Fig. 2.

Generation of focused sublibrary and validation of individual gene hits. (A) Validation of select genes from screen. Immunoblot analysis shows mTORC1 signaling is blunted, as detected by decreased phosphorylation of the direct mTORC1 substrate S6K1, in HEK293T cells expressing sgRNAs targeting indicated genes. Signaling was assayed 8 d posttransduction with the indicated sgRNA as described for the primary screen. (B) Categories of genes included in the focused sublibrary along with a schematic of how the focused screens were performed. (C) CS scores for the 24 top scoring genes from a focused sublibrary screen in wild-type HEK293T cells.