Force-induced dephosphorylation activates the cochaperone BAG3 to coordinate protein homeostasis and membrane traffic

Proteome maintenance in contracting skeletal and cardiac muscles depends on the chaperone-regulating protein BAG3. Reduced BAG3 activity leads to muscle weakness and heart failure in animal models and patients. BAG3 and its chaperone partners recognize mechanically damaged muscle proteins and initiate their disposal through chaperone-assisted selective autophagy (CASA). However, molecular details of the force-dependent regulation of BAG3 have remained elusive so far. Here, we demonstrate that mechanical stress triggers the dephosphorylation of BAG3 in human muscle and in isolated cells. We identify force-regulated phospho-switches in BAG3 that control CASA complex assembly and CASA activity. Differential proteomics reveal RAB GTPases, which organize membrane traffic and fusion, as dephosphorylation-dependent interactors of BAG3. In fact, RAB7A and RAB11B are shown here to be essential for CASA in skeletal muscle cells. Moreover, BAG3 dephosphorylation is also observed upon induction of mitophagy, suggesting an involvement of the cochaperone in the RAB7A-dependent autophagic engulfment of damaged mitochondria in exercised muscle. Cooperation of BAG3 with RAB7A relies on a direct interaction of both proteins, which is regulated by the nucleotide state of the GTPase and by association with the autophagosome membrane protein LC3B. Finally, we provide evidence that BAG3 and RAB7A also cooperate in non-muscle cells and propose that overactivation of CASA in RAB7A-L129F patients contributes to the loss of peripheral neurons in Charcot-Marie-Tooth neuropathy.