-
Meat Science Jul 2022The development of myofibrillar proteins drinks (MPDs) can provide meat protein nutrition to specific groups of people. However, one major challenge is that myofibrillar... (Review)
Review
The development of myofibrillar proteins drinks (MPDs) can provide meat protein nutrition to specific groups of people. However, one major challenge is that myofibrillar proteins (MPs) are insoluble in solutions with a low ionic strength. Another functional constraint is the susceptibility of MPs to heat-induced aggregation. Currently, the primary approach used to improve the water solubility of MPs is to inhibit the assembly of myofilaments. Increasing the thermostability of MPs primarily inhibits the aggregation of myosin or oxidizes myosin to soluble substances. This review focuses on the description of several chemical and physical strategies, with an emphasis on the advantages, disadvantages, and recent progress. Under the myosin filament assembly process and the cross-linking aggregation mechanism, this summary helps improve our understanding of the solution and thermostability of MPs in low-ionic-strength solutions, thus providing new ideas to the development of MPDs.
Topics: Humans; Solubility; Myosins; Myofibrils; Osmolar Concentration
PubMed: 35413661
DOI: 10.1016/j.meatsci.2022.108822 -
Life Science Alliance Apr 2022Protein isoform transitions confer muscle fibers with distinct properties and are regulated by differential transcription and alternative splicing. RNA-binding Fox...
Protein isoform transitions confer muscle fibers with distinct properties and are regulated by differential transcription and alternative splicing. RNA-binding Fox protein 1 (Rbfox1) can affect both transcript levels and splicing, and is known to contribute to normal muscle development and physiology in vertebrates, although the detailed mechanisms remain obscure. In this study, we report that Rbfox1 contributes to the generation of adult muscle diversity in Rbfox1 is differentially expressed among muscle fiber types, and RNAi knockdown causes a hypercontraction phenotype that leads to behavioral and eclosion defects. Misregulation of fiber type-specific gene and splice isoform expression, notably loss of an indirect flight muscle-specific isoform of Troponin-I that is critical for regulating myosin activity, leads to structural defects. We further show that Rbfox1 directly binds the 3'-UTR of target transcripts, regulates the expression level of myogenic transcription factors myocyte enhancer factor 2 and Salm, and both modulates expression of and genetically interacts with the CELF family RNA-binding protein Bruno1 (Bru1). Rbfox1 and Bru1 co-regulate fiber type-specific alternative splicing of structural genes, indicating that regulatory interactions between FOX and CELF family RNA-binding proteins are conserved in fly muscle. Rbfox1 thus affects muscle development by regulating fiber type-specific splicing and expression dynamics of identity genes and structural proteins.
Topics: Animals; Drosophila; Drosophila Proteins; Female; Gene Knockdown Techniques; Male; Myofibrils; Protein Isoforms; RNA-Binding Proteins
PubMed: 34996845
DOI: 10.26508/lsa.202101342 -
Journal of Food Science Aug 2018Bromelain was used to tenderize golden pomfrets (Trachinotus blochii). The enzyme kinetic model was x=2.447×ln[1+(1332.21×E0S0-1.74)t], which indicated that the degree...
UNLABELLED
Bromelain was used to tenderize golden pomfrets (Trachinotus blochii). The enzyme kinetic model was x=2.447×ln[1+(1332.21×E0S0-1.74)t], which indicated that the degree of hydrolysis (DH, x) was dependent on hydrolysis time (t), the initial concentration of myofibril (S ) and bromelain (E ). The relationship between the overall hydrolysis rate (v), S , E , and t is demonstrated as: v=(16.50(E0S0)-1.33)S0 exp {-2.447ln[1+(1332.21E0S0-1.74)t2]}. Sample of 0.40% E /S was further used to study the effects of hydrolysis time on the changes of proteins, peptides, free amino acids (FAA), and protein nanostructure. SDS-PAGE result showed that myosin heavy chain was degraded dramatically from 22.88% before treatment to 12.03% after 2 min bromelain treatment. Meanwhile, bromelain did not exhibit activity towards actin, trypomyosin, myosin light chain, and troponin C. A general increase of amino acids indicated the increased DH and the preferential cleavage sites of bromelain in the descending order of lysine, glutamic acid, glycine, ornithine, methionine sulfoxide, and alanine. Atomic force microscope images showed that the strip-like structure of myofibril was considerably degraded by bromelain, and the granulation of protein after 20 min indicated possible self-assembling of protein hydrolysate. Confocal laser scanning microscopy further confirmed the degradation of myofibril proteins and formation of protein aggregates.
