Force-induced dephosphorylation activates the cochaperone BAG3 to coordinate protein homeostasis and membrane traffic.

Proteome maintenance in contracting skeletal and cardiac muscles depends on the chaperone-regulating protein BAG3. Reduced BAG3 activity leads to muscle weakness and heart failure in animal models and patients. BAG3 and its chaperone partners recognize mechanically damaged muscle proteins and initiate their disposal through chaperone-assisted selective autophagy (CASA). However, molecular details of the force-dependent regulation of BAG3 have remained elusive so far. Here, we demonstrate that mechanical stress triggers the dephosphorylation of BAG3 in human muscle and in isolated cells. We identify force-regulated phospho-switches in BAG3 that control CASA complex assembly and CASA activity. Differential proteomics reveal RAB GTPases, which organize membrane traffic and fusion, as dephosphorylation-dependent interactors of BAG3. In fact, RAB7A and RAB11B are shown here to be essential for CASA in skeletal muscle cells. Moreover, BAG3 dephosphorylation is also observed upon induction of mitophagy, suggesting an involvement of the cochaperone in the RAB7A-dependent autophagic engulfment of damaged mitochondria in exercised muscle. Cooperation of BAG3 with RAB7A relies on a direct interaction of both proteins, which is regulated by the nucleotide state of the GTPase and by association with the autophagosome membrane protein LC3B. Finally, we provide evidence that BAG3 and RAB7A also cooperate in non-muscle cells and propose that overactivation of CASA in RAB7A-L129F patients contributes to the loss of peripheral neurons in Charcot-Marie-Tooth neuropathy.

Increased resting lactate levels and reduced carbohydrate intake cause νLa.max underestimation by reducing net lactate accumulation-A pilot study in young adults.

Modulation of testing conditions such as resting lactate (Larest) levels or carbohydrate intake may affect the calculation of the maximal glycolytic rate (νLa.max). To evaluate the impact of elevated Larest as well as reduced and increased carbohydrate availability on νLa.max in running sprints (RST), twenty-one participants completed five 15-s RST tests on a running track under five different conditions: (I). baseline: Larest ≤1.5 mmol·L-1; (II). Lactate+: Larest ≥2.5 mmol·L-1; (III). CHO-: carbohydrate intake: ≤ 1 g·kg-1 BW d-1 for 3 days; (IV). CHO+: carbohydrate intake: ≥ 9 g·kg-1 BW d-1 for one day; and (V). acuteCHO: 500 mL glucose containing beverage consumed before RST. νLa.max was significantly reduced in lactate+ and CHO- conditions compared to the baseline RST, due to a reduction in the arithmetic mean delta (∆) between Lapeak and Larest lactate concentration (Lapeak, mmol · L-1). AcuteCHO led to an increase in Larest compared to baseline, CHO- and CHO+ with a high interindividual variability but did not significantly reduce νLa.max. Therefore, avoiding low carbohydrate nutrition before νLa.max testing, along with carefully adjusting Larest to below ≤1.5 mmol·L-1, is crucial to prevent the unintentional underestimation of νLa.max.

microRNA-1 Regulates Metabolic Flexibility in Skeletal Muscle via Pyruvate Metabolism.

MicroRNA-1 (miR-1) is the most abundant miRNA in adult skeletal muscle. To determine the function of miR-1 in adult skeletal muscle, we generated an inducible, skeletal muscle-specific miR-1 knockout (KO) mouse. Integration of RNA-sequencing (RNA-seq) data from miR-1 KO muscle with Argonaute 2 enhanced crosslinking and immunoprecipitation sequencing (AGO2 eCLIP-seq) from human skeletal muscle identified miR-1 target genes involved with glycolysis and pyruvate metabolism. The loss of miR-1 in skeletal muscle induced cancer-like metabolic reprogramming, as shown by higher pyruvate kinase muscle isozyme M2 (PKM2) protein levels, which promoted glycolysis. Comprehensive bioenergetic and metabolic phenotyping combined with skeletal muscle proteomics and metabolomics further demonstrated that miR-1 KO induced metabolic inflexibility as a result of pyruvate oxidation resistance. While the genetic loss of miR-1 reduced endurance exercise performance in mice and in C. elegans, the physiological down-regulation of miR-1 expression in response to a hypertrophic stimulus in both humans and mice causes a similar metabolic reprogramming that supports muscle cell growth. Taken together, these data identify a novel post-translational mechanism of adult skeletal muscle metabolism regulation mediated by miR-1.

Acute exercise activates the AHR in peripheral blood mononuclear cells in an intensity-dependent manner.

The kynurenine pathway (KP) of tryptophan degradation generates several metabolites such as kynurenine (KYN) or kynurenic acid (KA) that serve as endogenous ligands of the aryl hydrocarbon receptor (AHR). Due to its distinct biological roles particularly modulating the immune system, the AHR is a current therapeutic target across different inflammation-related diseases. Here, we show an acute exercise-induced increase in AHR ligand availability on a systemic level and a kynurenine pathway activation in peripheral blood mononuclear cells (PBMCs). Concurrently, the AHR is activated in PBMCs following acute exercise. Exercise effects on both, kynurenic acid and AHR activation in PBMCs were greater in response to high-intensity interval exercise (HIIE) (50 min, six 3-min intervals at 90% V̇o2peak, and 3-min intervals at 50% V̇o2peak in between) compared with workload-matched moderate-intensity continuous exercise (MICE) (50 min). In conclusion, these data indicate a novel mechanistic link in how exercise modulates the immune system through the kynurenine pathway-AHR axis, potentially underlying exercise-induced benefits in various chronic diseases.NEW & NOTEWORTHY The findings of this study show that acute endurance exercise activates a receptor that has been described to integrate metabolic signals into the immune system. We uncover a potential mechanistic link on how exercise modulates the immune system through the kynurenine pathway-AHR axis, potentially underlying exercise-induced benefits in various chronic diseases and of relevance for other cell types.

