Myosin and actin expression and oxidation in aging muscle

Thompson, LaDora V.; Durand, Delphine; Fugere, Nicole A.; Ferrington, D.A.

doi:10.1016/j.exger.2006.06.028

Cited by 24 publications

(40 citation statements)

References 37 publications

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“…The overall strategy to identify proteins is the same as described previously (11,49). Briefly, we matched the individual bands on Western blots that showed HU-related changes in intensity of immune reaction with the bands on the silver-stained gels.…”

Section: Identification Of Proteinsmentioning

confidence: 99%

“…The muscle proteins within the soleus were separated experimentally into two fractions, soluble and myofibrillar protein fractions (49). Subsequent individual protein separation (SDS-PAGE), extent of oxidized proteins (Western blotting), and protein identification (mass spectroscopy) are facilitated when the muscles proteins are subfractionated (49).…”

Section: Overall Experimental Strategy To Determine Oxidized Proteinsmentioning

confidence: 99%

“…Buffer containing 2% 3-[(3-chloamidoprophyl)dimethylammonio]-2-hydroxy-1-propanesulfonate and 4 M urea was added into the collected supernatant to ensure complete protein solubilization and prevent aggregation. This fraction of homogenate is called the soluble fraction (49). The pellet was homogenized in buffer containing 10 mM tris(2-carboxyethyl)phosphine and 10% trifluoroacetic acid.…”

Section: Tissue Preparationmentioning

confidence: 99%

“…Muscle tissue was prepared as described (49). Briefly, the rats were anesthetized with pentobarbital sodium (35 mg/kg body wt) after the intervention.…”

Section: Tissue Preparationmentioning

confidence: 99%

“…Membranes probed with HNE antibody and NT antibody were then incubated with biotinylated goat anti-rabbit secondary antibody (Bio-Rad; 1:3,000) at room temperature for 1 h. After the incubation with secondary antibodies, membranes probed with either HNE or NT antibody were incubated with a streptavidin signal amplification solution (Sigma, St. Louis, MO; 1:3,000) and biotinylated alkaline phosphatase (Bio-Rad; 1:3,000) at room temperature for 1.5 h. Finally, substrate 5-bromo-4-chlor-3Ј-iodolylphosphate p-toluidine/nitro blue tetrazolium chloride (BCIP-NBT) was used to visualize the immunoreaction on membranes. Membranes were imaged using a GS-800 calibrated densitometer (Bio-Rad), and intensity (slot area and whole lane for slot blot and SDS-PAGE, respectively) was quantified by densitometry (optical density) using Sigma Scan Pro (Systat, Point Richmond, CA) (11,49).…”

Section: Evaluation Of the Global Accumulation Of Oxidized Proteinsmentioning

confidence: 99%

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Carbonic anhydrase III and four-and-a-half LIM protein 1 are preferentially oxidized with muscle unloading

Chen¹,

Ferrington²,

Thompson³

2008

Journal of Applied Physiology

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The identities of proteins that show disuse-related changes in the content of oxidative modification are unknown. Furthermore, it is unknown whether the global accumulation of oxidized proteins is greater in aged animals with muscle disuse. The purposes of this study are 1) to identify the exact proteins that show disuse-related changes in oxidation levels and 2) to test the hypothesis that the global accumulation of oxidized proteins with muscle disuse would be greater in aged animals. Adult and old rats were randomized into four groups: weight bearing and 3, 7, or 14 days of hindlimb unloading. Soleus muscles were harvested to investigate the protein oxidation with unloading. Slot blot, SDS-PAGE, and Western blot analyses were used to detect the accumulation of 4-hydroxy-2-nonenol (HNE)- and nitrotyrosine (NT)-modified proteins. Matrix-assisted laser desorption ionization-time of flight and tandem mass spectroscopy were used to identify modified proteins. We found that global HNE- and NT-modified proteins accumulated significantly with aging but not with muscle unloading. Two HNE and NT target proteins, four-and-a-half LIM protein 1 (FHL1) and carbonic anhydrase III (CAIII), showed changes in the oxidation levels with muscle unloading. The changes in the oxidation levels happened to adult rats but not old rats. However, old rats had higher baseline levels of HNE-modified FHL1. In summary, the data suggest that the muscle unloading-related changes of protein oxidation are more significant in specific proteins and that the changes are age related.

