG

Roles of Neddylation Against Viral Infections

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Roles of Neddylation Against Viral Infections Cellular & Molecular Immunology (2018) 15, 292–294 & 2018 CSI and USTC All rights reserved 2042-0226/18 $32.00 www.nature.com/cmi LETTER TO THE EDITOR Roles of neddylation against viral infections Kun Han and Jiyan Zhang Cellular & Molecular Immunology (2018) 15, 292–294; doi:10.1038/cmi.2017.100; published online 27 November 2017 he ubiquitin-dependent proteasome lentivirus by SAMHD1,7 or Rift Valley is carried out by certain CRLs members. Tpathway can be hijacked by certain fever virus by protein kinase R.8 Both Therefore, we speculated that neddyla- viruses to maintain viral genome ampli- CRLs-dependent and CRLs-independent tion should promote the HSV-1-induced fication. A key class of ubiquitin-E3s mechanisms have been suggested in this production of type I interferons, and this involved in this pathway is Cullin- regard.9 hypothesis was confirmed in our study, RING ligases (CRLs), which are activated As for the innate immune response, which was recently published in Cellular by an additional ubiquitin-like protein the involvement of neddylation was sug- & Molecular Immunology (Figure 1).14 NEDD8.1 The process by which the gested as early as the year 2006.10 The With primary macrophages deficient of ubiquitin-like protein NEDD8 is conju- transcription of type I interferon genes is the catalytic subunit of neddylation E1, gated to its target proteins is officially controlled by nuclear factor-κB (NF-κB) we have provided the first evidence that named ‘neddylation’. Consequently, ned- and interferon regulatory factor (IRF) neddylation indeed plays a role in type I dylation inhibition, through the use family members including IRF3. Many interferon production. Interestingly, ned- of the pharmacological inhibitor viruses execute immune-evasive activities dylation blockade only delays but does MLN4924, might prevent viral genome by targeting the type I interferon signal- not completely abrogate HSV-1-induced amplification.2,3 Herpesviruses, adeno- ing pathway. RNA viruses such as Sendi p65 nuclear translocation.14 Conse- virus, and influenza virus are MLN4924 virus (SeV), vesicular stomatitis virus, quently, the early phase type I interferon susceptible, whereas vaccinia virus and and rotavirus induce the degradation of production in HSV-1 infection is much – vesicular stomatitis virus are not.2,3 Ned- IRF3.10 12 However, a herpesvirus family more impaired than the late phase.14 dylation might also promote viral repli- member, herpes simplex virus type I Recently, the notion that the host also cation independent of CRLs.4 For (HSV-1), a DNA virus, has no such exploits the neddylation pathway to fight example, hepatitis B virus-encoded X effect.13 Even though the degradation of against virus infection has been sup- protein can act as an NEDD8 substrate IRF3 upon infection with rotavirus has ported by additional evidence. Zhang T and its neddylation by E3 ligase HDM2 been demonstrated to be independent of et al. reported that the polymerase basic enhances its stability and function in viral CRLs,11,12 a Cullin-based ubiquitin ligase protein 2 of influenza A virus can be replication and hepatocarcinogenesis.4 On pathway was reported to be involved in covalently modified by NEDD8, which the other hand, blockade of neddylation SeV-induced IRF3 degradation in 2006.10 reduces its stability and blocks the repli- prevents the degradation of host restric- Therefore, neddylation promotes IRF3 cation of influenza A virus.15 tion factors and thereby restores the degradation upon SeV infection.10 The- Another group later reported that restriction of human immunodeficiency oretically, neddylation should promote NEDD8 knockdown showed no effect virus by APOBEC3G and SAMHD1,5,6 IRF3-mediated transcription of type I on type I interferon production triggered interferon genes. by lipopolysaccharide (LPS), poly (I:C), Department of Molecular Immunology, Institute of In addition to IRF family members, or SeV.16 Since poly (I:C) and SeV Basic Medical Sciences, 27 Taiping Road, Beij- NF-κB also plays a key role in mediating induce IRF3 degradation, whereas ing 100850, China 10 Correspondence: Dr J Zhang, PhD, MD, the transcription of type I interferon HSV-1 does not, perhaps the interplay Department of Molecular Immunology, Institute genes. It is demonstrated that neddyla- between the NF-κB pathway and IRF of Basic