Mislocalization of protein kinase A drives pathology in Cushing’s syndrome

Mutations in the catalytic subunit of protein kinase A (PKAc) drive the stress hormone disorder adrenal Cushing’s syndrome. We define mechanisms of action for the PKAc-L205R and W196R variants. Proximity proteomic techniques demonstrate that both Cushing’s mutants are excluded from A kinase-anchoring protein (AKAP)-signaling islands, whereas live-cell photoactivation microscopy reveals that these kinase mutants indiscriminately diffuse throughout the cell. Only cAMP analog drugs that displace native PKAc from AKAPs enhance cortisol release. Rescue experiments that incorporate PKAc mutants into AKAP complexes abolish cortisol overproduction, indicating that kinase anchoring restores normal endocrine function. Analyses of adrenal-specific PKAc-W196R knockin mice and Cushing’s syndrome patient tissue reveal defective signaling mechanisms of the disease. Surprisingly each Cushing’s mutant engages a different mitogenic-signaling pathway, with upregulation of YAP/TAZ by PKAc-L205R and ERK kinase activation by PKAc-W196R. Thus, aberrant spatiotemporal regulation of each Cushing’s variant promotes the transmission of distinct downstream pathogenic signals. [Display omitted] •PKAc mutants found in adrenal Cushing’s syndrome are chronically mislocalized•Displacement from AKAPs is required for stress hormone overproduction by mutants•A PKAc-W196R knockin mouse recapitulates adrenal Cushing’s syndrome hallmarks•PKAc-L205R and PKAc-W196R drive distinct downstream signaling pathways Mutations in the catalytic subunit of protein kinase A (PKAc) cause a stress hormone disorder called adrenal Cushing’s syndrome. Omar et al. uncover mislocalization as a required component of mutant-kinase pathology. Remarkably, two seemingly similar disease-causing PKAc variants diverge in subcellular localization, physiochemical properties, and downstream signaling pathways.