Novel Sarcopenia-related Alterations in Sarcomeric Protein Post-translational Modifications (PTMs) in Skeletal Muscles Identified by Top-down Proteomics

Abstract

Sarcopenia, the age-related loss of skeletal muscle mass and strength, is a significant cause of morbidity in the elderly and is a major burden on health care systems. Unfortunately, the underlying molecular mechanisms in sarcopenia remain poorly understood. Herein, we utilized top-down proteomics to elucidate sarcopenia-related changes in the fast- and slow-twitch skeletal muscles of aging rats with a focus on the sarcomeric proteome, which includes both myofilament and Z-disc proteins-the proteins that constitute the contractile apparatuses. Top-down quantitative proteomics identified significant changes in the post-translational modifications (PTMs) of critical myofilament proteins in the fast-twitch skeletal muscles of aging rats, in accordance with the vulnerability of fast-twitch muscles to sarcopenia. Surprisingly, age-related alterations in the phosphorylation of Cypher isoforms, proteins that localize to the Z-discs in striated muscles, were also noted in the fast-twitch skeletal muscle of aging rats. This represents the first report of changes in the phosphorylation of Z-disc proteins in skeletal muscle during aging. In addition, increased glutathionylation of slow skeletal troponin I, a novel modification that may help protect against oxidative damage, was observed in slow-twitch skeletal muscles. Furthermore, we have identified and characterized novel muscle type-specific proteoforms of myofilament proteins and Z-disc proteins, including a novel isoform of the Z-disc protein Enigma. The finding that the phosphorylation of Z-disc proteins is altered in response to aging in the fast-twitch skeletal muscles of aging rats opens new avenues for the investigation of the role of Z-discs in age-related muscle dysfunction.

A mini review: Proteomics approaches to understand disused vs. exercised human skeletal muscle

“…Recently, Wei et al (63) reported novel phosphorylation of myofilament proteins, such as cypher, is increased in skeletal muscle of aged rats compared with young rats, by a top-down quantitative proteomics approach that injects intact proteins into the mass spectrometer (63). Given myofilament proteins play a key role in force generation and skeletal muscle function, these results raise the possibility that age-dependent phosphorylation of myofilament proteins could contribute to age-related muscle dysfunction (63). Beyond the effects of aging that can be associated with sarcopenia, Hoffman et al (30) studied global phosphorylation induced by acute exercise in human skeletal muscle.…”

“…A large number of studies have shown that function of proteins and enzymes is regulated by a wide variety of posttranslational modifications, such as phosphorylation, glycosylation, ubiquitination, and acetylation, among others (34). Recently, Wei et al (63) reported novel phosphorylation of myofilament proteins, such as cypher, is increased in skeletal muscle of aged rats compared with young rats, by a top-down quantitative proteomics approach that injects intact proteins into the mass spectrometer (63). Given myofilament proteins play a key role in force generation and skeletal muscle function, these results raise the possibility that age-dependent phosphorylation of myofilament proteins could contribute to age-related muscle dysfunction (63).…”

Simultaneous Quantification of Protein Expression and Modifications by Top-down Targeted Proteomics: A Case of the Sarcomeric Subproteome

“…MS/MS (either online or offline) were performed to identify proteins and subsequently associated PTMs and sequence variations. Offline MS/MS results of rat skeletal muscles have been previously reported (36). Online MS/MS data of sheep cardiac sarcomeric proteins was output as an .msalign file from the DataAnalysis software for protein identification using MS-Alignϩ (37).…”

Expression of SUMO enzymes is fiber type dependent in skeletal muscles and is dysregulated in muscle disuse

“…The ability of skeletal muscles to quickly adapt their functions as a consequence of alterations in posture, training, inactivity, microgravity, or injuries has been studied for decades . While the most relevant effect is a gradual change in the fiber type composition, the molecular mechanisms and events underlying muscle adaptation to different conditions have only started to be elucidated, and recent studies provided a biological link between muscle plasticity and post‐translational modifications PTM(s) on specific proteins …”

“…[1][2][3] While the most relevant effect is a gradual change in the fiber type composition, the molecular mechanisms and events underlying muscle adaptation to different conditions have only started to be elucidated, and recent studies provided a biological link between muscle plasticity and post-translational modifications PTM(s) on specific proteins. 4,5 Post-translational modifications of substrates further modulate and extend the range of possible protein and organs functions by covalently attaching small chemical moieties to selected amino acid residues. More than a 100 different types of PTMs have been identified that affect many aspects of cellular functionalities, including metabolism, signal transduction, protein interaction, and stability.…”