DOCSLIB.ORG

Prokaryotic Viperins Produce Diverse Antiviral Molecules

Total Page：16

File Type：pdf, Size：1020Kb

Download full-text PDF Read full-text
Prokaryotic Viperins Produce Diverse Antiviral Molecules Article Prokaryotic viperins produce diverse antiviral molecules https://doi.org/10.1038/s41586-020-2762-2 Aude Bernheim1, Adi Millman1, Gal Ofir1, Gilad Meitav1, Carmel Avraham1, Helena Shomar2, Masha M. Rosenberg2, Nir Tal2, Sarah Melamed1, Gil Amitai1 & Rotem Sorek1 ✉ Received: 25 February 2020 Accepted: 27 August 2020 Viperin is an interferon-induced cellular protein that is conserved in animals1. It Published online: xx xx xxxx has previously been shown to inhibit the replication of multiple viruses by producing Check for updates the ribonucleotide 3′-deoxy-3′,4′-didehydro (ddh)-cytidine triphosphate (ddhCTP), which acts as a chain terminator for viral RNA polymerase2. Here we show that eukaryotic viperin originated from a clade of bacterial and archaeal proteins that protect against phage infection. Prokaryotic viperins produce a set of modifed ribonucleotides that include ddhCTP, ddh-guanosine triphosphate (ddhGTP) and ddh-uridine triphosphate (ddhUTP). We further show that prokaryotic viperins protect against T7 phage infection by inhibiting viral polymerase-dependent transcription, suggesting that it has an antiviral mechanism of action similar to that of animal viperin. Our results reveal a class of potential natural antiviral compounds produced by bacterial immune systems. Viperin is an antiviral protein that becomes highly expressed in cells genes involved in defence systems is a strong predictor that the genes in stimulated by interferons1. In humans, this protein has broad antiviral the cluster have a role in phage resistance8,9. We denoted the genes in the activity against DNA and RNA viruses, including human cytomegalovi- defensive cluster pVips (for prokaryotic viperin homologues). As pVips rus, West Nile virus, dengue virus, hepatitis C virus and HIV1,3. Viperin are relatively rare in prokaryotic genomes (164 genes in the cluster), we was recently shown to catalyse the conversion of CTP to ddhCTP2. This performed an online homology search with additional genomes that modified nucleotide lacks a hydroxyl group at the 3′ carbon of the were not included in our original database, retrieving 86 additional ribose, and when the viral polymerase incorporates it into the nas- such genes, resulting in a total of 250 pVips (Supplementary Table 2). cent chain of the viral RNA, it acts as a chain terminator that does not To determine whether pVips can defend against phages, we selected allow further polymerization of the RNA chain2. Thus, ddhCTP directly 59 genes that span the space of the pVip sequence diversity (Supple- inhibits the RNA-dependent replication of RNA viruses such as Zika mentary Table 2) and cloned them in Escherichia coli under the con- virus in vivo. trol of an inducible promoter. Similarly, we also cloned GFP and the Some bacteria and archaea encode genes that have marked sequence MoaA gene from E. coli as negative controls. We then challenged the similarity to vertebrate viperins, although their roles have remained pVip-expressing bacteria with an array of phages spanning several unknown4–6. We set out to examine whether prokaryotic homologues major phage families (Myoviridae: P1; Siphoviridae: lambda, SECphi6, of human viperin participate in defence against phages. We first per- SECphi18 and SECphi27; Podoviridae: T7; Leviviridae: MS2 and Qβ) formed a profile-based search for viperin homologues in a database of (Fig. 1b, c, Extended Data Fig. 1). more than 38,000 bacterial and archaeal genomes. This search yielded Around half of the tested pVips conferred clearly identifiable activ- 1,724 genes (1,112 non-redundant sequences) homologous to the human ity against phages. Most of these protected against T7, as evidenced viperin gene (RSAD2), which we aggregated into 17 clusters on the basis by plaque assays (up to tenfold reduction in T7 plaque sizes) (Fig. 1b) of sequence similarity (Methods) (Supplementary Table 1). and by a delay or absence of culture collapse in T7 infection assays in Viperin is a member of the radical S-adenosyl-methionine (SAM) liquid culture (Fig. 1c, Extended Data Fig. 2a). Mutations in the cysteine family of enzymes, and shows both sequence and structural homology residues predicted to coordinate the iron–sulfur cluster in the Cxxx- to other members of that family, particularly the housekeeping