Medical Sciences, 27 Taiping Road, tion contributes to NF-κB activity by pathway determines the outcome. Intri- Beijing, PA 100850, China fi E-mail: [email protected] promoting the ubiquitination and sub- guingly, MLN4924 signi cantly inhibited Received: 2 August 2017; Accepted: 14 August sequent degradation of IκBproteins type I interferon production triggered by 2017 since the ubiquitination of IκBproteins LPS, poly (I:C), or SeV,16 which has been Neddylation in viral infection K Han and J Zhang 293 Figure 1 Neddylation contributes to DNA virus-induced production of type I interferons. The transcription of type I interferon genes is controlled by nuclear factor-κB (NF-κB) and interferon regulatory factor (IRF) family members IRF3 and IRF7. DNA virus-induced NF-κB activation depends on the ubiquitination and subsequent degradation of IκB proteins. The ubiquitination of IκBproteinssuchasIκBα is carried out by a certain member of the Cullin-RING ligases (CRLs), that is, SKP1/Cullin-1/F-box protein β-TrCP/Roc1 complex. As CRLs are activated by the covalent conjugation of an additional ubiquitin-like protein NEDD8 to Cullins, neddylation promotes DNA virus-induced NF-κB activation and thereby promotes DNA virus-induced production of type I interferons. attributed by the authors to its off-target SAMHD1-mediated restriction of HIV-1. – effects. However, the potential off-target 1 Enchev RI, Schulman BA, Peter M. Protein J Virol 2013; 87: 11741 11750. neddylation: beyond cullin-RING ligases. Nat 7 Wei W, Guo H, Liu X, Zhang H, Qian L, Luo K effects of small interfering RNAs and Rev Mol Cell Biol 2015; 16:30–44. et al. A first-in-class NAE inhibitor, the relatively low efficiency of small 2 Hughes DJ, Wood JJ, Jackson BR, Baquero- MLN4924, blocks lentiviral infection in mye- loid cells by disrupting neddylation- RNA interference cannot be excluded. Perez B, Whitehouse A. NEDDylation is essential for Kaposi's sarcoma-associated dependent Vpx-mediated SAMHD1 degrada- Future studies are required to address herpesvirus latency and lytic reactivation tion. JVirol2014; 88:745–751. this issue. and represents a novel anti-KSHV target. 8 Mudhasani R, Tran JP, Retterer C, Kota KP, Whitehouse CA, Bavari S. Protein kinase R Together, these 10 years of studies Plos Pathog 2015; 11: e1004771. 3 Le-Trilling VT, Megger DA, Katschinski B, degradation is essential for rift valley fever suggest that neddylation might exert Landsberg CD, Rückborn MU, Tao S et al. virus infection and is regulated by SKP1- distinct roles in the infection of different Broad and potent antiviral activity of the NAE CUL1-F-box (SCF)FBXW11-NSs E3 ligase. inhibitor MLN4924. Sci Rep 2016; 6: Plos Pathog 2016; 12: e1005437. viruses. Targeting the neddylation 19977. 9 Ramirez PW, DePaula-Silva AB, Szaniawski M, pathway against virus infection, though 4 Liu N, Zhang J, Yang X, Jiao T, Zhao X, Li W Barker E, Bosque A, Planelles V. HIV-1 Vpu hopeful under certain circumstances, et al. HDM2 promotes NEDDylation of HBV utilizes both cullin-RING ligase (CRL) HBx to enhance its stability and function. dependent and independent mechanisms should be cast with caution and JVirol2017; 91:e00340–17. to downmodulate host proteins. Retrovirol- requires extensive studies to clarify how 5 Stanley DJ, Bartholomeeusen K, Crosby DC, ogy 2015; 12: 65. neddylation works in each step of virus Kim DY, Kwon E, Yen L et al. Inhibition of a 10 Bibeau-Poirier A, Gravel SP, Clément JF, NEDD8 cascade restores restriction of HIV by Rolland S, Rodier G, Coulombe P et al. infection. APOBEC3G. Plos Pathog 2012; 8: Involvement of the IkappaB kinase (IKK)- e1003085. related kinases tank-binding kinase 1/IKKi 6 Hofmann H, Norton TD, Schultz ML, Polsky SB, and cullin-based ubiquitin ligases in IFN CONFLICT OF INTEREST Sunseri N, Landau NR. Inhibition of CUL4A regulatory factor-3 degradation. JImmunol The authors declare no conflict of interest. Neddylation causes a reversible block to 2006; 177:5059–5067. Cellular & Molecular Immunology Neddylation in viral infection K Han and J Zhang 294 11 Ding S, Mooney N, Li B, Kelly MR, Feng N, 13 Collins SE, Noyce RS, Mossman KL. Innate 15 Zhang T, Ye Z, Yang X, Qin Y, Hu Y, Tong X Loktev AV et al. Comparative proteomics cellular response to virus particle entry et al. NEDDylation of PB2 reduces its reveals strain-specific beta-TrCP degradation requires IRF3 but not virus replication. stability and blocks the replication of via rotavirus NSP1 hijacking a host Cullin-3- J Virol 2004; 78:1706–1717. influenza A virus. Sci Rep 2017; 7: Rbx1 complex. Plos Pathog 2016; 12: 14 Zhang X, Ye Z, Pei Y, Qiu G, Wang Q, 43691. e1005929. Xu Y et al. Neddylation is required for 16 Song H, Huai W, Yu Z, Wang W, Zhao J, 12 Lutz LM, Pace CR, Arnold MM. Rotavirus herpes simplex virus type I (HSV-1)- Zhang L et al. MLN4924, a first-in-class NSP1 associates with components of the induced early phase interferon-beta pro- NEDD8-activating enzyme inhibitor, attenu- Cullin RING ligase family of E3 ubiquitin duction. Cell Mol Immunol 2016; 13: ates IFN-beta production. JImmunol2016; ligases. JVirol2016; 90:6036–6048. 578–583. 196:3117–3123. Cellular & Molecular Immunology.
