Decoding RNA metabolism by RNA-linked CRISPR screening in human cells

Abstract

RNAs undergo a complex choreography of metabolic processes in human cells that are regulated by thousands of RNA-associated proteins. While individual RNA-associated proteins and their effects on specific RNA metabolic events have been extensively characterized, the full complement of regulators for most RNA metabolic events remain unknown. Here we present a massively parallel RNA-linked CRISPR (ReLiC) screening approach to measure the response of diverse RNA metabolic events to knockout of 2,092 human genes encoding all known RNA-associated proteins. ReLiC screens highlight widespread yet modular interactions between cellular networks regulating splicing, nuclear export, translation, and decay of mRNAs. Isoform-specific ReLiC screens reveal differential regulation of intron retention and exon skipping by SF3b complex subunits. Combining ReLiC with polysome fractionation uncovers translation repression by the trimeric eIF2 initiation complex during nonsense-mediated mRNA decay. ReLiC identifies an unexpected role for GCN1 in suppressing ribosome collisions during treatment with the anti-leukemic drug homoharringtonine. Our work demonstrates ReLiC as a versatile platform for discovering and dissecting regulators of human RNA metabolism.