Discovery of new intracellular junctions: The calcium entry units (CEUs)

In 2017, Boncompagni, Michelucci et al. demonstrated that during exercise the sarcotubular system of extensor digitorum longus (EDL) fibers undergoes a profound remodeling that leads to the assembly of new junctions between T-tubule extensions at the I band and sarcoplasmic reticulum (SR) stacks. As these junctions contain colocalized STIM1 and Orai1 and enhance store-operated Ca2+ entry (SOCE), they have been named Ca2+ entry units (CEUs). In addition, it has been more recently shown that (1) CEUs disassemble following recovery, with T-tubules retraction from the I band faster than SR stacks disassembly, and (2) lack of calsequestrin-1 (CASQ1) induces a constitutive assembly of CEUs, resulting in enhanced SOCE that counteracts the SR Ca2+ depletion. We have now analyzed (1) CEUs during postnatal maturation (at 2 wk of age) and (2) whether CEUs form in slow-twitch fibers (soleus). (a) Compared with adult (4 mo) EDL fibers of resting WT mice, at 2 wk of age we found a greater longitudinal disposition of T-tubules associated to SR membranes forming junctions virtually identical to CEUs in adult EDLs of exercised WT mice, which promote increased STIM1/Orai1-mediated SOCE. (b) We also compared structure and function of soleus (which also express the cardiac isoform CASQ2) from WT mice and from mice lacking either CASQ1 (CASQ1-null) or CASQ1/2 (dCASQ-null). In soleus from both genotypes, CEUs are constitutively assembled although they appear structurally smaller than those described previously in exercised WT or CASQ1-null EDLs. A detailed EM quantitative analysis revealed that CEUs were more abundant in dCASQ-null than CASQ1-null mice. The amount of CEUs strictly correlated with the ability of soleus fibers to recover extracellular Ca2+ via SOCE to support contractility during high-frequency stimulation. These data were supported by molecular analysis of Western blots, showing that Orai1 expression was enhanced following ablation of CASQ.

Effect of marketable ages on meat quality through fiber characteristics in goose

Background: Goose meat is more and more popular among consumers because of its good quality. The fiber characteristics, as a key determinant factor, contributing to meat quality has been well demonstrated, and the marketable ages are also closely related to meat quality in livestock and chicken. However, little is known about the effect of different marketable ages on the meat quality through fiber characteristics in goose. Results: Here, 1-day, 28-day and three market-age old (70, 90 & 120 days) Yangzhou geese were selected and their fiber characteristics were investigated. The results showed that only fast-twitch fibers were observed in the breast muscle irrespective of the ages, while little slow-twitch fibers could be identified in leg muscle in three marketable ages, especially in gastrocnemius and extensor digitorum longus. As for the fiber diameter, a rapid upward trend was observed in breast muscle from 70 days to 90 days, and the corresponding values were 19.88 to 26.27μm, respectively, and it remained stable 90d thereafter. While the diameter and cross-sectional area of muscle fiber in leg muscle increased with ages. In addition, the proximate composition and physical properties was measured at different ages. 120-day-old geese had richer intramuscular fat and protein content both in breast and leg meat, as well as lower moisture content among three marketable ages. The higher lightness and pressing loss and the lower redness and shear force was observed in the breast and leg meat of 70-day-old geese compared with the 90 and 120-day-old geese. Conclusions: Taken together, although longer marketable age did not affect muscle fiber type in geese, it resulted in a thicker muscle fiber area, richer intramuscular fat and protein content, redder and chewier meat. As a result, the reasonable marketable age should be taken into account to improve the meat quality in the goose production.

Age-Related Maintenance of the Autophagy-Lysosomal System Is Dependent on Skeletal Muscle Type

The skeletal muscle plays an important role in maintaining whole-body mechanics, metabolic homeostasis, and interorgan crosstalk. However, during aging, functional and structural changes such as fiber integrity loss and atrophy can occur across different species. A commonly observed hallmark of aged skeletal muscle is the accumulation of oxidatively modified proteins and protein aggregates which point to an imbalance in proteostasis systems such as degradation machineries. Recently, we showed that the ubiquitin-proteasomal system was impaired. Specifically, the proteasomal activity, which was declining in aged M. soleus (SOL) and M. extensor digitorum longus (EDL). Therefore, in order to understand whether another proteolytic system would compensate the decline in proteasomal activity, we aimed to investigate age-related changes in the autophagy-lysosomal system (ALS) in SOL, mostly consisting of slow-twitch fibers, and EDL, mainly composed of fast-twitch fibers, from young (4 months) and old (25 months) C57BL/6JRj mice. Here, we focused on changes in the content of modified proteins and the ALS. Our results show that aged SOL and EDL display high levels of protein modifications, particularly in old SOL. While autophagy machinery appears to be functional, lysosomal activity declines gradually in aged SOL. In contrast, in old EDL, the ALS seems to be affected, demonstrated by an increased level of key autophagy-related proteins, which are known to accumulate when their delivery or degradation is impaired. In fact, lysosomal activity was significantly decreased in old EDL. Results presented herein suggest that the ALS can compensate the high levels of modified proteins in the more oxidative muscle, SOL, while EDL seems to be more prone to ALS age-related alterations.