Load more

Recommended publications
  • The Aryl Hydrocarbon Receptor: Structural Analysis and Activation Mechanisms
    The Aryl Hydrocarbon Receptor: Structural Analysis and Activation Mechanisms This thesis is submitted in fulfilment of the requirements for the degree of Doctor of Philosophy in the School of Molecular and Biomedical Sciences (Biochemistry), The University of Adelaide, Australia Fiona Whelan, B.Sc. (Hons) 2009 2 Table of Contents THESIS SUMMARY................................................................................. 6 DECLARATION....................................................................................... 7 PUBLICATIONS ARISING FROM THIS THESIS.................................... 8 ACKNOWLEDGEMENTS...................................................................... 10 ABBREVIATIONS ................................................................................. 12 CHAPTER 1: INTRODUCTION ............................................................. 17 1.1 BHLH.PAS PROTEINS ............................................................................................17 1.1.1 General background..................................................................................17 1.1.2 bHLH.PAS Class I Proteins.........................................................................18 1.2 THE ARYL HYDROCARBON RECEPTOR......................................................................19 1.2.1 Domain Structure and Ligand Activation ..............................................19 1.2.2 AhR Expression and Developmental Activity .......................................21 1.2.3 Mouse AhR Knockout Phenotype ...........................................................23
    [Show full text]
  • THE ROLE of the HERPES SIMPLEX VIRUS TYPE 1 UL28 PROTEIN in TERMINASE COMPLEX ASSEMBLY and FUNCTION by Jason Don Heming Bachelor
    THE ROLE OF THE HERPES SIMPLEX VIRUS TYPE 1 UL28 PROTEIN IN TERMINASE COMPLEX ASSEMBLY AND FUNCTION by Jason Don Heming Bachelor of Science, Clarion University of Pennsylvania, 2004 Submitted to the Graduate Faculty of the School of Medicine in partial fulfillment of the requirements for the degree of Doctor of Philosophy University of Pittsburgh 2013 UNIVERSITY OF PITTSBURGH SCHOOL OF MEDICINE This dissertation was presented by Jason Don Heming It was defended on April 18, 2013 and approved by Michael Cascio, Associate Professor, Bayer School of Natural and Environmental Sciences James Conway, Associate Professor, Department of Structural Biology Neal DeLuca, Professor, Department of Microbiology and Molecular Genetics Saleem Khan, Professor, Department of Microbiology and Molecular Genetics Dissertation Advisor: Fred Homa, Associate Professor, Department of Microbiology and Molecular Genetics ii Copyright © by Jason Don Heming 2013 iii THE ROLE OF THE HERPES SIMPLEX VIRUS TYPE 1 UL28 PROTEIN IN TERMINASE COMPLEX ASSEMBLY AND FUNCTION Jason Don Heming, PhD University of Pittsburgh, 2013 Herpes simplex virus type I (HSV-1) is the causative agent of several pathologies ranging in severity from the common cold sore to life-threatening encephalitic infection. During productive lytic infection, over 80 viral proteins are expressed in a highly regulated manner, resulting in the replication of viral genomes and assembly of progeny virions. Cleavage and packaging of replicated, concatemeric viral DNA into newly assembled capsids is critical to virus proliferation and requires seven viral genes: UL6, UL15, UL17, UL25, UL28, UL32, and UL33. Analogy with the well-characterized cleavage and packaging systems of double-stranded DNA bacteriophage suggests that HSV-1 encodes for a viral terminase complex to perform these essential functions, and several studies have indicated that this complex consists of the viral UL15, UL28, and UL33 proteins.
