Abstract
Restriction of nutrients and oxygen in the tumor microenvironment disrupts ER homeostasis and adaptation to such stress is mediated by the key UPR effector PERK. Given its pro-tumorigenic activity, significant efforts have been made to elucidate the molecular mechanisms that underlie PERK function. Chemical-genetic approaches have recently proven instrumental in pathway mapping and interrogating kinase function. To enable a detailed study of PERK signaling we have generated an analog-sensitive PERK allele that accepts N(6)-alkylated ATP analogs. We find that this allele can be regulated by bulky ATP-competitive inhibitors, confirming the identity of the PERK gatekeeper residue as methionine 886. Furthermore, this analog-sensitive allele can be used to specifically label substrates with thiophosphate both in vitro and in cells. These data highlight the potential for using chemical-genetic techniques to identify novel PERK substrates, thereby providing an expanded view of PERK function and further definition of its signaling networks.