Journal of General Physiology最新文献

Mitochondrial fission is a key trigger of cardiac ischemia-reperfusion injuries (IR). Exercise training is an efficient cardioprotective strategy, but its impact on mitochondrial fragmentation during IR remains unknown. Using isolated rat hearts, we found that exercise training limited the activation of dynamin-like protein 1 and limited mitochondrial fragmentation during IR. These results support the hypothesis that exercise training contributes to cardioprotection through its capacity to modulate the mitochondrial fragmentation during IR.

Accurate models of muscle contraction are necessary for understanding muscle performance and the molecular modifications that enhance it (e.g., therapeutics, posttranslational modifications, etc.). As a thermal system containing millions of randomly fluctuating atoms that on the thermal scale of a muscle fiber generate unidirectional force and power output, muscle mechanics are constrained by the laws of thermodynamics. According to a thermodynamic muscle model, muscle's power stroke occurs with the shortening of an entropic spring consisting of an ensemble of force-generating myosin motor switches, each induced by actin binding and gated by inorganic phosphate release. This model differs fundamentally from conventional molecular power stroke models that assign springs to myosin motors in that it is physically impossible to describe an entropic spring in terms of the springs of its molecular constituents. A simple two-state thermodynamic model (a binary mechanical system) accurately accounts for muscle force-velocity relationships, force transients following rapid mechanical and chemical perturbations, and a thermodynamic work loop. Because this model transforms our understanding of muscle contraction, it must continue to be tested. Here, we show that a simple stochastic kinetic simulation of isometric muscle force predicts four phases of a force-generating loop that bifurcates between periodic and stochastic beating through mechanisms framed by two thermodynamic equations. We compare these model predictions with experimental data including observations of spontaneous oscillatory contractions (SPOCs) in muscles and periodic force generation in small myosin ensembles.

Tripartite ATP-independent periplasmic (TRAP) transporters are widespread in prokaryotes and are responsible for the transport of a variety of different ligands, primarily organic acids. TRAP transporters can be divided into two subclasses; DctP-type and TAXI type, which share the same overall architecture and substrate-binding protein requirement. DctP-type transporters are very well studied and have been shown to transport a range of compounds including dicarboxylates, keto acids, and sugar acids. However, TAXI-type transporters are relatively poorly understood. To address this gap in our understanding, we have structurally and biochemically characterized VC0430 from Vibrio cholerae. We show it is a monomeric, high affinity glutamate-binding protein, which we thus rename VcGluP. VcGluP is stereoselective, binding the L-isomer preferentially, and can also bind L-glutamine and L-pyroglutamate with lower affinity. Structural characterization of ligand-bound VcGluP revealed details of its binding site and biophysical characterization of binding site mutants revealed the substrate binding determinants, which differ substantially from those of DctP-type TRAPs. Finally, we have analyzed the interaction between VcGluP and its cognate membrane component, VcGluQM (formerly VC0429) in silico, revealing an architecture hitherto unseen. To our knowledge, this is the first transporter in V. cholerae to be identified as specific to glutamate, which plays a key role in the osmoadaptation of V. cholerae, making this transporter a potential therapeutic target.

Myosin-binding protein H (MyBP-H) is a component of the vertebrate skeletal muscle sarcomere with sequence and domain homology to myosin-binding protein C (MyBP-C). Whereas skeletal muscle isoforms of MyBP-C (fMyBP-C, sMyBP-C) modulate muscle contractility via interactions with actin thin filaments and myosin motors within the muscle sarcomere "C-zone," MyBP-H has no known function. This is in part due to MyBP-H having limited expression in adult fast-twitch muscle and no known involvement in muscle disease. Quantitative proteomics reported here reveal that MyBP-H is highly expressed in prenatal rat fast-twitch muscles and larval zebrafish, suggesting a conserved role in muscle development and prompting studies to define its function. We take advantage of the genetic control of the zebrafish model and a combination of structural, functional, and biophysical techniques to interrogate the role of MyBP-H. Transgenic, FLAG-tagged MyBP-H or fMyBP-C both localize to the C-zones in larval myofibers, whereas genetic depletion of endogenous MyBP-H or fMyBP-C leads to increased accumulation of the other, suggesting competition for C-zone binding sites. Does MyBP-H modulate contractility in the C-zone? Globular domains critical to MyBP-C's modulatory functions are absent from MyBP-H, suggesting that MyBP-H may be functionally silent. However, our results suggest an active role. In vitro motility experiments indicate MyBP-H shares MyBP-C's capacity as a molecular "brake." These results provide new insights and raise questions about the role of the C-zone during muscle development.

Decades of research into striated muscle have provided a robust understanding of the structure and function of the sarcomere and its protein constituents. However, a handful of sarcomere proteins remain that have had little to no functional characterization. These are typically proteins that are highly muscle-type specific or are products of alternative start sites or alternative splicing.

The ryanodine receptor type 1 (RyR1) is a Ca2+ release channel that regulates skeletal muscle contraction by controlling Ca2+ release from the sarcoplasmic reticulum (SR). Posttranslational modifications (PTMs) of RyR1, such as phosphorylation, S-nitrosylation, and carbonylation are known to increase RyR1 open probability (Po), contributing to SR Ca2+ leak and skeletal muscle dysfunction. PTMs on RyR1 have been linked to muscle dysfunction in diseases like breast cancer, rheumatoid arthritis, Duchenne muscle dystrophy, and aging. While reactive oxygen species (ROS) and oxidative stress induce PTMs, the impact of stable oxidative modifications like 3-nitrotyrosine (3-NT) and malondialdehyde adducts (MDA) on RyR1 gating remains unclear. Mass spectrometry and single-channel recordings were used to study how 3-NT and MDA modify RyR1 and affect Po. Both modifications increased Po in a dose-dependent manner, with mass spectrometry identifying 30 modified residues out of 5035 amino acids per RyR1 monomer. Key modifications were found in domains critical for protein interaction and channel activation, including Y808/3NT in SPRY1, Y1081/3NT and H1254/MDA in SPRY2&3, and Q2107/MDA and Y2128/3NT in JSol, near the binding site of FKBP12. Though these modifications did not directly overlap with FKBP12 binding residues, they promoted FKBP12 dissociation from RyR1. These findings provide detailed insights into how stable oxidative PTMs on RyR1 residues alter channel gating, advancing our understanding of RyR1-mediated Ca2+ release in conditions associated with oxidative stress and muscle weakness.

JGP study (Steinz et al. https://doi.org/10.1085/jgp.202313515) reveals that oxidative stress can induce stable posttranslational modifications of RyR1 that increase the channel's open probability and could therefore disrupt muscle contractility.