Dnajb5 From Antarctic Fish Reveals a Redox‐Sensitive Mechanism Coordinating Muscle Regeneration via mTORC1 and HDAC4

Background Dnajb5, a member of the heat shock protein family, has not been previously reported to play a role in muscle differentiation. We identify Dnajb5 as a negative regulator of myogenesis via mammalian target of rapamycin (mTOR) and histone deacetylase 4 (HDAC4) signalling, functioning as a central controller of muscle growth and metabolism. Methods mTOR‐binding proteins were screened by mass spectrometry in Antarctic fish ( Notothenia coriiceps ) muscle extracts. Functional roles of Dnajb5 were examined in C2C12 and primary myoblasts using shRNA‐mediated knockdown. In vivo effects were analysed using a BaCl 2 ‐induced muscle injury model in mice ( n = 6–12 per group). Results Mass spectrometry identified Dnajb5 as an mTOR‐binding protein. This interaction was conserved in mammalian cells and weakened by hydrogen peroxide ( p < 0.05). Dnajb5 knockdown in C2C12 myoblasts selectively increased S6K1 phosphorylation without altering basal Akt phosphorylation. Myogenic differentiation increased, characterized by upregulated Myog , Igf2 and Ckm expression ( p < 0.05) and elevated differentiation index ( p < 0.05). In vivo, Dnajb5 depletion improved tibialis anterior muscle regeneration, increasing regenerating fibre cross‐sectional area by approximately 26% at Day 5 post‐injury ( p < 0.05). Dnajb5 knockdown increased Ppargc1a expression via release of mTORC1 inhibition and preventing the nuclear retention of HDAC4. Mitochondrial DNA copy number increased 1.3‐fold ( p < 0.05), and oxidative enzyme activity rose significantly (succinate dehydrogenase‐positive fibres: 2.8‐fold ( p < 0.01), cytochrome c oxidase‐positive fibres: 1.4‐fold ( p < 0.05). Grip strength improved by 14% ( p < 0.05), and rotarod endurance increased by 31% ( p < 0.01) relative to controls. Conclusions Dnajb5 functions as a negative regulator of muscle differentiation and regeneration by coordinating mTORC1 signalling and the HDAC4‐MEF2 axis. These findings reveal a redox‐sensitive dual‐pathway mechanism linking protein synthesis and mitochondrial biogenesis.