Running title: PKCε controls INrf2:Nrf2 interaction SUMMARY

The INrf2 (Keap1):Nrf2 complex plays a critical role in protection against chemical and radiation-induced oxidative stress and cellular transformation. INrf2 in association with Cul3-Rbx1 ubiquitinylates and degrades Nrf2. Exposure to stressors leads to Nrf2 stabilization and coordinated activation of cytoprotective proteins and cellular protection. However, the molecular signal(s) that regulate INrf2 control of Nrf2 remains elusive. In this report, we demonstrate that oncoprotein PKCε phosphorylation of INrf2 at Ser599 and Ser602 is essential for specific INrf2:Nrf2 interaction and Nrf2 ubiquitination/degradation. Inhibition of PKCε, MEFs lacking PKCε, and the INrf2S602A mutant all failed to phosphorylate INrf2 leading to loss of INrf2:Nrf2 interaction, Nrf2 degradation and enhanced cytoprotection/drug resistance. Molecular modeling analyses revealed that phosphorylation of S599 exposes the deeply buried S602 for phosphorylation and enhanced INrf2:Nrf2 interaction. Analysis of human lung and liver tumor protein arrays showed lower PKCε and higher Nrf2 that presumably promoted cancer cell survival and drug resistance. In conclusion, phosphorylation of INrf2 by PKCε leads to regulation of Nrf2 with significant implications in survival of cancer cells that often express lower levels of PKCε