The copper chaperone CCS facilitates copper binding to MEK1/2 to promote kinase activation.

Abstract Normal physiology relies on the precise coordination of intracellular signal transduction pathways that respond to nutrient availability to balance cell growth and cell death. The canonical MAPK pathway consists of the RAF-MEK-ERK signaling cascade and represents one of the most well-defined axes within eukaryotic cells to promote cell proliferation, which underscores its frequent mutational activation in the majority of human cancers. Our recent studies illuminated a function for the redox-active micronutrient copper (Cu) as an intracellular mediator of signaling by connecting Cu to the amplitude of MAPK signaling via a direct interaction between Cu and the kinases MEK1 and MEK2. Given the large quantities of molecules like glutathione and metallothionein that limit cellular toxicity from free Cu ions, evolutionarily conserved Cu chaperones facilitate the efficient delivery of Cu to cuproenzymes. Thus, a dedicated cellular delivery mechanism of Cu to MEK1/2 is likely to exist. Using surface plasmon resonance and proximity-dependent biotin ligase studies, we report here that the Cu chaperone CCS selectively bound to and facilitated Cu transfer to MEK1. Mutations in CCS that disrupt Cu(I) acquisition and exchange or a CCS small molecule inhibitor were employed and resulted in reduced Cu-stimulated MEK1 kinase activity. Our findings indicate that the Cu chaperone CCS provides fidelity within a complex biological system to achieve appropriate installation of Cu within the MEK1 kinase active site that in turn modulates kinase activity and support the development of novel MEK1/2 inhibitors that target the Cu structural interface or blunt dedicated Cu delivery mechanisms via CCS.