molyb- CxxC motif in pVips resulted in loss of defensive capacity against T7, denum cofactor biosynthesis enzyme MoaA4. To differentiate between suggesting that the catalytic activity of pVips is necessary for defence viperin homologues with housekeeping properties and homologues (Extended Data Fig. 2b). A subset of the pVips also protected against that may participate in defence against phages, we took advantage of P1, lambda, SECphi6 and SECphi18 phages, reducing the observed the fact that, in prokaryotes, genes involved in antiviral activity tend number of plaques by between 10-fold and 10,000-fold (Extended to colocalize next to one another on the genome, forming ‘defence Data Fig. 1). Of note, when the human viperin gene was cloned and islands’7,8. Most clusters of viperin homologues did not show a tendency expressed in E. coli under the same conditions, it protected against T7 to colocalize with defence genes (Supplementary Table 1). However, infection in a similar manner to that observed for many pVips (Fig. 1, in one of the clusters, 60% of the genes were found in the vicinity of Extended Data Figs. 1, 2). CRISPR–cas systems, restriction-modification systems and other bacte- The pVips that we identified are present in phylogenetically dis- rial defence genes (Fig. 1a). This high propensity for colocalization with tant organisms, suggesting an ancient evolutionary origin, frequent 1Department of Molecular Genetics, Weizmann Institute of Science, Rehovot, Israel. 2Pantheon Biosciences, Yavne, Israel. ✉e-mail: [email protected] Nature | www.nature.com | 1 Article a Homologues of human viperin pVip2 Type I RM Type IV RM Nucleotide kinase TA TA Chryseobacterium gambrini DSM 18014 Ga0111674_101 10k 20k 30k pVip8 CRISPR–cas type III Psychrobacter lutiphocae DSM 21542 H140DRAFT_ scaffold00013.13 12k 22k 32k pVip68 Type I RM Nucleotide kinase Transposase Vibrio vulnificus YJ016 chromosome I: NC_005139 2206k 2196k pVip147 Type I RM TA Pseudomonas sp. 1-7 Ga0057553_134 153k 163k 173k pVip208 Nucleotide kinase Gabija Aliivibrio sp. 1S128 Ga0166917_1058 133k 143k 153k b 140 ******* *** * ** * * 120 100 80 60 40 T7 plaque size (mm) 20 0 GFP MoaA pVip6pVip7pVip8pVip9pVip1pVip120 pVip15pVip19pVip21pVip27pVip32pVip34pVip3pVip397 pVip42pVip4pVip464 pVip47pVip48pVip50pVip54pVip5pVip576 pVip58pVip6pVip610 pVip6pVip632 Human viperin c Human Viperin pVip9 pVip21 pVip34 Homo sapiens Vibrio porteresiae Lewinella persica Cryomorphaceae bacterium 0.8 0.6 0 0.4 OD60 0.2 0.0 0 100200 0 100200 0 100200 0 100200 Time post infection (min)Time post infection (min)Time post infection (min)Time post infection (min) Control uninfected Control infected Vip uninfected Vip infected Fig. 1 | pVips and human viperin exhibit antiviral activity in bacteria. viperin were grown on agar plates and phage lysate was added onto the plates. a, Representative instances of pVip genes and their genomic neighbourhoods. Graphs show mean of three replicates with individual data points overlaid. Homologues of human viperin are shown in red, genes annotated as nucleotide Asterisks indicate statistically significant differences compared with the GFP kinase are brown, genes known to be involved in defence are yellow and genes negative control. Two-sided t-test; *P < 0.0001 except for pVip46 (P = 0.0034)). with mobile genetic elements are dark grey. RM, restriction modification; TA, c, Growth curves for E. coli strains expressing viperins that were infected with toxin–antitoxin; Gabija is a recently described defence system8. The bacterial phage T7. Negative controls are GFP-expressing cells and are the same in all species, and the accession of the relevant genomic scaffold in the Integrated four graphs. Each growth curve represents the mean of 3 biological replicates, Microbial Genomes (IMG) database20, are indicated on the left. b, Sizes of each with an average of 2 technical replicates, and the shaded area corresponds plaques caused by T7 phage infecting E. coli strains that express different to the 95% confidence interval. viperins. Bacteria expressing pVips, negative controls (GFP or MoaA) or human horizontal gene transfer or both. We found pVips in 176 species overall, during the interferon response2,10. This kinase phosphorylates cyti- belonging to 14 bacterial and archaeal phyla that include Proteobacte- dine monophosphate to CTP, thus generating the substrate for viperin ria, Firmicutes, Cyanobacteria, Actinobacteria, Bacteriodetes, Euryar- activity2. We found that 47 of the 250 pVips (19%) genes