    [Show full text]
  • S41598-018-28214-2.Pdf
    www.nature.com/scientificreports OPEN Dissecting Distinct Roles of NEDDylation E1 Ligase Heterodimer APPBP1 and UBA3 Received: 7 November 2017 Accepted: 25 May 2018 Reveals Potential Evolution Process Published: xx xx xxxx for Activation of Ubiquitin-related Pathways Harbani Kaur Malik-Chaudhry1, Zied Gaieb1, Amanda Saavedra1, Michael Reyes1, Raphael Kung1, Frank Le1, Dimitrios Morikis1,2 & Jiayu Liao1,2 Despite the similar enzyme cascade in the Ubiquitin and Ubiquitin-like peptide(Ubl) conjugation, the involvement of single or heterodimer E1 activating enzyme has been a mystery. Here, by using a quantitative Förster Resonance Energy Transfer (FRET) technology, aided with Analysis of Electrostatic Similarities Of Proteins (AESOP) computational framework, we elucidate in detail the functional properties of each subunit of the E1 heterodimer activating-enzyme for NEDD8, UBA3 and APPBP1. In contrast to SUMO activation, which requires both subunits of its E1 heterodimer AOS1-Uba2 for its activation, NEDD8 activation requires only one of two E1 subunits, UBA3. The other subunit, APPBP1, only contributes by accelerating the activation reaction rate. This discovery implies that APPBP1 functions mainly as a scafold protein to enhance molecular interactions and facilitate catalytic reaction. These fndings for the frst time reveal critical new mechanisms and a potential evolutionary pathway for Ubl activations. Furthermore, this quantitative FRET approach can be used for other general biochemical pathway analysis in a dynamic mode. Ubiquitin and Ubls are peptides that are conjugated to various target proteins to either lead the targeted protein to degradation or changes of activities in vivo, and their dysregulations ofen leads to various diseases, such as can- cers or neurodegenerative diseases1–3.
    [Show full text]
  • The Anti-Tumor Activity of the NEDD8 Inhibitor Pevonedistat in Neuroblastoma
    International Journal of Molecular Sciences Article The Anti-Tumor Activity of the NEDD8 Inhibitor Pevonedistat in Neuroblastoma Jennifer H. Foster 1,* , Eveline Barbieri 1, Linna Zhang 1, Kathleen A. Scorsone 1, Myrthala Moreno-Smith 1, Peter Zage 2,3 and Terzah M. Horton 1,* 1 Texas Children’s Cancer and Hematology Centers, Department of Pediatrics, Section of Hematology and Oncology, Baylor College of Medicine, Houston, TX 77030, USA; [email protected] (E.B.); [email protected] (L.Z.); [email protected] (K.A.S.); [email protected] (M.M.-S.) 2 Department of Pediatrics, Division of Hematology-Oncology, University of California San Diego, La Jolla, CA 92024, USA; [email protected] 3 Peckham Center for Cancer and Blood Disorders, Rady Children’s Hospital, San Diego, CA 92123, USA * Correspondence: [email protected] (J.H.F.); [email protected] (T.M.H.); Tel.: +1-832-824-4646 (J.H.F.); +1-832-824-4269 (T.M.H.) Abstract: Pevonedistat is a neddylation inhibitor that blocks proteasomal degradation of cullin–RING ligase (CRL) proteins involved in the degradation of short-lived regulatory proteins, including those involved with cell-cycle regulation. We determined the sensitivity and mechanism of action of pevonedistat cytotoxicity in neuroblastoma. Pevonedistat cytotoxicity was assessed using cell viability assays and apoptosis. We examined mechanisms of action using flow cytometry, bromod- eoxyuridine (BrDU) and immunoblots. Orthotopic mouse xenografts of human neuroblastoma were generated to assess in vivo anti-tumor activity. Neuroblastoma cell lines were very sensitive to pevonedistat (IC50 136–400 nM). The mechanism of pevonedistat cytotoxicity depended on p53 Citation: Foster, J.H.; Barbieri, E.; status.
