| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
|
|
|
|||||||
|
|||||||||
J. Biol. Chem., Vol. 280, Issue 43, 36417-36428, October 28, 2005
| ||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||||
1
From the
Department of Biochemistry and Molecular Biology, Pennsylvania State University, University Park, Pennsylvania 16802 and the Department of Biochemistry and Molecular Biology, University of Arkansas for Medical Sciences, Little Rock, Arkansas 72205
Hepatitis C virus (HCV) nonstructural protein 5A (NS5A) has been shown to antagonize numerous cellular pathways, including the antiviral interferon-
response. However, the capacity of this protein to interact with the viral polymerase suggests a more direct role for NS5A in genome replication. In this study, we employed two bacterially expressed, soluble derivatives of NS5A to probe for novel functions of this protein. We find that NS5A has the capacity to bind to the 3'-ends of HCV plus and minus strand RNAs. The high affinity binding site for NS5A in the 3'-end of plus strand RNA maps to the polypyrimidine tract, an element known to be essential for genome replication and infectivity. NS5A has a preference for single-stranded RNA containing stretches of uridine or guanosine. Values for the equilibrium dissociation constants for high affinity binding sites were in the 10 nM range. Two-dimensional gel electrophoresis followed by Western blotting revealed the presence of unphosphorylated NS5A in Huh-7 cells stably expressing the subgenomic replicon. Moreover, RNA immunoprecipitation and NS5A pull-down experiments showed the capacity of replicon-derived NS5A to bind to synthetic RNA and the HCV genome, respectively. Deletion of all of the casein kinase II phosphorylation sites in NS5A supported stable replication of a subgenomic replicon in Huh-7. However, this derivative could not be labeled with inorganic phosphate, suggesting that extensive phosphorylation of NS5A is not required for the replication functions of NS5A. The discovery that NS5A is an RNA-binding protein defines a new functional target for development of agents to treat HCV infection and a new structural class of RNA-binding proteins.
Received for publication, July 26, 2005 , and in revised form, August 25, 2005.
* This work was supported in part by NIAID Grant AI66919 from the National Institutes of Health (to K. D. R. and C. E. C.). The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
1 To whom correspondence should be addressed: Dept. of Biochemistry and Molecular Biology, Pennsylvania State University, 201 Althouse Laboratory, University Park, PA 16802. Tel.: 814-876-8705; Fax: 814-865-7927; E-mail: cec9{at}psu.edu.
CiteULike 
Complore 
Connotea 
Del.icio.us 
Digg 
Reddit 
Technorati What's this?
This article has been cited by other articles:
|
|
 |
|
  T. Masaki, R. Suzuki, K. Murakami, H. Aizaki, K. Ishii, A. Murayama, T. Date, Y. Matsuura, T. Miyamura, T. Wakita, et al. Interaction of Hepatitis C Virus Nonstructural Protein 5A with Core Protein Is Critical for the Production of Infectious Virus Particles J. Virol., August 15, 2008; 82(16): 7964 - 7976. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
P. Targett-Adams, S. Boulant, and J. McLauchlan Visualization of Double-Stranded RNA in Cells Supporting Hepatitis C Virus RNA Replication J. Virol., March 1, 2008; 82(5): 2182 - 2195. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 G. Cheng, A. Montero, P. Gastaminza, C. Whitten-Bauer, S. F. Wieland, M. Isogawa, B. Fredericksen, S. Selvarajah, P. A. Gallay, M. R. Ghadiri, et al. A virocidal amphipathic {alpha}-helical peptide that inhibits hepatitis C virus infection in vitro PNAS, February 26, 2008; 105(8): 3088 - 3093. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. L. Tellinghuisen, K. L. Foss, J. C. Treadaway, and C. M. Rice Identification of Residues Required for RNA Replication in Domains II and III of the Hepatitis C Virus NS5A Protein J. Virol., February 1, 2008; 82(3): 1073 - 1083. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