were adjacent chaeota and others (Supplementary Table 2). To better understand their to genes annotated as a nucleotide kinase in their genome of origin diversity and phylogenetic distribution, we generated a phylogenetic (Fig. 1a, Fig. 2) and that, in some cases, the kinase was fused to the pVip tree of the viperin family, including pVips and eukaryotic viperins (Fig. 2, gene (Fig. 2, Supplementary Table 2). This further strengthens the Supplementary Table 3). MoaA genes from bacteria and eukaryotes hypothesis that the pVip substrate is a triphosphorylated nucleotide. were added to the tree as an outgroup. We found that pVips are grouped Whereas some pVip-associated kinases were annotated as cytidylate into seven major
Load more

Recommended publications
  • Polymerase Ribozyme with Promoter Recognition
    In vitro Evolution of a Processive Clamping RNA Polymerase Ribozyme with Promoter Recognition by Razvan Cojocaru BSc, Simon Fraser University, 2014 Thesis Submitted in Partial Fulfillment of the Requirements for the Degree of Doctor of Philosophy in the Department of Molecular Biology and Biochemistry Faculty of Science © Razvan Cojocaru 2021 SIMON FRASER UNIVERSITY Summer 2021 Copyright in this work is held by the author. Please ensure that any reproduction or re-use is done in accordance with the relevant national copyright legislation. Declaration of Committee Name: Razvan Cojocaru Degree: Doctor of Philosophy Title: In vitro Evolution of a Processive Clamping RNA Polymerase Ribozyme with Promoter Recognition Committee: Chair: Lisa Craig Professor, Molecular Biology and Biochemistry Peter Unrau Supervisor Professor, Molecular Biology and Biochemistry Dipankar Sen Committee Member Professor, Molecular Biology and Biochemistry Michel Leroux Committee Member Professor, Molecular Biology and Biochemistry Mani Larijani Internal Examiner Associate Professor, Molecular Biology and Biochemistry Gerald Joyce External Examiner Professor, Jack H. Skirball Center for Chemical Biology and Proteomics Salk Institute for Biological Studies Date Defended/Approved: August 12, 2021 ii Abstract The RNA World hypothesis proposes that the early evolution of life began with RNAs that can serve both as carriers of genetic information and as catalysts. Later in evolution, these functions were gradually replaced by DNA and enzymatic proteins in cellular biology. I start by reviewing the naturally occurring catalytic RNAs, ribozymes, as they play many important roles in biology today. These ribozymes are central to protein synthesis and the regulation of gene expression, creating a landscape that strongly supports an early RNA World.
    [Show full text]
  • Chikungunya Fever: Epidemiology, Clinical Syndrome, Pathogenesis
    Antiviral Research 99 (2013) 345–370 Contents lists available at SciVerse ScienceDirect Antiviral Research journal homepage: www.elsevier.com/locate/antiviral Review Chikungunya fever: Epidemiology, clinical syndrome, pathogenesis and therapy ⇑ Simon-Djamel Thiberville a,b, , Nanikaly Moyen a,b, Laurence Dupuis-Maguiraga c,d, Antoine Nougairede a,b, Ernest A. Gould a,b, Pierre Roques c,d, Xavier de Lamballerie a,b a UMR_D 190 ‘‘Emergence des Pathologies Virales’’ (Aix-Marseille Univ. IRD French Institute of Research for Development EHESP French School of Public Health), Marseille, France b University Hospital Institute for Infectious Disease and Tropical Medicine, Marseille, France c CEA, Division of Immuno-Virologie, Institute of Emerging Diseases and Innovative Therapies, Fontenay-aux-Roses, France d UMR E1, University Paris Sud 11, Orsay, France article info abstract Article history: Chikungunya virus (CHIKV) is the aetiological agent of the mosquito-borne disease chikungunya fever, a Received 7 April 2013 debilitating arthritic disease that, during the past 7 years, has caused immeasurable morbidity and some Revised 21 May 2013 mortality in humans, including newborn babies, following its emergence and dispersal out of Africa to the Accepted 18 June 2013 Indian Ocean islands and Asia. Since the first reports of its existence in Africa in the 1950s, more than Available online 28 June 2013 1500 scientific publications on the different aspects of the disease and its causative agent have been pro- duced. Analysis of these publications shows that, following a number of studies in the 1960s and 1970s, Keywords: and in the absence of autochthonous cases in developed countries, the interest of the scientific commu- Chikungunya virus nity remained low.