Muscle fiber typology substantially influences time to recover from high-intensity exercise

Human fast-twitch muscle fibers generate high power in a short amount of time but are easily fatigued, whereas slow-twitch fibers are more fatigue resistant. The transfer of this knowledge to coaching is hampered by the invasive nature of the current evaluation of muscle typology by biopsies. Therefore, a noninvasive method was developed to estimate muscle typology through proton magnetic resonance spectroscopy in the gastrocnemius. The aim of this study was to investigate whether male subjects with an a priori-determined fast typology (FT) are characterized by a more pronounced Wingate exercise-induced fatigue and delayed recovery compared with subjects with a slow typology (ST). Ten subjects with an estimated higher percentage of fast-twitch fibers and 10 subjects with an estimated higher percentage of slow-twitch fibers underwent the test protocol, consisting of three 30-s all-out Wingate tests. Recovery of knee extension torque was evaluated by maximal voluntary contraction combined with electrical stimulation up to 5 h after the Wingate tests. Although both groups delivered the same mean power across all Wingates, the power drop was higher in the FT group (−61%) compared with the ST group (−41%). The torque at maximal voluntary contraction had fully recovered in the ST group after 20 min, whereas the FT group had not yet recovered 5 h into recovery. This noninvasive estimation of muscle typology can predict the extent of fatigue and time to recover following repeated all-out exercise and may have applications as a tool to individualize training and recovery cycles. NEW & NOTEWORTHY A one-fits-all training regime is present in most sports, though the same training implies different stimuli in athletes with a distinct muscle typology. Individualization of training based on this muscle typology might be important to optimize performance and to lower the risk for accumulated fatigue and potentially injury. When conducting research, one should keep in mind that the muscle typology of participants influences the severity of fatigue and might therefore impact the results.

Pterostilbene Enhances Endurance Capacity via Promoting Skeletal Muscle Adaptations to Exercise Training in Rats

It has been demonstrated that skeletal muscle adaptions, including muscle fibers transition, angiogenesis, and mitochondrial biogenesis are involved in the regular exercise-induced improvement of endurance capacity and metabolic status. Herein, we investigated the effects of pterostilbene (PST) supplementation on skeletal muscle adaptations to exercise training in rats. Six-week-old male Sprague Dawley rats were randomly divided into a sedentary control group (Sed), an exercise training group (Ex), and exercise training combined with 50 mg/kg PST (Ex + PST) treatment group. After 4 weeks of intervention, an exhaustive running test was performed, and muscle fiber type transformation, angiogenesis, and mitochondrial content in the soleus muscle were measured. Additionally, the effects of PST on muscle fiber transformation, paracrine regulation of angiogenesis, and mitochondrial function were tested in vitro using C2C12 myotubes. In vivo study showed that exercise training resulted in significant increases in time-to-exhaustion, the proportion of slow-twitch fibers, muscular angiogenesis, and mitochondrial biogenesis in rats, and these effects induced by exercise training could be augmented by PST supplementation. Moreover, the in vitro study showed that PST treatment remarkably promoted slow-twitch fibers formation, angiogenic factor expression, and mitochondrial function in C2C12 myotubes. Collectively, our results suggest that PST promotes skeletal muscle adaptations to exercise training thereby enhancing the endurance capacity.

lncRNA-Six1 Is a Target of miR-1611 that Functions as a ceRNA to Regulate Six1 Protein Expression and Fiber Type Switching in Chicken Myogenesis

Emerging studies indicate important roles for non-coding RNAs (ncRNAs) as essential regulators in myogenesis, but relatively less is known about their function. In our previous study, we found that lncRNA-Six1 can regulate Six1 in cis to participate in myogenesis. Here, we studied a microRNA (miRNA) that is specifically expressed in chickens (miR-1611). Interestingly, miR-1611 was found to contain potential binding sites for both lncRNA-Six1 and Six1, and it can interact with lncRNA-Six1 to regulate Six1 expression. Overexpression of miR-1611 represses the proliferation and differentiation of myoblasts. Moreover, miR-1611 is highly expressed in slow-twitch fibers, and it drives the transformation of fast-twitch muscle fibers to slow-twitch muscle fibers. Together, these data demonstrate that miR-1611 can mediate the regulation of Six1 by lncRNA-Six1, thereby affecting proliferation and differentiation of myoblasts and transformation of muscle fiber types.

Erratum: Polge, C., et al. UBE2E1 Is Preferentially Expressed in the Cytoplasm of Slow-Twitch Fibers and Protects Skeletal Muscles from Exacerbated Atrophy upon Dexamethasone Treatment. Cells 2018, 7, 214

Change in author names (order). [...]