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Section: Identification Of Proteinsmentioning

confidence: 99%

Section: Overall Experimental Strategy To Determine Oxidized Proteinsmentioning

confidence: 99%

Section: Tissue Preparationmentioning

confidence: 99%

“…Muscle tissue was prepared as described (49). Briefly, the rats were anesthetized with pentobarbital sodium (35 mg/kg body wt) after the intervention.…”

Section: Tissue Preparationmentioning

confidence: 99%

Section: Evaluation Of the Global Accumulation Of Oxidized Proteinsmentioning

confidence: 99%

See 3 more Smart Citations

Carbonic anhydrase III and four-and-a-half LIM protein 1 are preferentially oxidized with muscle unloading

Chen¹,

Ferrington²,

Thompson³

2008

Journal of Applied Physiology

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Oxidative stress‐induced dysregulation of excitation–contraction coupling contributes to muscle weakness

Qaisar

Bhaskaran

Premkumar

et al. 2018

J cachexia sarcopenia muscle

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BackgroundWe have previously shown that the deletion of the superoxide scavenger, CuZn superoxide dismutase, in mice (Sod1 −/− mice) results in increased oxidative stress and an accelerated loss of skeletal muscle mass and force that mirror the changes seen in old control mice. The goal of this study is to define the effect of oxidative stress and ageing on muscle weakness and the Excitation Contraction (EC) coupling machinery in age‐matched adult (8–10 months) wild‐type (WT) and Sod1 −/− mice in comparison with old (25–28 months) WT mice.Methods In vitro contractile assays were used to measure muscle contractile parameters. The activity of the sarcoplasmic reticulum Ca2+ ATPase (SERCA) pump was measured using an NADH‐linked enzyme assay. Immunoblotting and immunofluorescence techniques were used to measure protein expression, and real‐time reverse transcription PCR was used to measure gene expression.ResultsThe specific force generated by the extensor digitorum longus muscle was reduced in the Sod1 −/− and old WT mice compared with young WT mice along with significant prolongation of time to peak force, increased half relaxation time, and disruption of intracellular calcium handling. The maximal activity of the SERCA calcium uptake pump was significantly reduced in gastrocnemius muscle from both old WT (≈14%) and adult Sod1 −/− (≈33%) mice compared with young WT mice along with increased expression of sarcolipin, a known inhibitor of SERCA activity. Protein levels of the voltage sensor and calcium uptake channel proteins dihydropyridine receptor α1 and SERCA2 were significantly elevated (≈45% and ≈57%, respectively), while the ratio of calstabin, a channel stabilizing protein, to ryanodine receptor was significantly reduced (≈21%) in Sod1 −/− mice compared with young WT mice. The changes in calcium handling were accompanied by substantially elevated levels of global protein carbonylation and lipid peroxidation.ConclusionsOur data suggest that the muscle weakness in Sod1 −/− and old WT mice is in part driven by reactive oxygen species‐mediated EC uncoupling and supports a role for reduced SERCA pump activity in compromised muscle function. The novel quantitative mechanistic data provided here can lead to potential therapeutic interventions of SERCA dysfunction for sarcopenia and muscle diseases.

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Fast and slow skeletal myosin binding protein-C and aging

2022

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Aging is associated with skeletal muscle strength decline and cardiac diastolic dysfunction. The structural arrangements of the sarcomeric proteins, such as myosin binding protein-C (MyBP-C) are shown to be pivotal in the pathogenesis of diastolic dysfunction. Yet, the role of fast (fMyBP-C) and slow (sMyBP-C) skeletal muscle MyBP-C remains to be elucidated. Herein, we aimed to characterize MyBP-C and its paralogs in the fast tibialis anterior (TA) muscle from adult and old mice. Immunoreactivity preparations showed that the relative abundance of the fMyBP-C paralog was greater in the TA of both adult and old, but no differences were noted between groups. We further found that the expression level of cardiac myosin binding protein-C (cMyBP-C), an important modulator of cardiac output, was lowered by age. Standard SDS-PAGE along with Pro-Q Diamond phosphoprotein staining did not identify age-related changes in phosphorylated MyBP-C proteins from TA and cardiac muscles; however, it revealed that MyBP-C paralogs in fast skeletal and cardiac muscle were highly phosphorylated. Mass spectrometry further identified glycogen phosphorylase, desmin, actin, troponin T, and myosin regulatory light chain 2 as phosphorylated myofilament proteins in both ages. MyBP-C protein-bound carbonyls were determined using anti-DNP immunostaining and found the carbonyl level of fMyBP-C, sMyBP-C, and cMyBP-C to be similar between old and adult animals. In summary, our data showed some differences regarding the MyBP-C paralog expression and identified an age-related reduction of cMyBP-C expression. Future studies are needed to elucidate which are the age-driven post-translational modifications in the MyBP-C paralogs.

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Myosin and actin expression and oxidation in aging muscle

Cited by 24 publications

References 37 publications

Carbonic anhydrase III and four-and-a-half LIM protein 1 are preferentially oxidized with muscle unloading

Carbonic anhydrase III and four-and-a-half LIM protein 1 are preferentially oxidized with muscle unloading

Oxidative stress‐induced dysregulation of excitation–contraction coupling contributes to muscle weakness

Fast and slow skeletal myosin binding protein-C and aging

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