Inhibition of DNAJ-HSP70 interaction improves strength in muscular dystrophy
Rocio Bengoechea, … , Heather L. True, Conrad C. Weihl
Rocio Bengoechea, … , Heather L. True, Conrad C. Weihl
Published May 19, 2020
Citation Information: J Clin Invest. 2020;130(8):4470-4485. https://doi.org/10.1172/JCI136167.
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Research Article Cell biology Muscle biology

Inhibition of DNAJ-HSP70 interaction improves strength in muscular dystrophy

Abstract

Dominant mutations in the HSP70 cochaperone DNAJB6 cause a late-onset muscle disease termed limb-girdle muscular dystrophy type D1 (LGMDD1), which is characterized by protein aggregation and vacuolar myopathology. Disease mutations reside within the G/F domain of DNAJB6, but the molecular mechanisms underlying dysfunction are not well understood. Using yeast, cell culture, and mouse models of LGMDD1, we found that the toxicity associated with disease-associated DNAJB6 required its interaction with HSP70 and that abrogating this interaction genetically or with small molecules was protective. In skeletal muscle, DNAJB6 localizes to the Z-disc with HSP70. Whereas HSP70 normally diffused rapidly between the Z-disc and sarcoplasm, the rate of diffusion of HSP70 in LGMDD1 mouse muscle was diminished, probably because it had an unusual affinity for the Z-disc and mutant DNAJB6. Treating LGMDD1 mice with a small-molecule inhibitor of the DNAJ-HSP70 complex remobilized HSP70, improved strength, and corrected myopathology. These data support a model in which LGMDD1 mutations in DNAJB6 are a gain-of-function disease that is, counterintuitively, mediated via HSP70 binding. Thus, therapeutic approaches targeting HSP70-DNAJB6 may be effective in treating this inherited muscular dystrophy.

Authors

Rocio Bengoechea, Andrew R. Findlay, Ankan K. Bhadra, Hao Shao, Kevin C. Stein, Sara K. Pittman, Jil A.W. Daw, Jason E. Gestwicki, Heather L. True, Conrad C. Weihl

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Figure 2

LGMDD1 mutations in DNAJB6b impair TDP43 disaggregation, which is corrected with a second H31Q mutation.

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LGMDD1 mutations in DNAJB6b impair TDP43 disaggregation, which is correc...
(A–C) HeLa cells were cotransfected with mCherry-TDP43 and WT or LGMDD1-mutant (F89I, F93L, P96R, or F100I) GFP-DNAJB6b. In some cases, a second mutation in the J domain (H31Q) was also present. Transfected cells were heat shocked at 42°C for 1 hour before analysis. (A) Quantification of the percentage of transfected cells with TDP43 aggregates. **P < 0.005, ***P < 0.0005, and ****P < 0.00005, by paired Student’s t test for comparisons between groups; a 1-way between-conditions ANOVA was performed to compare the effects of mutations. There was a significant effect at P < 0.05 for the 5 conditions [F(4,15) = 54.5, P = 9.3 × 10–9]. n = 300 cells analyzed per condition; the experiment was conducted 3 times. (B) Representative fluorescence images of the mCherry-TDP43–positive nuclei quantified in A. Scale bar: 10 μm. (C) Lysates from cells in A were separated into total (T), soluble (S), and pellet (P) fractions and then subjected to SDS-PAGE and Western blotting using anti-TDP43 and anti-GAPDH antibodies. The immunoblot (IB) is representative of 3 independent experiments. Ch-TDP43, mCherry-tagged TDP43.