Eukaryotic cells use autophagy and the ubiquitin-proteasome system as their major protein degradation pathways. Upon proteasomal impairment, cells switch to autophagy to ensure proper clearance of clients (the proteasome-to-autophagy switch). The HSPA8 and HSPA1A cochaperone BAG3 has been suggested to be involved in this switch. However, at present it is still unknown whether and to what extent BAG3 can indeed reroute proteasomal clients to the autophagosomal pathway. Here, we show that BAG3 induces the sequestration of ubiquitinated clients into cytoplasmic puncta colabeled with canonical autophagy linkers and markers. Following proteasome inhibition, BAG3 upregulation significantly contributes to the compensatory activation of autophagy and to the degradation of the (poly)ubiquitinated proteins. BAG3 binding to the ubiquitinated clients occurs through the BAG domain, in competition with BAG1, another BAG family member, that normally directs ubiquitinated clients to the proteasome. Therefore, we propose that following proteasome impairment, increasing the BAG3/BAG1 ratio ensures the "BAG-instructed proteasomal to autophagosomal switch and sorting" (BIPASS).
BAG3 is a multi-domain hub that connects two classes of chaperones, small heat shock proteins (sHSPs) via two isoleucine-proline-valine (IPV) motifs and Hsp70 via a BAG domain. Mutations in either the IPV or BAG domain of BAG3 cause a dominant form of myopathy, characterized by protein aggregation in both skeletal and cardiac muscle tissues. Surprisingly, for both disease mutants, impaired chaperone binding is not sufficient to explain disease phenotypes. Recombinant mutants are correctly folded, show unaffected Hsp70 binding but are impaired in stimulating Hsp70-dependent client processing. As a consequence, the mutant BAG3 proteins become the node for a dominant gain of function causing aggregation of itself, Hsp70, Hsp70 clients and tiered interactors within the BAG3 interactome. Importantly, genetic and pharmaceutical interference with Hsp70 binding completely reverses stress-induced protein aggregation for both BAG3 mutations. Thus, the gain of function effects of BAG3 mutants act as Achilles heel of the HSP70 machinery.
The family of the mammalian small heat-shock proteins consists of 10 members (sHSPs/HSPBs: HSPB1–HSPB10) that all share a highly conserved C-terminal alpha-crystallin domain, important for the modulation of both their structural and functional properties. HSPB proteins are biochemically classified as molecular chaperones and participate in protein quality control, preventing the aggregation of unfolded or misfolded proteins and/or assisting in their degradation. Thus, several members of the HSPB family have been suggested to be protective in a number of neurodegenerative and neuromuscular diseases that are characterized by protein misfolding. However, the pro-refolding, anti-aggregation or pro-degradative properties of the various members of the HSPB family differ largely, thereby influencing their efficacy and protective functions. Such diversity depends on several factors, including biochemical and physical properties of the unfolded/misfolded client, the expression levels and the subcellular localization of both the chaperone and the client proteins. Furthermore, although some HSPB members are inefficient at inhibiting protein aggregation, they can still exert neuroprotective effects by other, as yet unidentified, manners; e.g. by maintaining the proper cellular redox state or/and by preventing the activation of the apoptotic cascade. Here, we will focus our attention on how the differences in the activities of the HSPB proteins can influence neurodegenerative and neuromuscular disorders characterized by accumulation of aggregate-prone proteins. Understanding their mechanism of action may allow us to target a specific member in a specific cell type/disease for therapeutic purposes.
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