    [Show full text]
  • HIV Accessory Proteins: Review Leading Roles for the Supporting Cast
    View metadata, citation and similar papers at core.ac.uk brought to you by CORE provided by Elsevier - Publisher Connector Cell, Vol. 82, 189-192, July 28, 1995, Copyright © 1995 by Cell Press HIV Accessory Proteins: Review Leading Roles for the Supporting Cast Didier Trono ment into coated pits. The lymphoid-specific p56~k tyrosine Infectious Disease Laboratory kinase bound to the CD4 cytoplasmic tail plays such an The Salk Institute for Biological Studies inhibitory role. While Lck is released by Nef, this seems 10010 North Torrey Pines Road to be an epiphenomenon, because Nef-induced CD4 La Jolla, California 92037 down-modulation is observed in cells that do not express Lck and on truncated CD4 molecules that do not bind the kinase. In yet another model that would be unprecedented, Considering that a set of three genes is sufficient for the Nef could physically connect CD4 with a component of replication of most retroviruses, the human and simian the internalization pathway. Future experiments should immunodeficiency viruses (HIV and SIV) exhibit a surpris- test these various possibilities, in particular by determining ing degree of genomic complexity. HIV-1 contains no less conclusively whether Nef, directly or indirectly, interacts than six open reading frames in addition to the prototypic with CD4 or with the endocytic apparatus. Irrespective of gag, pol, and env coding sequences (see Figure 1). Two its mechanism, what' advantage does Nef-mediated CD4 of these supplementary genes, tat and rev, are absolutely down-regulation confer to the virus? Does it rapidly block necessary for virus growth: Tat is a major transactivator detrimental superinfection events? Does it prevent virus of the proviral long terminal repeat (LTR), and Rev acts aggregation on the surface of cells during budding, analo- posttranscriptionally to ensure the switch from the early gous to the role played by neuraminidase for influenza to the late phase of viral gene expression.
    [Show full text]
  • Chlamydia Trachomatis-Containing Vacuole Serves As Deubiquitination
    RESEARCH ARTICLE Chlamydia trachomatis-containing vacuole serves as deubiquitination platform to stabilize Mcl-1 and to interfere with host defense Annette Fischer1, Kelly S Harrison2, Yesid Ramirez3, Daniela Auer1, Suvagata Roy Chowdhury1, Bhupesh K Prusty1, Florian Sauer3, Zoe Dimond2, Caroline Kisker3, P Scott Hefty2, Thomas Rudel1* 1Department of Microbiology, Biocenter, University of Wu¨ rzburg, Wu¨ rzburg, Germany; 2Department of Molecular Biosciences, University of Kansas, lawrence, United States; 3Rudolf Virchow Center for Experimental Biomedicine, University of Wu¨ rzburg, Wu¨ rzburg, Germany Abstract Obligate intracellular Chlamydia trachomatis replicate in a membrane-bound vacuole called inclusion, which serves as a signaling interface with the host cell. Here, we show that the chlamydial deubiquitinating enzyme (Cdu) 1 localizes in the inclusion membrane and faces the cytosol with the active deubiquitinating enzyme domain. The structure of this domain revealed high similarity to mammalian deubiquitinases with a unique a-helix close to the substrate-binding pocket. We identified the apoptosis regulator Mcl-1 as a target that interacts with Cdu1 and is stabilized by deubiquitination at the chlamydial inclusion. A chlamydial transposon insertion mutant in the Cdu1-encoding gene exhibited increased Mcl-1 and inclusion ubiquitination and reduced Mcl- 1 stabilization. Additionally, inactivation of Cdu1 led to increased sensitivity of C. trachomatis for IFNg and impaired infection in mice. Thus, the chlamydial inclusion serves as an enriched site for a *For correspondence: thomas. deubiquitinating activity exerting a function in selective stabilization of host proteins and [email protected]. protection from host defense. de DOI: 10.7554/eLife.21465.001 Competing interests: The authors declare that no competing interests exist.
