![]() Indeed, we did not detect viral protein expression by immunofluorescence assays in Drosophila D-Mel2 cells inoculated with influenza virus A/WSN/33 (WSN H1N1) (data not shown). 1), we predicted that wild-type human influenza virus would not be able to infect them. Thus, in principle, Drosophila RNAi studies could accelerate identification of host interactions essential for influenza virus replication.īecause Drosophila D-Mel2 cells do not express the human influenza virus receptor α2,6-linked sialic acid ( Supplementary Fig. Because of its powerful genetics and conservation with vertebrates, Drosophila has been used to make numerous critical contributions to mammalian cell biology 6, 7, 8, 9. Such analysis is facilitated by well-developed model systems such as Drosophila, the genome of which contains only ∼14,000 genes, nearly all of which can be specifically targeted for high efficiency messenger RNA depletion by double-stranded RNA (dsRNA) libraries 1. Systematic, genome-wide RNAi analysis offers an exciting tool to identify host genes that function in viral replication. Although most such host molecules remain elusive, emerging results indicate that their identification and characterization can provide new insights into the mechanisms by which viruses complete their life cycle, and hence illuminate potentially valuable targets for prophylactic and therapeutic intervention 3, 4, 5. Many steps in the viral life cycle, including intracellular trafficking, gene expression, replication and virion assembly, depend on interactions with specific host cell gene products. To provide rational bases for improved treatment and control of influenza virus infection, we sought to advance understanding of viral infection mechanisms by elucidating previously unknown virus–host cell interactions. Since their first lethal infection of humans in 1997, H5N1 influenza A viruses have spread throughout Asia and to Europe and Africa, posing a major risk for a new influenza pandemic 2. Influenza outbreaks kill millions of people worldwide during pandemic years and hundreds of thousands during other years. Influenza, a highly contagious disease of birds and mammals, is caused by negative-strand RNA viruses of the family Orthomyxoviridae. This could accelerate the development of new classes of antiviral drugs for chemoprophylaxis and treatment, which are urgently needed given the obstacles to rapid development of an effective vaccine against pandemic influenza and the probable emergence of strains resistant to available drugs. ![]() Thus, we have demonstrated the feasibility of using genome-wide RNAi screens in Drosophila to identify previously unrecognized host proteins that are required for influenza virus replication. The relevance of these findings to influenza virus infection of mammalian cells is illustrated for a subset of the Drosophila genes identified that is, for three implicated Drosophila genes, the corresponding human homologues ATP6V0D1, COX6A1 and NXF1 are shown to have key functions in the replication of H5N1 and H1N1 influenza A viruses, but not vesicular stomatitis virus or vaccinia virus, in human HEK 293 cells. After modifying influenza virus to allow infection of Drosophila cells and detection of influenza virus gene expression, we tested an RNAi library against 13,071 genes (90% of the Drosophila genome), identifying over 100 for which suppression in Drosophila cells significantly inhibited or stimulated reporter gene ( Renilla luciferase) expression from an influenza-virus-derived vector. Here we describe a novel genome-wide RNA interference (RNAi) screen in Drosophila 1 that can be used to identify host genes important for influenza virus replication. Identifying the host molecules that participate in each step of virus replication could provide valuable new targets for antiviral therapy, but this goal may take several decades to achieve with conventional forward genetic screening methods and mammalian cell cultures. ![]() All viruses rely on host cell proteins and their associated mechanisms to complete the viral life cycle.
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