    [Show full text]
  • The Zinc-Finger Antiviral Protein Recruits the RNA Processing Exosome to Degrade the Target Mrna
    The zinc-finger antiviral protein recruits the RNA processing exosome to degrade the target mRNA Xuemin Guo, Jing Ma, Jing Sun, and Guangxia Gao* Institute of Biophysics, Chinese Academy of Sciences, Beijing 100101, China Edited by John M. Coffin, Tufts University School of Medicine, Boston, MA, and approved November 3, 2006 (received for review August 14, 2006) Zinc-finger antiviral protein (ZAP) is a host antiviral factor that putative RNA helicase (Kiaa0052), and a protein that is specifically specifically inhibits the replication of Moloney murine leukemia phosphorylated in the M phase of the cell cycle (Mpp6) (36, 38). virus (MLV) and Sindbis virus (SIN) by preventing accumulation of The RNase-PH domain subunits and the S1/KH RNA-binding the viral mRNA in the cytoplasm. In previous studies, we demon- domain subunits are considered to be the core components of the strated that ZAP directly binds to its specific target mRNAs. In this exosome, whereas Kiaa0052 and Mpp6 are considered to be article, we provide evidence indicating that ZAP recruits the RNA accessory factors (36, 38). Yeast PM/Scl-100 is found only in the processing exosome to degrade the target RNA. ZAP comigrated nuclear exosome (36, 46). with the exosome in sucrose or glycerol velocity gradient centrif- The structure of the exosome is not yet determined. Based on the ugation. Immunoprecipitation of ZAP coprecipitated the exosome results of mammalian-two-hybrid and yeast-two-hybrid experi- components. In vitro pull-down assays indicated that ZAP directly ments (38, 56–59), the six RNase-PH domain-containing subunits interacted with the exosome component hRrp46p and that the are thought to assemble into a doughnut-shaped ring.
    [Show full text]
  • Antibody-Mediated Enhancement Aggravates Chikungunya Virus
    www.nature.com/scientificreports OPEN Antibody-mediated enhancement aggravates chikungunya virus infection and disease severity Received: 14 July 2017 Fok-Moon Lum 1,2, Thérèse Couderc3,4, Bing-Shao Chia1,8, Ruo-Yan Ong1,9, Zhisheng Her1,10, Accepted: 17 January 2018 Angela Chow5, Yee-Sin Leo5, Yiu-Wing Kam1, Laurent Rénia1, Marc Lecuit 3,4,6 & Published: xx xx xxxx Lisa F. P. Ng1,2,7 The arthropod-transmitted chikungunya virus (CHIKV) causes a fu-like disease that is characterized by incapacitating arthralgia. The re-emergence of CHIKV and the continual risk of new epidemics have reignited research in CHIKV pathogenesis. Virus-specifc antibodies have been shown to control virus clearance, but antibodies present at sub-neutralizing concentrations can also augment virus infection that exacerbates disease severity. To explore this occurrence, CHIKV infection was investigated in the presence of CHIKV-specifc antibodies in both primary human cells and a murine macrophage cell line, RAW264.7. Enhanced attachment of CHIKV to the primary human monocytes and B cells was observed while increased viral replication was detected in RAW264.7 cells. Blocking of specifc Fc receptors (FcγRs) led to the abrogation of these observations. Furthermore, experimental infection in adult mice showed that animals had higher viral RNA loads and endured more severe joint infammation in the presence of sub-neutralizing concentrations of CHIKV-specifc antibodies. In addition, CHIKV infection in 11 days old mice under enhancing condition resulted in higher muscles viral RNA load detected and death. These observations provide the frst evidence of antibody-mediated enhancement in CHIKV infection and pathogenesis and could also be relevant for other important arboviruses such as Zika virus.