    [Show full text]
  • The COP9 Signalosome Variant CSNCSN7A Stabilizes the Deubiquitylating Enzyme CYLD Impeding Hepatic Steatosis
    Article The COP9 Signalosome Variant CSNCSN7A Stabilizes the Deubiquitylating Enzyme CYLD Impeding Hepatic Steatosis Xiaohua Huang 1,* , Dawadschargal Dubiel 2 and Wolfgang Dubiel 2,3,*,† 1 Charité—Universitätsmedizin Berlin, Chirurgische Klinik, Campus Charité Mitte|Campus Virchow-Klinikum, Experimentelle Chirurgie und Regenerative Medizin, Augustenburger Platz 1, 13353 Berlin, Germany 2 Institute of Experimental Internal Medicine, Medical Faculty, Otto von Guericke University, Leipziger Str. 44, 39120 Magdeburg, Germany; [email protected] 3 State Key Laboratory of Cellular Stress Biology, Fujian Provincial Key Laboratory of Innovative Drug Target Research, School of Pharmaceutical Sciences, Xiamen University, Xiang’an South Road, Xiamen 361102, China * Correspondence: [email protected] (X.H.); [email protected] (W.D.) † Lead Contact. Abstract: Hepatic steatosis is a consequence of distorted lipid storage and plays a vital role in the pathogenesis of nonalcoholic fatty liver disease (NAFLD). This study aimed to explore the role of the COP9 signalosome (CSN) in the development of hepatic steatosis and its interplay with the deubiquitylating enzyme (DUB) cylindromatosis (CYLD). CSN occurs as CSNCSN7A and CSNCSN7B variants regulating the ubiquitin proteasome system. It is a deneddylating complex and associates with other DUBs. CYLD cleaves Lys63-ubiquitin chains, regulating a signal cascade that mitigates hepatic steatosis. CSN subunits CSN1 and CSN7B, as well as CYLD, were downregulated with specific siRNA in HepG2 cells and human primary hepatocytes. The same cells were transfected Citation: Huang, X.; Dubiel, D.; with Flag-CSN7A or Flag-CSN7B for pulldowns. Hepatic steatosis in cell culture was induced Dubiel, W. The COP9 Signalosome by palmitic acid (PA). Downregulation of CSN subunits led to reduced PPAR-γ expression.
    [Show full text]
  • NEDD8 Acts As a 'Molecular Switch' Defining the Functional Selectivity Of
    scientificscientificreport report NEDD8 acts as a ‘molecular switch’ defining the functional selectivity of VHL Ryan C. Russell & Michael Ohh+ Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, Ontario, Canada The von Hippel–Lindau (VHL) tumour suppressor protein is (HIF), which is the main transcription factor that activates the important in the E3 ubiquitin ligase ECV (Elongin B/C–CUL2– expression of many hypoxia-inducible genes to counter VHL)-mediated destruction of hypoxia-inducible factor and the the detrimental effects of compromised oxygen availability promotion of fibronectin (FN) extracellular matrix assembly. (Kaelin, 2002), and as a positive regulator of fibronectin (FN) Although the precise molecular mechanism controlling the extracellular matrix (ECM) assembly (Roberts & Ohh, 2008). selectivity of VHL function remains unknown, a failure in either ECV (Elongin B/C–CUL2–VHL) is an SCF (SKP1/CDC53 or process is associated with oncogenic progression. Here, we show CUL1/F-box protein)-like E3 ubiquitin ligase in which VHL acts as that VHL performs its FN-associated function independently of a substrate-conferring component that recruits the a-subunits of the ECV complex, highlighting the autonomy of these pathways. HIF. These have been modified with hydroxyl groups on Furthermore, we show that NEDD8, a ubiquitin-like molecule, conserved prolyl residues within the oxygen-dependent degrada- acts as a ‘molecular switch’ in which its covalent conjugation to tion domain (ODD) by a class of prolyl hydroxylases in an oxygen- VHL prohibits the engagement of the scaffold component CUL2 dependent manner (Kaelin, 2002). This mechanistic insight and, concomitantly, activates the association with FN.
