Redox state of Protein Kinase A RIα coordinates lysosomal calcium signaling and mitochondrial homeostasis

Protein Kinase A (PKA) Regulatory RIα cysteine redox state regulates docking to D-AKAP1 and may control Drp1 phosphorylation, a principal mediator of mitochondrial fission. RIα C17S knock-in (KI) mice, unable to form disulfides and therefore mimicking the reduced kinase, exhibited enlarged dysfunctional mitochondria. Unexpectedly, this abnormal mitochondrial morphology, which was accompanied by reduced respiration, decreased membrane potential, increased reactive oxygen species and impaired treadmill performance, was not explained by altered Drp1 Ser637 phosphorylation. Transcriptomic analysis revealed decreased mitochondrial biogenesis without loss of total mitochondrial mass, consistent with impaired mitochondrial turnover. KI cells showed impaired lysosomal proteolysis, altered lysosomal calcium homeostasis, reduced RIα-lysosome colocalization and markedly diminished TRPML1 abundance. Pharmacological modulation of lysosomal calcium pathways restored lysosomal calcium signaling in KI cells, whereas activation of TRPML1 signaling impaired mitochondrial respiration in wildtype cells toward KI levels. Together, these findings support a role for disulfide-RIα in coordinating lysosomal calcium signaling and mitochondrial quality control, such that loss of this oxidation state promotes accumulation of dysfunctional mitochondria.