    [Show full text]
  • NSP4)-Induced Intrinsic Apoptosis
    viruses Article Viperin, an IFN-Stimulated Protein, Delays Rotavirus Release by Inhibiting Non-Structural Protein 4 (NSP4)-Induced Intrinsic Apoptosis Rakesh Sarkar †, Satabdi Nandi †, Mahadeb Lo, Animesh Gope and Mamta Chawla-Sarkar * Division of Virology, National Institute of Cholera and Enteric Diseases, P-33, C.I.T. Road Scheme-XM, Beliaghata, Kolkata 700010, India; [email protected] (R.S.); [email protected] (S.N.); [email protected] (M.L.); [email protected] (A.G.) * Correspondence: [email protected]; Tel.: +91-33-2353-7470; Fax: +91-33-2370-5066 † These authors contributed equally to this work. Abstract: Viral infections lead to expeditious activation of the host’s innate immune responses, most importantly the interferon (IFN) response, which manifests a network of interferon-stimulated genes (ISGs) that constrain escalating virus replication by fashioning an ill-disposed environment. Interestingly, most viruses, including rotavirus, have evolved numerous strategies to evade or subvert host immune responses to establish successful infection. Several studies have documented the induction of ISGs during rotavirus infection. In this study, we evaluated the induction and antiviral potential of viperin, an ISG, during rotavirus infection. We observed that rotavirus infection, in a stain independent manner, resulted in progressive upregulation of viperin at increasing time points post-infection. Knockdown of viperin had no significant consequence on the production of total Citation: Sarkar, R.; Nandi, S.; Lo, infectious virus particles. Interestingly, substantial escalation in progeny virus release was observed M.; Gope, A.; Chawla-Sarkar, M. upon viperin knockdown, suggesting the antagonistic role of viperin in rotavirus release. Subsequent Viperin, an IFN-Stimulated Protein, studies unveiled that RV-NSP4 triggered relocalization of viperin from the ER, the normal residence Delays Rotavirus Release by Inhibiting of viperin, to mitochondria during infection.
    [Show full text]
  • High Transdominant Revm10 Protein Levels Are Required to Inhibit HIV-1 Replication in Cell Lines and Primary T Cells: Implication for Gene Therapy of AIDS
    Gene Therapy (1997) 4, 128–139 1997 Stockton Press All rights reserved 0969-7128/97 $12.00 High transdominant RevM10 protein levels are required to inhibit HIV-1 replication in cell lines and primary T cells: implication for gene therapy of AIDS I Plavec1, M Agarwal1,KEHo2, M Pineda1, J Auten1, J Baker1, H Matsuzaki3, S Escaich4, M Bonyhadi1 and E Bo¨ hnlein1 1Progenesys Program, SyStemix Inc, 3155 Porter Drive, Palo Alto, CA 94304, USA Expression of antiviral genes in CD4+ T cells has been pro- uniformly higher than from internal promoters (eg CMV, posed as a strategy for gene therapy of AIDS. Over the PGK). Analysis of selected vectors in acutely and chron- past years, we and others have developed retroviral vec- ically HIV-infected cell lines suggested that threshold levels tors encoding the RevM10 protein, a dominant-negative of RevM10 expression are required to achieve inhibition of mutant of the HIV-1 Rev trans-activator protein. We could HIV replication. LTR-driven RevM10 expression also demonstrate gene transfer and inhibition of HIV-1 repli- yielded high steady-state protein levels in activated primary cation in cultured T cell lines and primary T cells. However, T cells resulting in inhibition of HIV replication, and there little is known about the levels of the antiviral protein was no apparent difference between the MoMLV, MPSV required to achieve a therapeutic effect, particularly in pri- and MESV-LTR vectors. However, RevM10 expression mary cells. In this report, we compare different vector was down-regulated in resting primary cells and conse- designs with regard to expression of the antiviral gene to quently anti-HIV efficacy was significantly reduced.