Resistance training does not increase myocellular garbage dumps: A pilot study on lipofuscin in skeletal muscle fibers of resistance trained young men.

Lipofuscin (LF) is an intracellular aggregate associated with proteostatic impairments, especially prevalent in nondividing skeletal muscle fibers. Reactive oxygen species (ROS) drive LF-formation. Resistance training (RT) improves muscle performance but also increases ROS production, potentially promoting LF-formation. Thus, we aimed to investigate if RT of a mesocycle duration increases LF-formation in type-I and II muscle fibers and whether RT increases the antioxidant capacity (AOC) in terms of SOD1 and SOD2 content. An intervention group (IG) performed 14 eccentrically accented RT-sessions within 7 weeks. Vastus lateralis muscle biopsies were collected before and after the intervention from IG as well as from a control group (CG) which refrained from RT for the same duration. LF was predominantly found near nuclei, followed by membrane-near and a minor amount in the fiber core, with corresponding spot sizes. Overall, LF-content was higher in type-I than type-II fibers (p < 0.05). There was no increase in LF-content in type-I or IIA fibers, neither for the IG following RT nor for the CG. The same is valid for SOD1/2. We conclude that, in healthy subjects, RT can be safely performed, without adverse effects on increased LF-formation.

Resistance exercise: a mighty tool that adapts, destroys, rebuilds and modulates the molecular and structural environment of skeletal muscle.

PURPOSE: Skeletal muscle regulates health and performance by maintaining or increasing strength and muscle mass. Although the molecular mechanisms in response to resistance exercise (RE) significantly target the activation of protein synthesis, a plethora of other mechanisms and structures must be involved in orchestrating the communication, repair, and restoration of homeostasis after RE stimulation. In practice, RE can be modulated by variations in intensity, continuity and volume, which affect molecular responses and skeletal muscle adaptation. Knowledge of these aspects is important with respect to planning of training programs and assessing the impact of RE training on skeletal muscle. METHODS: In this narrative review, we introduce general aspects of skeletal muscle substructures that adapt in response to RE. We further highlighted the molecular mechanisms that control human skeletal muscle anabolism, degradation, repair and memory in response to acute and repeated RE and linked these aspects to major training variables. RESULTS: Although RE is a key stimulus for the activation of skeletal muscle anabolism, it also induces myofibrillar damage. Nevertheless, to increase muscle mass accompanied by a corresponding adaptation of the essential substructures of the sarcomeric environment, RE must be continuously repeated. This requires the permanent engagement of molecular mechanisms that re-establish skeletal muscle integrity after each RE-induced muscle damage. CONCLUSION: Various molecular regulators coordinately control the adaptation of skeletal muscle after acute and repeated RE and expand their actions far beyond muscle growth. Variations of key resistance training variables likely affect these mechanisms without affecting muscle growth.

Repeated and Interrupted Resistance Exercise Induces the Desensitization and Re-Sensitization of mTOR-Related Signaling in Human Skeletal Muscle Fibers.

The acute resistance exercise (RE)-induced phosphorylation of mTOR-related signaling proteins in skeletal muscle can be blunted after repeated RE. The time frame in which the phosphorylation (p) of mTORS2448, p70S6kT421/S424, and rpS6S235/236 will be reduced during an RE training period in humans and whether progressive (PR) loading can counteract such a decline has not been described. (1) To enclose the time frame in which pmTORS2448, prpS6S235/236, and pp70S6kT421/S424 are acutely reduced after RE occurs during repeated RE. (2) To test whether PR will prevent that reduction compared to constant loading (CO) and (3) whether 10 days without RE may re-increase blunted signaling. Fourteen healthy males (24 ± 2.8 yrs.; 1.83 ± 0.1 cm; 79.3 ± 8.5 kg) were subjected to RE with either PR (n = 8) or CO (n = 6) loading. Subjects performed RE thrice per week, conducting three sets with 10−12 repetitions on a leg press and leg extension machine. Muscle biopsies were collected at rest (T0), 45 min after the first (T1), seventh (T7), 13th (T13), and 14th (X-T14) RE session. No differences were found between PR and CO for any parameter. Thus, the groups were combined, and the results show the merged values. prpS6S235/236 and pp70s6kT421/S424 were increased at T1, but were already reduced at T7 and up to T13 compared to T1. Ten days without RE re-increased prpS6S235/236 and pp70S6kT421/S424 at X-T14 to a level comparable to that of T1. pmTORS2448 was increased from T1 to X-T14 and did not decline over the training period. Single-fiber immunohistochemistry revealed a reduction in prpS6S235/236 in type I fibers from T1 to T13 and a re-increase at X-T14, which was more augmented in type II fibers at T13 (p < 0.05). The entity of myofibers revealed a high heterogeneity in the level of prpS6S235/236, possibly reflecting individual contraction-induced stress during RE. The type I and II myofiber diameter increased from T0 and T1 to T13 and X-T14 (p < 0.05) prpS6S235/236 and pp70s6kT421/S424 reflect RE-induced states of desensitization and re-sensitization in dependency on frequent loading by RE, but also by its cessation.