RNA interference (RNAi) is a double-stranded RNA (dsRNA) guided mechanism that mediates sequence specific degradation of RNA. The recent demonstration that RNA interference can be used to inhibit virus replication has initiated an exciting field of research. First, it provides a novel antiviral therapeutic approach and, second, it may constitute a hitherto unrecognized natural antiviral defense mechanism. The long-term goal of this proposal is to gain a better understanding of the significance and therapeutic potential of RNA interference (RNAi) in the inhibition of viral infections. To this end we propose three specific aims to: (1) understand the determinants of siRNA-based viral inhibition;(2) investigate RNAi as a natural antiviral defense mechanism;and (3) identify and characterize novel components of the RNAi machinery. We will employ well-characterized positive stranded RNA viruses (picornaviruses,alphaviruses, and flaviviruses) as model viruses, and employ Drosophila and mammalian cells as model hosts. (1) To understand the determinants of siRNA-based viral inhibition, we will define the genome regions of RNA viruses that are susceptible to RNAi, and explore the efficiency of different siRNA design approaches. We will also investigate the ability of viruses to escape RNAi. (2) To investigate RNAi as a natural antiviral defense mechanism, we examine whether viral infection of animal cells produces siRNAs and analyze whether inhibiting components of the RNAi machinery enhances viral replication in mammalian cells. We will also determine whether viral infection induces changes in the RNAi machinery. (3) To identify and characterize novel components of the RNAi machinery, we carry out genome-wide screens in Drosophila cells. We will then employ cell biology and biochemistry to characterize the mechanism of entry and the role of the genes identified in our screens, and isolate the mammalian orthologs of RNAi entry components identified in our screens and explore their role in RNAi entry in mammalian systems.

National Institute of Health (NIH)
National Institute of Allergy and Infectious Diseases (NIAID)
Research Project (R01)
Project #
Application #
Study Section
Virology - A Study Section (VIRA)
Program Officer
Tseng, Christopher K
Project Start
Project End
Budget Start
Budget End
Support Year
Fiscal Year
Total Cost
Indirect Cost
University of California San Francisco
Schools of Medicine
San Francisco
United States
Zip Code
Nayak, Arabinda; Tassetto, Michel; Kunitomi, Mark et al. (2013) RNA interference-mediated intrinsic antiviral immunity in invertebrates. Curr Top Microbiol Immunol 371:183-200
Burrill, Cecily P; Strings, Vanessa R; Andino, Raul (2013) Poliovirus: generation, quantification, propagation, purification, and storage. Curr Protoc Microbiol Chapter 15:Unit 15H.1
Nayak, Arabinda; Andino, Raul (2011) Slicer activity in Drosophila melanogaster S2 extract. Methods Mol Biol 721:231-44
Vogt, Dorothee A; Andino, Raul (2010) An RNA element at the 5'-end of the poliovirus genome functions as a general promoter for RNA synthesis. PLoS Pathog 6:e1000936
Nayak, Arabinda; Berry, Bassam; Tassetto, Michel et al. (2010) Cricket paralysis virus antagonizes Argonaute 2 to modulate antiviral defense in Drosophila. Nat Struct Mol Biol 17:547-54
Saleh, Maria-Carla; Tassetto, Michel; van Rij, Ronald P et al. (2009) Antiviral immunity in Drosophila requires systemic RNA interference spread. Nature 458:346-50
Ylosmaki, Erkko; Hakkarainen, Tanja; Hemminki, Akseli et al. (2008) Generation of a conditionally replicating adenovirus based on targeted destruction of E1A mRNA by a cell type-specific MicroRNA. J Virol 82:11009-15
Barnes, Dwight; Kunitomi, Mark; Vignuzzi, Marco et al. (2008) Harnessing endogenous miRNAs to control virus tissue tropism as a strategy for developing attenuated virus vaccines. Cell Host Microbe 4:239-48
van Rij, Ronald P; Saleh, Maria-Carla; Berry, Bassam et al. (2006) The RNA silencing endonuclease Argonaute 2 mediates specific antiviral immunity in Drosophila melanogaster. Genes Dev 20:2985-95