    [Show full text]
  • (12) Patent Application Publication (10) Pub. No.: US 2010/0317005 A1 Hardin Et Al
    US 20100317005A1 (19) United States (12) Patent Application Publication (10) Pub. No.: US 2010/0317005 A1 Hardin et al. (43) Pub. Date: Dec. 16, 2010 (54) MODIFIED NUCLEOTIDES AND METHODS (22) Filed: Mar. 15, 2010 FOR MAKING AND USE SAME Related U.S. Application Data (63) Continuation of application No. 11/007,794, filed on Dec. 8, 2004, now abandoned, which is a continuation (75) Inventors: Susan H. Hardin, College Station, in-part of application No. 09/901,782, filed on Jul. 9, TX (US); Hongyi Wang, Pearland, 2001. TX (US); Brent A. Mulder, (60) Provisional application No. 60/527,909, filed on Dec. Sugarland, TX (US); Nathan K. 8, 2003, provisional application No. 60/216,594, filed Agnew, Richmond, TX (US); on Jul. 7, 2000. Tommie L. Lincecum, JR., Publication Classification Houston, TX (US) (51) Int. Cl. CI2O I/68 (2006.01) Correspondence Address: (52) U.S. Cl. ............................................................ 435/6 LIFE TECHNOLOGES CORPORATION (57) ABSTRACT CFO INTELLEVATE Labeled nucleotide triphosphates are disclosed having a label P.O. BOX S2OSO bonded to the gamma phosphate of the nucleotide triphos MINNEAPOLIS, MN 55402 (US) phate. Methods for using the gamma phosphate labeled nucleotide are also disclosed where the gamma phosphate labeled nucleotide are used to attach the labeled gamma phos (73) Assignees: LIFE TECHNOLOGIES phate in a catalyzed (enzyme or man-made catalyst) reaction to a target biomolecule or to exchange a phosphate on a target CORPORATION, Carlsbad, CA biomolecule with a labeled gamme phosphate. Preferred tar (US); VISIGEN get biomolecules are DNAs, RNAs, DNA/RNAs, PNA, BIOTECHNOLOGIES, INC. polypeptide (e.g., proteins enzymes, protein, assemblages, etc.), Sugars and polysaccharides or mixed biomolecules hav ing two or more of DNAs, RNAs, DNA/RNAs, polypeptide, (21) Appl.
    [Show full text]
  • Characterization of Human UMP/CMP Kinase and Its Phosphorylation of D- and 1 L-Form Deoxycytidine Analogue Monophosphates
    [CANCER RESEARCH 62, 1624–1631, March 15, 2002] Characterization of Human UMP/CMP Kinase and Its Phosphorylation of D- and 1 L-Form Deoxycytidine Analogue Monophosphates Jieh-Yuan Liou, Ginger E. Dutschman, Wing Lam, Zaoli Jiang, and Yung-Chi Cheng2 Department of Pharmacology, Yale University School of Medicine, New Haven, Connecticut 06520 ABSTRACT with leukemia, lymphoma, or solid tumors (11). Deoxycytidine ana- logues, such as ␤-D-2Ј,3Ј-dideoxycytidine and L-(Ϫ)-SddC (Lamivu- Pyrimidine nucleoside monophosphate kinase [UMP/CMP kinase dine), have been shown to have anti-HIV and antihuman hepatitis B (UMP/CMPK); EC 2.7.4.14] plays a crucial role in the formation of UDP, virus activities (12–17). L-(Ϫ)-SddC was the first nucleoside analogue CDP, and dCDP, which are required for cellular nucleic acid synthesis. Several cytidine and deoxycytidine analogues are important anticancer with an L configuration to show therapeutic activity and, thus, defined ␤ Ј Ј and antiviral drugs. These drugs require stepwise phosphorylation to their a new category for the design of nucleoside analogues. -L-2 ,3 - triphosphate forms to exert their therapeutic effects. The role of UMP/ dideoxy-5-fluoro-3Ј-thia-cytidine and ␤-L-2Ј,3Ј-dideoxy-2Ј,3Ј-dide- CMPK for the phosphorylation of nucleoside analogues has been indi- hydro-5-fluorocytidine have been shown to be potent antihuman hep- cated. Thus, we cloned the human UMP/CMPK gene, expressed it in atitis B virus agents in vitro and in animal studies (18–22). In studies Escherichia coli, and purified it to homogeneity. Its kinetic properties of other ␤-L-(Ϫ)-2Ј,3Ј-dideoxycytidine analogues, it was observed were determined.
