Biology subjects

Stead, C. A.

Publications and source records attributed to Stead, C. A..

6 recordsLinked to original sources

bioRxiv2026

Dynamic proteomic profiling reveals protein-specific regulation of synthesis rates underpinning the divergent adaptation of human muscle to endurance and resistance training

Endurance (END) or resistance exercise (RE) training results in adaptations that give rise to distinct skeletal muscle phenotypes. Hallmarks of RE include increases in muscle fibre and muscle cross-sectional area and strength, whereas END increases mitochondrial content. Such distinct phenotypes arise from differential metabolic and mechanical signal transduction, transcriptional, and protein translation pathways, culminating in exercise mode-specific adaptations in the muscle proteome. However, little empirical data exist on the protein-specific dynamic responses underlying training-mode-specific adaptations in humans. Using a model of unilateral exercise combined with stable isotope labelling with deuterium oxide, we measured changes in synthesis and abundance from baseline and during early (week 1) and later (week 10) periods of adaptation to END and RE training in young healthy adults (n = 14; 8 female, 6 male; 20 {+/-} 1 y, 70 {+/-} 10 kg). We quantified changes in the abundance (n = 1146 proteins) and synthesis (n = 247 proteins) profiles of skeletal muscle across a 5-day pre-training baseline period and during early and later adaptation to RE and END. Abundance profiling revealed mode-specific proteome remodelling, whereby RE increased ribosomal and contractile protein networks, whereas END increased mitochondrial inner membrane proteins after 10 weeks of training. The protein-specific synthesis rates of 119 proteins showed training-induced differences (P < 0.1 and log2 fold change > 1), including subsets of structural proteins that responded differently to RE and END training modes. Notably, distinct Z-disc proteins, such as XIRP1 (RE-specific) and LDB3 (END-specific), exhibited mode-specific regulation despite sharing a similar subcellular localisation. We report, for the first time, that divergent phenotypic adaptations to RE and END extend beyond changes in bulk fraction-specific synthesis rates and are regulated by training-mode-specific adaptations in distinct protein subsets within similar subcellular protein locations.

physiology↗

bioRxiv2025

Dynamic proteome profiling uncovers age-related impairments in proteostasis and the protective effects of resistance exercise in human skeletal muscle

A loss of proteostasis is a primary hallmark of ageing that has emerged from mechanistic studies in model organisms, but little is currently known about changes to proteostasis in the muscle of older humans. We used stable isotope labelling (deuterium oxide; D2O) in vivo, and peptide mass spectrometry of muscle samples to investigate differences in proteome dynamics between the muscle of younger (28 {+/-} 5 y; n=4) and older (69 {+/-} 3 y; n=4) men during either habitual activity or resistance exercise training. We quantified the abundance of 1787 proteins and the turnover rate of 1046 proteins in bi-lateral samples of vastus lateralis (n=32 samples total) taken before and after a 15-day program including 5 sessions of unilateral leg-press exercise (3 sets of 10 repetitions at 90% of 10 RM). Our protein abundance profiling revealed a stoichiometric imbalance within the proteostasis network in aged skeletal muscle, including subunits of eIF3, subunits of 40S and 60S ribosomal proteins. The rate of bulk, mixed-protein synthesis was not different between younger and older men, but most ribosomal proteins were less abundant in the muscle of older participants, suggesting ribosomes in older muscle may exhibit increased translational efficiency to maintain similar levels of protein turnover compared to ribosomes in younger muscle. Resistance exercise partially restored age-related disruptions to the proteostasis network. In older skeletal muscle, resistance exercise specifically increased the absolute turnover rate (ATR) of mixed mitochondrial proteins, with increased fractional turnover rate (FTR) of prohibitin 1 (PHB1) and profilin-1 (PROF1), and increased abundance of prohibitin 2 (PHB2). These adaptations may suggest resistance exercise promotes mitochondrial proteostasis by facilitating the synthesis and maintenance of key mitochondrial proteins. Thus, our Dynamic Proteome Profiling data provide an impetus for further exploration of the role of proteostasis in maintaining skeletal muscle quality and supports resistance exercise as a potential therapeutic strategy to promote healthy skeletal muscle ageing in humans. In BriefNishimura et al. used Dynamic Proteome Profiling to uncover whether the distorted proteomic landscape of ageing skeletal muscle is associated with altered turnover of specific proteins. Basal muscle from older men exhibits a divergence in protein abundance between subunits of eIF3 and subunits of 40S and 60S ribosomal proteins, whereas resistance exercise partially restored age-related disruptions in the muscle proteome. In older muscle, protein-specific turnover generally increases after resistance exercise, independent of changes in protein abundance, suggesting improved protein quality and renewal. Created in BioRender. Nishimura, Y. (2025) https://BioRender.com/p2a1aio HighlightsO_LIDynamic Proteome Profiling in human skeletal muscle ageing C_LIO_LIAgeing alters muscle proteostasis C_LIO_LIMixed-muscle protein synthesis does not differ between younger and older men C_LIO_LIResistance exercise increased mitochondrial protein turnover specifically in older muscle C_LIO_LIProtein-specific responses to resistance exercise differed between age groups C_LI O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=200 SRC="FIGDIR/small/684531v1_ufig1.gif" ALT="Figure 1"> View larger version (48K): org.highwire.dtl.DTLVardef@11d0c8corg.highwire.dtl.DTLVardef@22f451org.highwire.dtl.DTLVardef@d62118org.highwire.dtl.DTLVardef@16eda9_HPS_FORMAT_FIGEXP M_FIG C_FIG

physiology↗

bioRxiv2025

Adaptive time course of the skeletal muscle proteome during programmed resistance training in rats

