Noroviruses subvert the core stress granule component G3BP1 to promote viral VPg-dependent translation

Myra Hosmillo; Jia Lu; Michael R. McAllaster; James B. Eaglesham; Xinjie Wang; Edward Emmott; Patricia Domingues; Yasmin Chaudhry; Timothy J Fitzmaurice; Matthew K.H. Tung; Marc Panas; Gerald McInerney; Nicholas Locker; Craig B. Willen; Ian Goodfellow

previously with a number of modifications that included the use 775 of the Brie gRNA library to reduce off target effects and shorter 776 infection times to improve the recovery of gRNAs that may also compromise cell 777 viability. BV-2 cells stably expressing Cas9 nuclease were 778 transduced with the Brie library using previously described protocols (Doench et al., 779 2016) . MNV strains CW3 and CR6 were used to infect BV-2 CRISPR library at MOI 780 5 pfu/cell and cells were isolated 24 hours post infection and preparation of gDNA 781 for sequencing as described previously . The screen relies on 782 the premise that guide RNAs targeting genes that are overrepresented following 783 infection represent genes that when disrupted are protected against infection and 784 therefore likely represent factors with pro-viral activity. Likewise, genes for which 785 guide RNA are underrepresented suggest that infection had proceeded faster and 786 the gene is antiviral. Following sequencing, the data was analyzed by STARS 787 method as previously described Orchard et al., 2016) . The copyright holder for this preprint (which was not peer-reviewed) is the author/funder. It . https://doi.org/10.1101/571455 doi: bioRxiv preprint 35 1X penicillin/streptomycin. SILAC media were supplemented with Light (R0K0), 796