    [Show full text]
  • Identification and Characterisation of Human Cytomegalovirus-Mediated Degradation of Helicase-Like Transcription Factor
    Identification and Characterisation of Human Cytomegalovirus-Mediated Degradation of Helicase-Like Transcription Factor Kai-Min Lin Department of Medicine Cambridge Institute for Medical Research University of Cambridge This dissertation is submitted for the degree of Doctor of Philosophy Fitzwilliam College September 2020 Declaration I hereby declare, that except where specific reference is made to the work of others, the contents of this dissertation are original and have not been submitted in whole or in part for consideration for any other degree of qualification in this, or any other university. This dissertation is the result of my own work and includes nothing which is the outcome of work done in collaboration except as where specified in the text and acknowledgments. This dissertation does not exceed the specified word limit of 60,000 words as defined by the Degree Committee, excluding figures, photographs, tables, appendices and bibliography. Kai-Min Lin September, 2020 I Summary Identification and characterisation of human cytomegalovirus-mediated degradation of helicase-like transcription factor Kai-Min Lin Viruses are known to degrade host factors that are important in innate antiviral immunity in order to infect successfully. To systematically identify host proteins targeted for early degradation by human cytomegalovirus (HCMV), the lab developed orthogonal screens using high resolution multiplexed mass spectrometry. Taking advantage of broad and selective proteasome and lysosome inhibitors, proteasomal degradation was found to be heavily exploited by HCMV. Several known antiviral restriction factors, including components of cellular promyelocytic leukemia (PML) were enriched in a shortlist of proteasomally degraded proteins during infection. A particularly robust novel ‘hit’ was helicase-like transcription factor (HLTF), a DNA repair protein that participates in error-free repair of stalled replication forks.
    [Show full text]
  • Antivirals Against the Chikungunya Virus
    Preprints (www.preprints.org) | NOT PEER-REVIEWED | Posted: 10 June 2021 Review Antivirals against the Chikungunya Virus Verena Battisti 1, Ernst Urban 2 and Thierry Langer 3,* 1 University of Vienna, Department of Pharmaceutical Sciences, Pharmaceutical Chemistry Division, A-1090 Vienna, Austria; [email protected] 2 University of Vienna, Department of Pharmaceutical Sciences, Pharmaceutical Chemistry Division, A-1090 Vienna, Austria; [email protected] 3 University of Vienna, Department of Pharmaceutical Sciences, Pharmaceutical Chemistry Division, A-1090 Vienna, Austria; * Correspondence: [email protected] Abstract: Chikungunya virus (CHIKV) is a mosquito-transmitted alphavirus that has re-emerged in recent decades, causing large-scale epidemics in many parts of the world. CHIKV infection leads to a febrile disease known as chikungunya fever (CHIKF), which is characterised by severe joint pain and myalgia. As many patients develop a painful chronic stage and neither antiviral drugs nor vac- cines are available, the development of a potent CHIKV inhibiting drug is crucial for CHIKF treat- ment. A comprehensive summary of current antiviral research and development of small-molecule inhibitor against CHIKV is presented in this review. We highlight different approaches used for the identification of such compounds and further discuss the identification and application of promis- ing viral and host targets. Keywords: Chikungunya virus ; alphavirus; antiviral therapy; direct-acting antivirals; host-directed antivirals; in silico screening; in vivo validation, antiviral drug development 1. Introduction Chikungunya virus (CHIKV) is a mosquito-borne alphavirus and belongs to the Togaviridae family. The virus was first isolated from a febrile patient in 1952/53 in the Makonde plateau (Tanzania) and has been named after the Makonde word for “that which bends you up”, describing the characteristic posture of patients suffering severe joint pains due to the CHIKV infection [1].
