Abstract
Objective To understand DNAJB6's function in skeletal muscle and identify therapeutic targets for limb-girdle muscular dystrophy 1D (LGMD1D).
Methods DNAJB6 knockout (KO) myoblasts were generated with Crispr/cas9 technology, and differentially accumulated proteins were identified using stable isotope labeling, followed by quantitative mass spectrometry. Cultured KO myotubes and mouse muscle from DNAJB6b-WT or DNAJB6b-F93L mice were analyzed using histochemistry, immunohistochemistry, and immunoblot. Mouse functional strength measures included forelimb grip strength and inverted wire hang.
Results DNAJB6 inactivation leads to the accumulation of sarcomeric proteins and hypertrophic myotubes with an enhanced fusion index. The increased fusion in DNAJB6 KO myotubes correlates with diminished glycogen synthase kinase-β (GSK3β) activity. In contrast, LGMD1D mutations in DNAJB6 enhance GSK3β activation and suppress β-catenin and NFAT3c signaling. GSK3β inhibition with lithium chloride improves muscle size and strength in an LGMD1D preclinical mouse model.
Conclusions Our results suggest that DNAJB6 facilitates protein quality control and negatively regulates myogenic signaling. In addition, LGMD1D-associated DNAJB6 mutations inhibit myogenic signaling through augmented GSK3β activity. GSK3β inhibition with lithium chloride may be a therapeutic option in LGMD1D.
Glossary
- CSA=
- cross-sectional area;
- GFP=
- green fluorescent protein;
- GO=
- Gene Ontology;
- GSK3β=
- glycogen synthase kinase-β;
- HSP=
- heat shock protein;
- KO=
- knockout;
- LGMD1D=
- limb-girdle muscular dystrophy 1D;
- LC-MS/MS=
- Liquid chromatography with tandem mass spectrometry;
- NFATc3=
- nuclear factor of activated T cells cytoplasmic 3;
- PBS=
- phosphate-buffered saline;
- SILAC=
- stable isotope labeling with amino acids in cell culture;
- WT=
- wild type
Footnotes
Funding information and disclosures are provided at the end of the article. Full disclosure form information provided by the authors is available with the full text of this article at Neurology.org/NG.
↵* These authors contributed equally to the manuscript.
The Article Processing Charge was funded by the authors.
- Received October 19, 2018.
- Accepted in final form February 4, 2019.
- Copyright © 2019 The Author(s). Published by Wolters Kluwer Health, Inc. on behalf of the American Academy of Neurology.
This is an open access article distributed under the terms of the Creative Commons Attribution-NonCommercial-NoDerivatives License 4.0 (CC BY-NC-ND), which permits downloading and sharing the work provided it is properly cited. The work cannot be changed in any way or used commercially without permission from the journal.
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