S. You and C. M. Rice 3' RNA Elements in Hepatitis C Virus Replication: Kissing Partners and Long Poly(U) J. Virol., January 1, 2008; 82(1): 184 - 195. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. L. Tellinghuisen, M. J. Evans, T. von Hahn, S. You, and C. M. Rice Studying Hepatitis C Virus: Making the Best of a Bad Virus J. Virol., September 1, 2007; 81(17): 8853 - 8867. [Full Text] [PDF]
|
|
|
|
 |
|
 H. B. Pathak, J. J. Arnold, P. N. Wiegand, M. R. S. Hargittai, and C. E. Cameron Picornavirus Genome Replication: ASSEMBLY AND ORGANIZATION OF THE VPg URIDYLYLATION RIBONUCLEOPROTEIN (INITIATION) COMPLEX J. Biol. Chem., June 1, 2007; 282(22): 16202 - 16213. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
V. S. Korneeva and C. E. Cameron Structure-Function Relationships of the Viral RNA-dependent RNA Polymerase: FIDELITY, REPLICATION SPEED, AND INITIATION MECHANISM DETERMINED BY A RESIDUE IN THE RIBOSE-BINDING POCKET J. Biol. Chem., June 1, 2007; 282(22): 16135 - 16145. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Binder, D. Quinkert, O. Bochkarova, R. Klein, N. Kezmic, R. Bartenschlager, and V. Lohmann Identification of Determinants Involved in Initiation of Hepatitis C Virus RNA Synthesis by Using Intergenotypic Replicase Chimeras J. Virol., May 15, 2007; 81(10): 5270 - 5283. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
T. Schaller, N. Appel, G. Koutsoudakis, S. Kallis, V. Lohmann, T. Pietschmann, and R. Bartenschlager Analysis of Hepatitis C Virus Superinfection Exclusion by Using Novel Fluorochrome Gene-Tagged Viral Genomes J. Virol., May 1, 2007; 81(9): 4591 - 4603. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
V. Brass, Z. Pal, N. Sapay, G. Deleage, H. E. Blum, F. Penin, and D. Moradpour Conserved Determinants for Membrane Association of Nonstructural Protein 5A from Hepatitis C Virus and Related Viruses J. Virol., March 15, 2007; 81(6): 2745 - 2757. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Quintavalle, S. Sambucini, V. Summa, L. Orsatti, F. Talamo, R. De Francesco, and P. Neddermann Hepatitis C Virus NS5A Is a Direct Substrate of Casein Kinase I-{alpha}, a Cellular Kinase Identified by Inhibitor Affinity Chromatography Using Specific NS5A Hyperphosphorylation Inhibitors J. Biol. Chem., February 23, 2007; 282(8): 5536 - 5544. [Abstract] [Full Text] [PDF]
|
|
|
|
 |
|
 D. M. Jones, S. N. Gretton, J. McLauchlan, and P. Targett-Adams Mobility analysis of an NS5A-GFP fusion protein in cells actively replicating hepatitis C virus subgenomic RNA J. Gen. Virol., February 1, 2007; 88(2): 470 - 475. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
Y. Nakatsu, M. Takeda, S. Ohno, R. Koga, and Y. Yanagi Translational Inhibition and Increased Interferon Induction in Cells Infected with C Protein-Deficient Measles Virus J. Virol., December 1, 2006; 80(23): 11861 - 11867. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
M. Quintavalle, S. Sambucini, C. Di Pietro, R. De Francesco, and P. Neddermann The {alpha} Isoform of Protein Kinase CKI Is Responsible for Hepatitis C Virus NS5A Hyperphosphorylation J. Virol., November 15, 2006; 80(22): 11305 - 11312. [Abstract] [Full Text] [PDF]
|
|
|
|
|
|
N. Appel, T. Schaller, F. Penin, and R. Bartenschlager From Structure to Function: New Insights into Hepatitis C Virus RNA Replication J. Biol. Chem., April 14, 2006; 281(15): 9833 - 9836. [Full Text] [PDF]
|
|
| HOME | HELP | FEEDBACK | SUBSCRIPTIONS | ARCHIVE | SEARCH | TABLE OF CONTENTS |
| All ASBMB Journals | Molecular and Cellular Proteomics |
| Journal of Lipid Research | ASBMB Today |
