Chemical genetics reveals a kinase-independent role for protein kinase R in pyroptosis

Formation of the inflammasome, a scaffolding complex that activates Caspase-1, is important in numerous diseases. Pyroptotic cell death induced by anthrax lethal toxin (LT) is a model for inflammasome-mediated Caspase-1 activation. We discovered 7-desacetoxy-6,7-dehydrogedunin (7DG) in a phenotypic screen as a small molecule that protects macrophages from LT-induced death. Using chemical proteomics, we identified protein kinase R (PKR) as the target of 7DG and show that RNAi knockdown of PKR phenocopies treatment with 7DG. Further, we show that PKR's role in ASC assembly and Caspase-1 activation induced by several different inflammasome stimuli is independent of PKR's kinase activity, demonstrating that PKR has a previously uncharacterized role in Caspase-1 activation and Pyroptosis that is distinct from its reported kinase-dependent roles in Apoptosis and inflammasome formation in lipopolysaccharide-primed cells. Remarkably, PKR has different roles in two distinct cell death pathways and has a broad role in inflammasome function relevant in Other Diseases.