    [Show full text]
  • The Impact of Herpes Therapy on Genital and Systemic Immunology
    The impact of herpes therapy on genital and systemic immunology by Tae Joon Yi A thesis submitted in conformity with the requirements for the degree of Doctor of Philosophy Department of Immunology University of Toronto © Copyright by Tae Joon Yi 2013 The impact of herpes therapy on genital and systemic immunology Tae Joon Yi Doctor of Philosophy Department of Immunology University of Toronto 2013 Abstract HIV infects more than 34 million people globally. Herpes simplex virus type 2 (HSV-2) has been associated with a 3-fold increase in the rate of HIV acquisition, which may be due to physical breaks in the mucosal barrier during symptomatic episodes and/or an increased number of HIV target cells being exposed in the genital tract. Although clinical trials have demonstrated that herpes suppression in HIV uninfected individuals had no impact on HIV incidence, acyclovir was associated with a decrease in plasma HIV viral load and delayed disease progression in HSV-2 co-infected individuals. In addition, many HIV infected individuals display persistent systemic immune activation, which correlates with disease progression, despite successful antiretroviral therapy (ART). It is hypothesized that HSV could be one of the drivers of this activation. To further assess these relationships, my thesis focused on examining the impact of herpes therapy on genital tract immunology and systemic immune activation in HIV uninfected women and a comparison of systemic immune activation in individuals co-infected with HIV and HSV-2 on ART randomized to valacyclovir or placebo. A randomized, placebo-controlled, double-blinded, cross-over trial was conducted in HSV-2 infected, HIV uninfected, women using valacyclovir.
    [Show full text]
  • Recombinant Human NEDD8 E1/APPBP1/UBA3 Protein Catalog Number: ATGP1425
    Recombinant human NEDD8 E1/APPBP1/UBA3 protein Catalog Number: ATGP1425 PRODUCT INPORMATION Expression system E.coli Domain 1-463aa UniProt No. Q8TBC4 NCBI Accession No. AAH22853 Alternative Names NEDD8-activating enzyme E1 catalytic subunit, uBE1C PRODUCT SPECIFICATION Molecular Weight 54.4 kDa (487aa) Concentration 1mg/ml (determined by Bradford assay) Formulation Liquid in. 20mM Tris-HCl buffer (pH 8.0) containing 20% glycerol, 1mM DTT Purity > 90% by SDS-PAGE Tag His-Tag Application SDS-PAGE Storage Condition Can be stored at +2C to +8C for 1 week. For long term storage, aliquot and store at -20C to -80C. Avoid repeated freezing and thawing cycles. BACKGROUND Description uBA3, also known as NEDD8-activating enzyme E1 catalytic subunit, is the catalytic subunit of the dimeric uBA3- NAE1 E1 enzyme. E1 activates NEDD8 by first adenylating its C-terminal glycine residue with ATP, thereafter linking this residue to the side chain of the catalytic cysteine, yielding a NEDD8-uBA3 thioester and free AMP. E1 finally transfers NEDD8 to the catalytic cysteine of uBE2M. Recombinant human uBA3 protein, fused to His-tag at N-terminus, was expressed in E. coli and purified by using conventional chromatography. 1 Recombinant human NEDD8 E1/APPBP1/UBA3 protein Catalog Number: ATGP1425 Amino acid Sequence MGSSHHHHHH SSGLVPRGSH MGSHMADGEE PERKRRRIEE LLAEKMAVDG GCGDTGDWEG RWNHVKKFLE RSGPFTHPDF EPSTESLQFL LDTCKVLVIG AGGLGCELLK NLALSGFRQI HVIDMDTIDV SNLNRQFLFR PKDIGRPKAE VAAEFLNDRV PNCNVVPHFN KIQDFNDTFY RQFHIIVCGL DSIIARRWIN GMLISLLNYE DGVLDPSSIV PLIDGGTEGF KGNARVILPG MTACIECTLE LYPPQVNFPM CTIASMPRLP EHCIEYVRML QWPKEQPFGE GVPLDGDDPE HIQWIFQKSL ERASQYNIRG VTYRLTQGVV KRIIPAVAST NAVIAAVCAT EVFKIATSAY IPLNNYLVFN DVDGLYTYTF EAERKENCPA CSQLPQNIQF SPSAKLQEVL DYLTNSASLQ MKSPAITATL EGKNRTLYLQ SVTSIEERTR PNLSKTLKEL GLVDGQELAV ADVTTPQTVL FKLHFTS General References Gong L., et al.