PRACTICAL APPLICATION
Meat of golden pomfrets is tough, thus not idea for fish balls or fish cakes. Tenderization is essential to achieve desired texture and consumer acceptance, especially for this fish meat with intrinsic hard texture. Bromelain can be extracted from pineapple processing waste. Enzymatic kinetics was studied to instruct industry to control the tenderness of the processed fish meat. The microstructural and mechanism study elucidate the process, thus could be applied to improve the quality of the seafood products correspondingly.
Topics: Amino Acids; Ananas; Animals; Bromelains; Fishes; Food Handling; Hydrolysis; Kinetics; Meat; Myofibrils; Protein Hydrolysates; Proteins; Seafood
PubMed: 30020543
DOI: 10.1111/1750-3841.14212 -
Journal of Cell Science Oct 2022Myofibrils are the intracellular structures formed by actin and myosin filaments. They are paracrystalline contractile cables with unusually well-defined dimensions. The...
Myofibrils are the intracellular structures formed by actin and myosin filaments. They are paracrystalline contractile cables with unusually well-defined dimensions. The sliding of actin past myosin filaments powers contractions, and the entire system is held in place by a structure called the Z-disc, which anchors the actin filaments. Myosin filaments, in turn, are anchored to another structure called the M-line. Most of the complex architecture of myofibrils can be reduced to studying the Z-disc, and recently, important advances regarding the arrangement and function of Z-discs in insects have been published. On a very small scale, we have detailed protein structure information. At the medium scale, we have cryo-electron microscopy maps, super-resolution microscopy and protein-protein interaction networks, while at the functional scale, phenotypic data are available from precise genetic manipulations. All these data aim to answer how the Z-disc works and how it is assembled. Here, we summarize recent data from insects and explore how it fits into our view of the Z-disc, myofibrils and, ultimately, muscles.
Topics: Actins; Animals; Biology; Cryoelectron Microscopy; Insecta; Myofibrils; Myosins; Sarcomeres
PubMed: 36226637
DOI: 10.1242/jcs.260179 -
Annals of Anatomy = Anatomischer... Aug 2023The force a muscle exerts is partly determined by anatomical parameters, such as its physiological cross-section. The temporal muscle is structurally heterogeneous. To...
BACKGROUND
The force a muscle exerts is partly determined by anatomical parameters, such as its physiological cross-section. The temporal muscle is structurally heterogeneous. To the authors' knowledge, the ultrastructure of this muscle has been poorly specifically studied.
METHODS
Five adult Wistar rats weighting 350-400 g were used as temporal muscle donors. Tissues were specifically processed and studied under transmission electron microscope.
RESULTS
On ultrathin cuts, the general ultrastructural pattern of striated muscles was observed. Moreover, pennate sarcomeres were identified, sharing a one-end insertion on the same Z-disc. Bipennate morphologies resulted when two neighbor sarcomeres, attached on different neighbor Z-discs and separated at that end by a triad, converged to the same Z-disc at the opposite ends, thus building a thicker myofibril distinctively flanked by triads. Tripennate morphologies were identified when sarcomeres from three different Z-discs converged to the same Z-disc at the opposite ends.
CONCLUSIONS
These results support recent evidence of sarcomeres branching gathered in mice. Adequate identification of the sites of excitation-contraction coupling should be on both sides of a myofibril, on bidimensional ultrathin cuts, to avoid false positive results due to putative longitudinal folds of myofibrils.
Topics: Animals; Rats; Mice; Myofibrils; Temporal Muscle; Rats, Wistar; Sarcomeres; Muscle, Skeletal; Muscle Contraction
PubMed: 37011827
DOI: 10.1016/j.aanat.2023.152096 -
BMC Developmental Biology Aug 2021Flying is an essential function for mosquitoes, required for mating and, in the case of females, to get a blood meal and consequently function as a vector. Flight...
BACKGROUND
Flying is an essential function for mosquitoes, required for mating and, in the case of females, to get a blood meal and consequently function as a vector. Flight depends on the action of the indirect flight muscles (IFMs), which power the wings beat. No description of the development of IFMs in mosquitoes, including Aedes aegypti, is available.
METHODS
A. aegypti thoraces of larvae 3 and larvae 4 (L3 and L4) instars were analyzed using histochemistry and bright field microscopy. IFM primordia from L3 and L4 and IFMs from pupal and adult stages were dissected and processed to detect F-actin labelling with phalloidin-rhodamine or TRITC, or to immunodetection of myosin and tubulin using specific antibodies, these samples were analyzed by confocal microscopy. Other samples were studied using transmission electron microscopy.
RESULTS
At L3-L4, IFM primordia for dorsal-longitudinal muscles (DLM) and dorsal-ventral muscles (DVM) were identified in the expected locations in the thoracic region: three primordia per hemithorax corresponding to DLM with anterior to posterior orientation were present. Other three primordia per hemithorax, corresponding to DVM, had lateral position and dorsal to ventral orientation. During L3 to L4 myoblast fusion led to syncytial myotubes formation, followed by myotendon junctions (MTJ) creation, myofibrils assembly and sarcomere maturation. The formation of Z-discs and M-line during sarcomere maturation was observed in pupal stage and, the structure reached in teneral insects a classical myosin thick, and actin thin filaments arranged in a hexagonal lattice structure.
