Identification of stress specific autophagy regulators from tandem CRISPR screens

Abstract

AbstractAutophagy is a conserved degradative process that promotes cellular homeostasis under stress conditions. Under nutrient starvation autophagy is largely non-selective, promoting the indiscriminate breakdown of cytosolic components. Conversely, selective autophagy is responsible for the specific turnover of damaged organelles including endoplasmic reticula, lysosomes, mitochondria, and peroxisomes. The mechanisms of selective autophagy are best understood through the activity of cargo-specific receptors called autophagy receptors, which facilitate the engulfment of the targeted cargo within autophagosomes, leading to subsequent degradation. We hypothesized that selective autophagy may be regulated by distinct upstream signaling from starvation induced autophagy, providing an additional layer of regulatory control to targeted autophagic degradation. To comprehensively address this question we conducted kinome-wide CRISPR screens to identify distinct signaling pathways responsible for the regulation of basal autophagy, starvation-induced autophagy, and two types of selective autophagy, ER-phagy and pexophagy. These parallel screens identified established and novel autophagy shared regulators under these conditions, as well as kinases specifically required for ER-phagy or pexophagy. More specifically, CDK11A and NME3 were further characterized to be selective ER-phagy regulators. Meanwhile, PAN3 and CDC42BPG were identified as activator or inhibitor of pexophagy, respectively. Collectively, these datasets provide the first comparative description of the kinase signaling specificity, separating regulation of selective autophagy and bulk autophagy.HighlightsParallel pooled kinome genetic knockout screens reveal known and novel regulators of autophagy under basal conditions, nutrient starvation, ER stress, and peroxisomal stressSelective ER and peroxisomal autophagy both have unique activators and inhibitors that distinguish them from bulk autophagyCDK11A and NME3 specifically induce and inhibit ER-phagy, respectivelyPAN3 and CDC42BPG specifically induce and inhibit pexophagy, respectively