TY - JOUR TI - Pooled genome-wide CRISPR activation screening for rapamycin resistance genes in Drosophila cells AU - Xia, Baolong AU - Viswanatha, Raghuvir AU - Hu, Yanhui AU - Mohr, Stephanie E AU - Perrimon, Norbert A2 - VijayRaghavan, K A2 - Padinjat, Raghu VL - 12 PY - 2023 DA - 2023/04/20 SP - e85542 C1 - eLife 2023;12:e85542 DO - 10.7554/eLife.85542 UR - https://doi.org/10.7554/eLife.85542 AB - Loss-of-function and gain-of-function genetic perturbations provide valuable insights into gene function. In Drosophila cells, while genome-wide loss-of-function screens have been extensively used to reveal mechanisms of a variety of biological processes, approaches for performing genome-wide gain-of-function screens are still lacking. Here, we describe a pooled CRISPR activation (CRISPRa) screening platform in Drosophila cells and apply this method to both focused and genome-wide screens to identify rapamycin resistance genes. The screens identified three genes as novel rapamycin resistance genes: a member of the SLC16 family of monocarboxylate transporters (CG8468), a member of the lipocalin protein family (CG5399), and a zinc finger C2H2 transcription factor (CG9932). Mechanistically, we demonstrate that CG5399 overexpression activates the RTK-Akt-mTOR signaling pathway and that activation of insulin receptor (InR) by CG5399 requires cholesterol and clathrin-coated pits at the cell membrane. This study establishes a novel platform for functional genetic studies in Drosophila cells. KW - CRISPR activation KW - genetic screening KW - rapamycin resistance gene JF - eLife SN - 2050-084X PB - eLife Sciences Publications, Ltd ER -