CONCLUSIONS
A general description of A. aegypti IFM development is presented, from the myoblast fusion at L3 to form myotubes, to sarcomere maturation at adult stage. Several differences during IFM development were observed between A. aegypti (Nematoceran) and Drosophila melanogaster (Brachyceran) and, similitudes with Chironomus sp. were observed as this insect is a Nematoceran, which is taxonomically closer to A. aegypti and share the same number of larval stages.
Topics: Aedes; Animals; Arboviruses; Drosophila melanogaster; Mosquito Vectors; Sarcomeres
PubMed: 34445959
DOI: 10.1186/s12861-021-00242-8 -
Cell and Tissue Research Jul 2023In vertebrate skeletal muscles, the architecture of myofibrils is particularly well conserved throughout the taxa. It is composed of suites of repeating functional units...
In vertebrate skeletal muscles, the architecture of myofibrils is particularly well conserved throughout the taxa. It is composed of suites of repeating functional units called sarcomeres which give the muscle its striated structure. Here, we show that the skeletal sound producing muscles of the cusk eel Parophidion vassali have a different organisation, distinct from the classical type found in textbooks. Within sarcomeres, filaments are not straight lines but have a Y-shaped structure. This looks like chicken wire, with one branch connecting to a branch from the myofibril above and the other connecting to a branch from the myofibril below. This organisation seems to be an adaptation to counteract a trade-off between the speed and force. The low ratio of myofibrils within cell muscles and the high volume of sarcoplasmic reticulum strongly suggest that these muscles are capable of fast contractions. In parallel, the Z-bands are quite wide about 30% of the sarcomere length. This extraordinary long Z-band could smooth out the tension variations found in high-speed muscle contraction, helping to produce sounds with low variabilities in the sound features. Simultaneously, the Y-shaped structure allows having more cross-bridges, increasing the force in this high-speed muscle.
Topics: Myofibrils; Sarcomeres; Muscle Fibers, Skeletal; Muscle Contraction; Muscle, Skeletal
PubMed: 37129618
DOI: 10.1007/s00441-023-03775-5 -
Circulation Jan 2022Titin truncation variants (TTNtvs) are the most common inheritable risk factor for dilated cardiomyopathy (DCM), a disease with high morbidity and mortality. The...
BACKGROUND
Titin truncation variants (TTNtvs) are the most common inheritable risk factor for dilated cardiomyopathy (DCM), a disease with high morbidity and mortality. The pathogenicity of TTNtvs has been associated with structural localization as A-band variants overlapping myosin heavy chain-binding domains are more pathogenic than I-band variants by incompletely understood mechanisms. Demonstrating why A-band variants are highly pathogenic for DCM could reveal new insights into DCM pathogenesis, titin (TTN) functions, and therapeutic targets.
METHODS
We constructed human cardiomyocyte models harboring DCM-associated TTNtvs within A-band and I-band structural domains using induced pluripotent stem cell and CRISPR technologies. We characterized normal TTN isoforms and variant-specific truncation peptides by their expression levels and cardiomyocyte localization using TTN protein gel electrophoresis and immunofluorescence, respectively. Using CRISPR to ablate A-band variant-specific truncation peptides through introduction of a proximal I-band TTNtv, we studied genetic mechanisms in single cardiomyocyte and 3-dimensional, biomimetic cardiac microtissue functional assays. Last, we engineered a full-length TTN protein reporter assay and used next-generation sequencing assays to develop a CRISPR therapeutic for somatic cell genome editing TTNtvs.
RESULTS
An A-band TTNtv dose-dependently impaired cardiac microtissue twitch force, reduced full-length TTN levels, and produced abundant TTN truncation peptides. TTN truncation peptides integrated into nascent myofibril-like structures and impaired myofibrillogenesis. CRISPR ablation of TTN truncation peptides using a proximal I-band TTNtv partially restored cardiac microtissue twitch force deficits. Cardiomyocyte genome editing using SpCas9 and a TTNtv-specific guide RNA restored the TTN protein reading frame, which increased full-length TTN protein levels, reduced TTN truncation peptides, and increased sarcomere function in cardiac microtissue assays.
CONCLUSIONS
An A-band TTNtv diminished sarcomere function greater than an I-band TTNtv in proportion to estimated DCM pathogenicity. Although both TTNtvs resulted in full-length TTN haploinsufficiency, only the A-band TTNtv produced TTN truncation peptides that impaired myofibrillogenesis and sarcomere function. CRISPR-mediated reading frame repair of the A-band TTNtv restored functional deficits, and could be adapted as a one-and-done genome editing strategy to target ≈30% of DCM-associated TTNtvs.
