BAG3 Pro209 mutants associated with myopathy and neuropathy relocate chaperones of the CASA-complex to aggresomes

Elias Adriaenssens,Angelo Poletti,Bob Asselbergh,Valeria Crippa,Barbara Tedesco,Francesco Antoniani,Laura Mediani,Vincent Timmerman,Serena Carra

doi:10.1038/s41598-020-65664-z

Abstract

Three missense mutations targeting the same proline 209 (Pro209) codon in the co-chaperone Bcl2-associated athanogene 3 (BAG3) have been reported to cause distal myopathy, dilated cardiomyopathy or Charcot-Marie-Tooth type 2 neuropathy. Yet, it is unclear whether distinct molecular mechanisms underlie the variable clinical spectrum of the rare patients carrying these three heterozygous Pro209 mutations in BAG3. Here, we studied all three variants and compared them to the BAG3_Glu455Lys mutant, which causes dilated cardiomyopathy. We found that all BAG3_Pro209 mutants have acquired a toxic gain-of-function, which causes these variants to accumulate in the form of insoluble HDAC6- and vimentin-positive aggresomes. The aggresomes formed by mutant BAG3 led to a relocation of other chaperones such as HSPB8 and Hsp70, which, together with BAG3, promote the so-called chaperone-assisted selective autophagy (CASA). As a consequence of their increased aggregation-proneness, mutant BAG3 trapped ubiquitinylated client proteins at the aggresome, preventing their efficient clearance. Combined, these data show that all BAG3_Pro209 mutants, irrespective of their different clinical phenotypes, are characterized by a gain-of-function that contributes to the gradual loss of protein homeostasis.

Highlights

Protein homeostasis is maintained by a complex network of molecular chaperones and co-chaperones providing protection to client proteins at every stage of their life-time[1]
In case the substrate cannot be refolded by Hsp[70], the client is directed towards autophagosomal degradation by CHIP and SQSTM1/p62
We studied the pathogenic consequences of 3 previously reported Bcl2associated athanogene 3 (BAG3) missense mutations, located in the second IPV-motif of BAG3 (Fig. 1a) which mediates the interaction with small heat shock proteins (sHSPs) family members, and causing distinct phenotypes: Pro209Leu, Pro209Gln, Pro209Ser

Summary

Introduction

Protein homeostasis is maintained by a complex network of molecular chaperones and co-chaperones providing protection to client proteins at every stage of their life-time[1]. BAG3 is a well-characterized family member that contains a number of additional protein domains besides the conserved BAG-domain, including two Ile-Pro-Val (IPV)-motifs, a PxxP domain and a WW-domain (Fig. 1a). Each of these domains is known to have specific interacting partners. In case the substrate cannot be refolded by Hsp[70], the client is directed towards autophagosomal degradation by CHIP (an E3 ubiquitin ligase that ubiquitinylates the substrate17–19) and SQSTM1/p62 (an autophagy receptor that binds to ubiquitinylated proteins and transfers them to autophagosomes[20,21,22]). By clustering the different components into a single complex, substrates are likely handed over faster to reduce the potentially dangerous dwell time

Methods

Results

Conclusion