    [Show full text]
  • The Microbiota-Produced N-Formyl Peptide Fmlf Promotes Obesity-Induced Glucose
    Page 1 of 230 Diabetes Title: The microbiota-produced N-formyl peptide fMLF promotes obesity-induced glucose intolerance Joshua Wollam1, Matthew Riopel1, Yong-Jiang Xu1,2, Andrew M. F. Johnson1, Jachelle M. Ofrecio1, Wei Ying1, Dalila El Ouarrat1, Luisa S. Chan3, Andrew W. Han3, Nadir A. Mahmood3, Caitlin N. Ryan3, Yun Sok Lee1, Jeramie D. Watrous1,2, Mahendra D. Chordia4, Dongfeng Pan4, Mohit Jain1,2, Jerrold M. Olefsky1 * Affiliations: 1 Division of Endocrinology & Metabolism, Department of Medicine, University of California, San Diego, La Jolla, California, USA. 2 Department of Pharmacology, University of California, San Diego, La Jolla, California, USA. 3 Second Genome, Inc., South San Francisco, California, USA. 4 Department of Radiology and Medical Imaging, University of Virginia, Charlottesville, VA, USA. * Correspondence to: 858-534-2230, [email protected] Word Count: 4749 Figures: 6 Supplemental Figures: 11 Supplemental Tables: 5 1 Diabetes Publish Ahead of Print, published online April 22, 2019 Diabetes Page 2 of 230 ABSTRACT The composition of the gastrointestinal (GI) microbiota and associated metabolites changes dramatically with diet and the development of obesity. Although many correlations have been described, specific mechanistic links between these changes and glucose homeostasis remain to be defined. Here we show that blood and intestinal levels of the microbiota-produced N-formyl peptide, formyl-methionyl-leucyl-phenylalanine (fMLF), are elevated in high fat diet (HFD)- induced obese mice. Genetic or pharmacological inhibition of the N-formyl peptide receptor Fpr1 leads to increased insulin levels and improved glucose tolerance, dependent upon glucagon- like peptide-1 (GLP-1). Obese Fpr1-knockout (Fpr1-KO) mice also display an altered microbiome, exemplifying the dynamic relationship between host metabolism and microbiota.
    [Show full text]
  • Lora Grainger Dissertation 6 23 10
    Characterization of the Transcripts that Encode pUL138, a Latency Determinant, During Human Cytomegalovirus Infection Item Type text; Electronic Dissertation Authors Grainger, Lora Ann Publisher The University of Arizona. Rights Copyright © is held by the author. Digital access to this material is made possible by the University Libraries, University of Arizona. Further transmission, reproduction or presentation (such as public display or performance) of protected items is prohibited except with permission of the author. Download date 24/09/2021 15:04:11 Link to Item http://hdl.handle.net/10150/195915 1 CHARACTERIZATION OF THE TRANSCRIPTS THAT ENCODE PUL138, A LATENCY DETERMINANT, DURING HUMAN CYTOMEGALOVIRUS INFECTION by Lora A. Grainger Copyright © Lora A. Grainger 2010 A Dissertation Submitted to the Faculty of the DEPARTMENT OF IMMUNOBIOLOGY In Partial Fulfillment of the Requirements for the Degree of DOCTOR OF PHILOSOPHY In the Graduate College THE UNIVERSITY OF ARIZONA 2010 2 THE UNIVERSITY OF ARIZONA GRADUATE COLLEGE As members of the Dissertation Committee, we certify that we have read the dissertation prepared by Lora A. Grainger entitled: Characterization of the Transcripts that Encode pUL138, a Latency Determinant, During Human Cytomegalovirus Infection. We recommend that it be accepted as fulfilling the dissertation requirement for the Degree of Doctor of Philosophy. _____________________________________________________Date: 6/15/10 Dr. Nafees Ahmad _____________________________________________________ Date: 6/15/10 Dr. Lonnie Lybarger _____________________________________________________ Date: 6/15/10 Dr. Carol Dieckmann Final approval and acceptance of this dissertation is contingent upon the candidate’s submission of the final copies of the dissertation to the Graduate College. I hereby certify that I have read this dissertation prepared under my direction and recommend that it be accepted as fulfilling the dissertation requirement _____________________________________________________Date: 6/15/10 Dissertation Director: Dr.
    [Show full text]