Myofibrillar regulatory mechanisms of stretch activation in mammalian striated muscle

Stretch activation is described as a delayed increase in force after an imposed stretch. This process is essential in the flight muscles of many insects and is also observed, to some degree, in mammalian striated muscles. The mechanistic basis for stretch activation remains uncertain, although it appears to involve cooperative activation of the thin filaments (12, 80). The purpose of this study was to address myofibrillar regulatory mechanisms of stretch activation in mammalian striated muscle. For these studies, permeabilized rat slow-twitch and fast-twitch skeletal muscle fibers were mounted between a force transducer and motor, and a slack-re-stretch maneuver was performed over a range of Ca[superscript 2+] activation levels. Following slack-re-stretch there was a stretch activation process that often resulted in a transient force overshoot (P[subscript TO]), which was quantified relative to steady-state isometric force. P[subscript TO] was highly dependent upon Ca[superscript 2+] activation level, and the relative magnitude of P[subscript TO] was greater in slow-twitch fibers than fast-twitch fibers. In both slow-twitch and fast-twitch fibers, force redevelopment involved a fast, Ca[superscript 2+] activation dependent process (k1) and a slower, less activation dependent process (k2). Interestingly, the two processes converged at low levels of Ca[superscript 2+] activation in both fiber types. P[subscript TO] also contained a relaxation phase, which progressively slowed as Ca[superscript 2+] activation levels increased and was more Ca[superscript 2+] activation dependent in slow-twitch fibers. These results suggest that stretch activation may not be solely regulated by the extent of apparent cooperative activation of force due to a higher relative level of stretch activation in the less cooperative slow-twitch skeletal muscle fiber. Next, we investigated an additional potential molecular mechanism by regulating stretch activation in mammalian striated muscle. Along these lines, our lab has previously observed that PKA-induced phosphorylation of cMyBP-C and cTnI elicited a significant increase in transient force overshoot following slack-re-stretch maneuver in permeabilized cardiac myocytes (29). Interestingly, in slow-twitch skeletal muscle fibers MyBP-C but not ssTnI is phosphorylated by PKA (28). We, thus, took advantage of this variation in substrates phosphorylated by PKA to investigate the effects of PKA-induced phosphorylation of MyBP-C on stretch activation in slow-twitch skeletal muscle fibers. Following PKA treatment of skinned slow-twitch skeletal muscle fibers, the magnitude of P[subscript TO] more than doubled, but this only occurred at low levels of Ca[superscript 2+] activation (i.e., [approximately]25% maximal Ca[superscript 2+] activated force). Also, force redevelopment rates were significantly increased over the entire range of Ca[superscript 2+] activation levels following PKA treatment. In a similar manner, force decay rates showed a tendency of being faster following PKA treatment, however, were only statistically significantly faster at 50% Ca[superscript 2+] activation. Overall, these results are consistent with a model whereby stretch transiently increases the number of cross-bridges made available for force generation and PKA phosphorylation of MyBP-C enhances these stretch activation processes.

S-nitrosylation and S-glutathionylation of Cys134 on troponin I have opposing competitive actions on Ca2+ sensitivity in rat fast-twitch muscle fibers

Nitric oxide is generated in skeletal muscle with activity and decreases Ca2+ sensitivity of the contractile apparatus, putatively by S-nitrosylation of an unidentified protein. We investigated the mechanistic basis of this effect and its relationship to the oxidation-induced increase in Ca2+ sensitivity in mammalian fast-twitch (FT) fibers mediated by S-glutathionylation of Cys134 on fast troponin I (TnIf). Force-[Ca2+] characteristics of the contractile apparatus in mechanically skinned fibers were assessed by direct activation with heavily Ca2+-buffered solutions. Treatment with S-nitrosylating agents, S-nitrosoglutathione (GSNO) or S-nitroso- N-acetyl-penicillamine (SNAP), decreased pCa50 ( = −log10 [Ca2+] at half-maximal activation) by ~−0.07 pCa units in rat and human FT fibers without affecting maximum force, but had no effect on rat and human slow-twitch fibers or toad or chicken FT fibers, which all lack Cys134. The Ca2+ sensitivity decrease was 1) fully reversed with dithiothreitol or reduced glutathione, 2) at least partially reversed with ascorbate, indicative of involvement of S-nitrosylation, and 3) irreversibly blocked by low concentration of the alkylating agent, N-ethylmaleimide (NEM). The biotin-switch assay showed that both GSNO and SNAP treatments caused S-nitrosylation of TnIf. S-glutathionylation pretreatment blocked the effects of S-nitrosylation on Ca2+ sensitivity, and vice-versa. S-nitrosylation pretreatment prevented NEM from irreversibly blocking S-glutathionylation of TnIf and its effects on Ca2+ sensitivity, and likewise S-glutathionylation pretreatment prevented NEM block of S-nitrosylation. Following substitution of TnIf into rat slow-twitch fibers, S-nitrosylation treatment caused decreased Ca2+ sensitivity. These findings demonstrate that S-nitrosylation and S-glutathionylation exert opposing effects on Ca2+ sensitivity in mammalian FT muscle fibers, mediated by competitive actions on Cys134 of TnIf.