An Evolutionarily Conserved Pathway Essential for Orsay Virus Infection of .

Many fundamental biological discoveries have been made in The discovery of Orsay virus has enabled studies of host-virus interactions in this model organism. To identify host factors critical for Orsay virus infection, we designed a forward genetic screen that utilizes a virally induced green fluorescent protein (GFP) reporter. Following chemical mutagenesis, two Viro (irus nduced eporter ff) mutants that failed to express GFP were mapped to , a nonreceptor tyrosine kinase, and B0280.13 (renamed ), an ortholog of human Wiskott-Aldrich syndrome protein (WASP). Both mutants yielded Orsay virus RNA levels comparable to that of the residual input virus, suggesting that they are not permissive for Orsay virus replication. In addition, we demonstrated that both genes affect an early prereplication stage of Orsay virus infection. Furthermore, it is known that the human ortholog of SID-3, activated CDC42-associated kinase (ACK1/TNK2), is capable of phosphorylating human WASP, suggesting that VIRO-2 may be a substrate for SID-3 in A targeted RNA interference (RNAi) knockdown screen further identified the gene , which has a human ortholog that interacts with TNK2 and WASP, as required for Orsay virus infection. Thus, genetic screening in identified critical roles in virus infection for evolutionarily conserved genes in a known human pathway. Orsay virus is the only known virus capable of naturally infecting the model organism , which shares many evolutionarily conserved genes with humans. We exploited the robust genetic tractability of to identify three host genes, , , and , which are essential for Orsay virus infection. Mutant animals that lack these three genes are highly defective in viral replication. Strikingly, the human orthologs of these three genes, activated CDC42-associated kinase (TNK2), Wiskott-Aldrich syndrome protein (WASP), and noncatalytic region of tyrosine kinase adaptor protein 1 (NCK1) are part of a known signaling pathway in mammals. These results suggest that TNK2, WASP, and NCK1 may play important roles in mammalian virus infection.