    [Show full text]
  • Chemotherapy Induces NEDP1-Mediated Destabilization of MDM2
    Oncogene (2010) 29, 297–304 & 2010 Macmillan Publishers Limited All rights reserved 0950-9232/10 $32.00 www.nature.com/onc SHORT COMMUNICATION Chemotherapy induces NEDP1-mediated destabilization of MDM2 IR Watson1,2,BKLi1,2, O Roche1, A Blanch2, M Ohh1 and MS Irwin1,2,3 1Department of Laboratory Medicine and Pathobiology, University of Toronto, Toronto, Ontario, Canada; 2Cell Biology Program, Hospital for Sick Children, Toronto, Ontario, Canada and 3Department of Paediatrics and Institute of Medical Science, University of Toronto, Toronto, Ontario, Canada MDM2 is an E3 ligase that promotes ubiquitin-mediated In response to DNA damage, p53 becomes phos- destruction of p53. Cellular stresses such as DNA damage phorylated by several kinases within the MDM2- can lead to p53 activation due in part to MDM2 binding domain, which prevents MDM2–p53 interac- destabilization. Here, we show that the stability of tion (Bode and Dong, 2004). The stabilization of p53 MDM2 is regulated by an ubiquitin-like NEDD8 pathway then leads to DNA repair, cell cycle arrest, senescence or and identify NEDP1 as a chemotherapy-induced isopepti- apoptosis. Recent studies have shown that MDM2 is dase that deneddylates MDM2, resulting in MDM2 destabilized in response to DNA damage, which promotes destabilization concomitant with p53 activation. Concor- p53 activation (Stommel and Wahl, 2004; Meulmeester dantly, RNAi-mediated knockdown of endogenous et al., 2005). NEDD8 is a ubiquitin-like protein that NEDP1 blocked diminution of MDM2 levels and regulates protein function through covalent modification increased chemoresistance of tumor cells. These findings of substrates such as Cullins, BCA3, EGFR, ribosomal unveil the regulation of MDM2 stability through NEDP1 L11 protein, VHL, p73 and p53 (Xirodimas, 2008).
    [Show full text]
  • E3 Ubiquitin Ligase Cullin-5 Modulates Multiple Molecular and Cellular Responses to Heat Shock Protein 90 Inhibition in Human Cancer Cells
    E3 ubiquitin ligase Cullin-5 modulates multiple molecular and cellular responses to heat shock protein 90 inhibition in human cancer cells Rahul S. Samant, Paul A. Clarke, and Paul Workman1 Cancer Research UK Cancer Therapeutics Unit, The Institute of Cancer Research, London SM2 5NG, UK Edited by Melanie H. Cobb, University of Texas Southwestern Medical Center, Dallas, TX, and approved April 3, 2014 (received for review December 24, 2013) The molecular chaperone heat shock protein 90 (HSP90) is required Given the link between CUL5 and the HSP90 inhibitor- for the activity and stability of its client proteins. Pharmacologic induced degradation of ERBB2 (12), we have investigated the inhibition of HSP90 leads to the ubiquitin-mediated degradation of role of Cullin-RING ligases with respect to HSP90’s protein kinase clients, particularly activated or mutant oncogenic protein kinases. clients in human cancer cell lines. Our initial focused siRNA Client ubiquitination occurs via the action of one or more E3 screen of 28 Cullin-RING ligase family members identified five ubiquitin ligases. We sought to identify the role of Cullin-RING fam- genes, including CUL5, that were required for ERBB2 degra- ily E3 ubiquitin ligases in the cellular response to HSP90 inhibition. dation following treatment with 17-AAG—which we use here as Through a focused siRNA screen of 28 Cullin-RING ligase family a representative HSP90 inhibitor and chemical tool to promote members, we found that CUL5 and RBX2 were required for degra- client protein turnover. We go on to show for the first time to our dation of several HSP90 clients upon treatment of human cancer knowledge that RNAi silencing of CUL5 reduces the 17-AAG– cells with the clinical HSP90 inhibitor 17-AAG.
    [Show full text]