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Short Communication |
1 International Centre for Genetic Engineering and Biotechnology, Padriciano 99, 34012 Trieste, Italy
2 Lay Line Genomics SpA, via di Castel Romano 100, 00128 Roma, Italy
3 Scuola Internazionale Superiore di Studi Avanzati, Ed. B, Padriciano 99, 34012 Trieste, Italy
Correspondence
Oscar R. Burrone
burrone{at}icgeb.org
Intracellular antibodies or intrabodies (ICAbs) have great potential in protein knockout strategies for intracellular antigens. In this study, they have been used to investigate the role of the rotavirus non-structural protein NSP5 in the virus replication cycle. Intracellular antibody-capture technology was used to select single-chain Fv format (scFv) ICAbs against an NSP5 mutant. Five different specific ICAbs were selected and expressed in MA104 cells, in the scFv format, as cytoplasmic- and nuclear-tagged forms. By confocal microscopy, it was found that three of these ICAbs recognized the full-length wild-type NSP5 specifically, forming antigen-specific aggresomes in the cytoplasm of cotransfected cells. Expression of the ICAbs in rotavirus-infected cells largely reduced the assembly of viroplasms and cellular cytopathic effect. Replication of dsRNA was partially inhibited, despite there being no reduction in virus titre. These results demonstrate for the first time a key role for NSP5 during the virus replicative cycle.
Present address: Institute Curie, INSERM U520, 12 Rue Lhomond, 75005 Paris, France.
This article has been cited by other articles:
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  A. Sen, N. Sen, and E. R. Mackow The Formation of Viroplasm-Like Structures by the Rotavirus NSP5 Protein Is Calcium Regulated and Directed by a C-Terminal Helical Domain J. Virol., November 1, 2007; 81(21): 11758 - 11767. [Abstract] [Full Text] [PDF]
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 M. Campagna, M. Budini, F. Arnoldi, U. Desselberger, J. E. Allende, and O. R. Burrone Impaired hyperphosphorylation of rotavirus NSP5 in cells depleted of casein kinase 1{alpha} is associated with the formation of viroplasms with altered morphology and a moderate decrease in virus replication J. Gen. Virol., October 1, 2007; 88(10): 2800 - 2810. [Abstract] [Full Text] [PDF]
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F. Arnoldi, M. Campagna, C. Eichwald, U. Desselberger, and O. R. Burrone Interaction of Rotavirus Polymerase VP1 with Nonstructural Protein NSP5 Is Stronger than That with NSP2 J. Virol., March 1, 2007; 81(5): 2128 - 2137. [Abstract] [Full Text] [PDF]
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A. Sen, D. Agresti, and E. R. Mackow Hyperphosphorylation of the Rotavirus NSP5 Protein Is Independent of Serine 67 or NSP2, and the Intrinsic Insolubility of NSP5 Is Regulated by Cellular Phosphatases J. Virol., February 15, 2006; 80(4): 1807 - 1816. [Abstract] [Full Text] [PDF]
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T. Lopez, M. Rojas, C. Ayala-Breton, S. Lopez, and C. F. Arias Reduced expression of the rotavirus NSP5 gene has a pleiotropic effect on virus replication J. Gen. Virol., June 1, 2005; 86(6): 1609 - 1617. [Abstract] [Full Text] [PDF]
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M. Campagna, C. Eichwald, F. Vascotto, and O. R. Burrone RNA interference of rotavirus segment 11 mRNA reveals the essential role of NSP5 in the virus replicative cycle J. Gen. Virol., May 1, 2005; 86(5): 1481 - 1487. [Abstract] [Full Text] [PDF]
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