Optimized CRISPR Perturbation Illuminates the Functions of RNA Modification Proteins

A new frontier in functional genomics is to construct “Perturbation Atlas” that systematically profiles the effects of perturbation of all the mammalian genes across various cell types in health and disease. To this end, we have previously developed an in situ CRISPR screening platform in mice named iMAP, where Cas9 guides are “stored” in germline-transmitted transgenes but inducibly “released” (expressed) via Cre-mediated recombination. A major limitation of iMAP is a strong bias in guide representation following the recombination. We now report that this bias was effectively mitigated by eliminating a recombination hotspot at the transgene. We next profiled, across 46 tissues, the abundance of the guides targeting 71 (52%) genes encoding writers, readers and erasers of RNA modifications, revealing their context- specific functions. Furthermore, in a tumor model, we identified regulators of NK cell activation, which was validated in human NK cells, thus offering potential targets for improving cancer immunotherapy. Finally, a public database was established for accessing and analyzing iMAP data. Our study marks an important step toward the creation of the Perturbation Atlas.