Gαq-mediated contact release is required for efficient mitochondrial quality control
Lysosomes and mitochondria are highly dynamic organelles whose coordinated interactions are essential for cellular homeostasis, while their dysfunction contributes to numerous human diseases. Here, we identify the mitochondrial outer membrane protein Fis1 and the RabGAPs TBC1D15 and TBC1D17 as novel interacting partners of the heterotrimeric G protein -subunit Gq. Using mitochondrial proteomics, co-immunoprecipitation, proximity ligation assays, and in vitro binding studies with purified proteins, we demonstrate that Gq forms a complex with Fis1 and TBC1D15/TBC1D17 through a non-canonical interaction interface. Gq localizes to mitochondria-lysosome contact sites, where it regulates the dynamic balance between contact formation and dissociation. Loss of Gq resulted in enlarged and clustered lysosomes and increased the extent and duration of mitochondria-lysosome contacts. Live-cell imaging revealed prolonged organelle tethering in Gq-deficient cells, while proximity ligation and three-dimensional reconstruction analyses localized Gq and TBC1D15 to mitochondrial tips closely associated with lysosomes. This localization, together with increased mitochondrial fission and altered mitophagic turnover following Gq loss, links the regulation of contacts at mitochondrial tips to mitochondrial fragmentation and quality control. Re-expression of Gq, but not the non-canonical interaction-deficient mutant, restored normal lysosomal morphology, contact-site dynamics, mitochondrial fragmentation, and mitophagy. Together, these findings identify a Gq as a dynamic regulator of mitochondria-lysosome untethering and connect contact site turnover with mitochondria fragmentation and mitophagy. Our work uncovers a novel link between G protein signaling, organelle communication, and mitochondrial quality control.