DNAJB6 mutants display toxic gain of function through unregulated interaction with Hsp70 chaperones

Abstract

Molecular chaperones are essential cellular components that aid in protein folding and preventing the abnormal aggregation of disease-associated proteins. Mutations in one such chaperone, DNAJB6, were identified in patients with LGMDD1, a dominant autosomal disorder characterized by myofibrillar degeneration and accumulations of aggregated protein within myocytes. The molecular mechanisms through which such mutations cause this dysfunction, however, are not well understood. Here we employ a combination of solution NMR and biochemical assays to investigate the structural and functional changes in LGMDD1 mutants of DNAJB6. Surprisingly, we find that DNAJB6 disease mutants show no reduction in their aggregation-prevention activity in vitro, and instead differ structurally from the WT protein, affecting their interaction with Hsp70 chaperones. While WT DNAJB6 contains a helical element regulating its ability to bind and activate Hsp70, in LGMDD1 disease mutants this regulation is disrupted. These variants can thus recruit and hyperactivate Hsp70 chaperones in an unregulated manner, depleting Hsp70 levels in myocytes, and resulting in the disruption of proteostasis. Interfering with DNAJB6-Hsp70 binding, however, reverses the disease phenotype, suggesting future therapeutic avenues for LGMDD1.

Tertiary structure and conformational dynamics of the anti-amyloidogenic chaperone DNAJB6b at atomistic resolution

“…Together, our data suggest that LGMD causing mutations in the G/F 1 domain have an impact on the stability of the autoinhibitory state potentially leading to release of helix V from helices II/III and hence allowing access of Hsp70 in the absence of substrates in line with the recent findings of Abayev-Avraham et al. in the accompanying paper [32].…”

“…However, LGMDD1-mutated DNAJB6b shows alterations in the interactions between helix V and helices II/III that ultimately can destabilize this autoinhibition. This is in line with the NMR data in the corresponding paper (Abayev-Avraham et al [32]), showing that this autoinhibitory regulation is disrupted in LGMDD1 mutants of DNAJB6b such that they recruit Hsp70 in an unregulated, non-functional manner.…”

Defining a small-molecule stimulator of the human Hsp70-disaggregase system with selectivity for DnaJB proteins

“…Indeed, overactivation of Hsp70 by specific Hsp40s can underlie disease. 93,94 Small-molecule inhibitors of Hsp70, which would presumably reduce disaggregase activity, can prevent pathological tau accumulation. [95][96][97][98][99] Moreover, the Hsp70-disaggregase system can promote protein aggregation and toxicity in C. elegans.…”

Unique characteristics of the J-domain proximal regions of Hsp70 cochaperone Apj1 in prion propagation/elimination and its overlap with Sis1 function

“…However, what Helix V is doing in the context of the short N-terminal segments of Class A/B JDPs is not well understood. Experimental evidence has led to the hypothesis that Helix V plays a role in modulating the substrate binding cycle of Hsp70 in both Class A and B JDPs ( Ciesielski, et al, 2024 ) while also playing an important intramolecular regulatory role in eukaryotic Class B JDPs ( Yu et al, 2015a ; Yu et al, 2015b ; Faust et al, 2020 ; Abayev-Avraham et al, 2023 ).…”