Resistance training (RT) promotes muscle protein accretion and myofiber hypertrophy, driven by dynamic processes of protein synthesis and degradation. While molecular studies have focused on acute signalling or long-term hypertrophy and strength gains, a critical gap remains in understanding the intermediate processes of muscle adaptation. Acute signalling does not always correlate directly with long-term outcomes, highlighting the need for a time-course analysis of protein abundance and turnover rates. To address this, we utilised deuterium oxide labelling and peptide mass spectrometry to quantify absolute protein content and synthesis rates in skeletal muscle. A daily programmed resistance training regimen was applied to the rat tibialis anterior (TA) via electrical stimulation of the left hind limb for 10, 20, and 30 days (5 sets of 10 repetitions daily). Muscle samples from stimulated (Stim) and contralateral control (Ctrl) limbs were analysed, quantifying 658 protein abundances and 215 protein synthesis rates. Unsupervised temporal clustering of protein responses revealed distinct phases of muscle adaptation, with early (0-10 days) and mid (10-20 days) responses driven by differential protein accretion rates in ribosomal and mitochondrial networks, respectively. These findings suggest that subsets of proteins exhibit distinct adaptation timelines due to variations in translation and/or degradation rates. A deeper understanding of these temporal shifts could improve strategies for optimising muscle growth and functional adaptation to resistance training.

physiology↗

bioRxiv2024

Turnover rates of human muscle proteins in vivo reported in fractional, mole and absolute units.

Protein fractional turnover rates (FTR) represent measurements of flux through a protein pool, i.e. net abundance (ABD) of the protein. If protein abundance is not measured or is different between experimental conditions the interpretation of FTR data may be confounded. This project investigates the consequences of reporting turnover rates of human muscle proteins in vivo in mole and absolute units (that incorporate protein abundance data) compared to fractional (%/d) data that ignore protein abundance. Three physically active males (21 {+/-} 1 years) were recruited and underwent a 12-d protocol of daily deuterium oxide (D2O) consumption and biopsies of vastus lateralis on days 8 and 12. Protein abundances were normalised to yeast alcohol dehydrogenase, added during sample preparation, and FTR was calculated from time-dependent changes in peptide mass isotopomer profiles. FTR and abundance data (fmol/ g protein) were combined to calculate mole turnover rates (MTR; fmol/ g protein/ d) and absolute turnover rates (ATR; ng/ g protein/ d). Abundance data were collected for 1,772 proteins and FTR data were calculated from 3,944 peptides representing 935 proteins (average 3 peptides per protein). The median (M), lower- (Q1) and upper-quartile (Q3) values for protein FTR (%/d) were M = 4.3, Q1 = 2.52, Q3 = 7.84. Our analyses suggest MTR data is preferred over FTR, particularly for studies on multiprotein complexes, wherein MTR takes account of potential differences amongst the molecular weight of the component subunits. ATR data may be preferred over MTR and FTR, particularly when comparing samples with different abundance profiles.

systems biology↗

bioRxiv2023

Muscle of obese insulin-resistant humans exhibits losses in proteostasis and attenuated proteome dynamics that are improved by exercise training.

We examined muscle proteostasis in obese insulin-resistant (OIR) individuals to determine whether endurance exercise could positively influence proteome dynamics in this population. Male OIR (n = 3) and lean, healthy controls (LHC; n = 4) were recruited and underwent a 14-d measurement protocol of daily deuterium oxide (D2O) consumption and serial biopsies of vastus lateralis muscle. The OIR group then completed 10-weeks of high-intensity interval training (HIIT), encompassing 3 sessions per week of cycle ergometer exercise with 1 min intervals at 100 % maximum aerobic power (Wmax) interspersed by 1 min recovery periods. The number of intervals per session progressed from 4 to 8, and during weeks 8-10 the 14-d measurement protocol was repeated. The abundance and turnover rates of 880 and 301 proteins, respectively, were measured. OIR and LHC muscle exhibited 352 differences (p < 0.05, false discovery rate (p < 0.05) differences in protein turnover. OIR muscle was enriched with markers of metabolic stress, protein misfolding and components of the ubiquitin-proteasome system, and the turnover rate of many of these proteins was less compared to LHC muscle. HIIT altered the abundance of 53 proteins and increased the turnover rate of 22 proteins (p < 0.05) in OIR muscle and tended to restore proteostasis, evidenced by increasing muscle protein turnover rates and normalizing proteasome composition in OIR participants. In conclusion, obesity and insulin resistance are associated with compromised muscle proteostasis, which can be partially restored by endurance exercise.

physiology↗

bioRxiv2022

Facioscapulohumeral muscular dystrophy is associated with altered myoblast proteome dynamics

Proteomic studies in facioscapulohumeral muscular dystrophy (FSHD) could offer new insight to disease mechanisms underpinned by post-transcriptional processes. We used stable isotope (deuterium oxide; D2O) labelling and peptide mass spectrometry to investigate the abundance and turnover rates of proteins in cultured muscle cells from 2 individuals affected by FSHD and their unaffected siblings (UASb). We measured the abundance of 4485 proteins and the turnover rate of 2324 proteins in each (n = 4) myoblast sample. FSHD myoblasts exhibited a greater abundance but slower turnover rate of subunits of mitochondrial respiratory complexes and mitochondrial ribosomal proteins, which may indicate an accumulation of older less viable mitochondrial proteins in myoblasts from individuals affected by FSHD. Our results highlight the importance of post-transcriptional processes and protein turnover in FSHD pathology and provide a resource for the FSHD research community to explore this burgeoning aspect of FSHD.

cell biology↗