Topics: Cardiomyopathy, Dilated; Connectin; Gene Editing; Genetic Variation; Humans; Induced Pluripotent Stem Cells; Myocytes, Cardiac; Myofibrils; Reading Frames
PubMed: 34905694
DOI: 10.1161/CIRCULATIONAHA.120.049997 -
Journal of Muscle Research and Cell... Aug 2017A basic goal in muscle research is to understand how the cyclic ATPase activity of cross-bridges is converted into mechanical force. A direct approach to study the... (Review)
Review
A basic goal in muscle research is to understand how the cyclic ATPase activity of cross-bridges is converted into mechanical force. A direct approach to study the chemo-mechanical coupling between P release and the force-generating step is provided by the kinetics of force response induced by a rapid change in [P]. Classical studies on fibres using caged-P discovered that rapid increases in [P] induce fast force decays dependent on final [P] whose kinetics were interpreted to probe a fast force-generating step prior to P release. However, this hypothesis was called into question by studies on skeletal and cardiac myofibrils subjected to P jumps in both directions (increases and decreases in [P]) which revealed that rapid decreases in [P] trigger force rises with slow kinetics, similar to those of calcium-induced force development and mechanically-induced force redevelopment at the same [P]. A possible explanation for this discrepancy came from imaging of individual sarcomeres in cardiac myofibrils, showing that the fast force decay upon increase in [P] results from so-called sarcomere 'give'. The slow force rise upon decrease in [P] was found to better reflect overall sarcomeres cross-bridge kinetics and its [P] dependence, suggesting that the force generation coupled to P release cannot be separated from the rate-limiting transition. The reasons for the different conclusions achieved in fibre and myofibril studies are re-examined as the recent findings on cardiac myofibrils have fundamental consequences for the coupling between P release, rate-limiting steps and force generation. The implications from P-induced force kinetics of myofibrils are discussed in combination with historical and recent models of the cross-bridge cycle.
Topics: Adenosine Triphosphate; Animals; Calcium; Humans; Isometric Contraction; Kinetics; Muscle Strength; Phosphates; Sarcomeres
PubMed: 28918606
DOI: 10.1007/s10974-017-9482-8 -
The Journal of Physiology Jan 2016The contractile properties of human fetal cardiac muscle have not been previously studied. Small-scale approaches such as isolated myofibril and isolated contractile...
KEY POINTS
The contractile properties of human fetal cardiac muscle have not been previously studied. Small-scale approaches such as isolated myofibril and isolated contractile protein biomechanical assays allow study of activation and relaxation kinetics of human fetal cardiac muscle under well-controlled conditions. We have examined the contractile properties of human fetal cardiac myofibrils and myosin across gestational age 59-134 days. Human fetal cardiac myofibrils have low force and slow kinetics of activation and relaxation that increase during the time period studied, and kinetic changes may result from structural maturation and changes in protein isoform expression. Understanding the time course of human fetal cardiac muscle structure and contractile maturation can provide a framework to study development of contractile dysfunction with disease and evaluate the maturation state of cultured stem cell-derived cardiomyocytes.
ABSTRACT
Little is known about the contractile properties of human fetal cardiac muscle during development. Understanding these contractile properties, and how they change throughout development, can provide valuable insight into human heart development, and provide a framework to study the early stages of cardiac diseases that develop in utero. We characterized the contractile properties of isolated human fetal cardiac myofibrils across 8-19 weeks of gestation. Mechanical measurements revealed that in early stages of gestation there is low specific force and slow rates of force development and relaxation, with increases in force and the rates of activation and relaxation as gestation progresses. The duration and slope of the initial, slow phase of relaxation, related to myosin detachment and thin filament deactivation rates, decreased with gestation age. F-actin sliding on human fetal cardiac myosin-coated surfaces slowed significantly from 108 to 130 days of gestation. Electron micrographs showed human fetal muscle myofibrils elongate and widen with age, but features such as the M-line and Z-band are apparent even as early as day 52. Protein isoform analysis revealed that β-myosin is predominantly expressed even at the earliest time point studied, but there is a progressive increase in expression of cardiac troponin I (TnI), with a concurrent decrease in slow skeletal TnI. Together, our results suggest that cardiac myofibril force production and kinetics of activation and relaxation change significantly with gestation age and are influenced by the structural maturation of the sarcomere and changes in contractile filament protein isoforms.
Topics: Actins; Adult; Female; Fetal Heart; Humans; Male; Myocardial Contraction; Myofibrils; Myosins; Troponin I
PubMed: 26460603
DOI: 10.1113/JP271290