Proteomic profiling of skeletal muscle ribosomes from higher versus lower responders to 10 weeks of resistance training

Michel, J. M.; Norton, S. C.; Bergamasco, J.; Scarpelli, M.; Chaves, T.; da Silva, D.; Bittencourt, D.; Ugochukwu, C.; Boersma, M.; Mobley, C. B.; Godwin, J.; Nader, G.; Libardi, C.; Roberts, M.

doi:10.1101/2025.11.04.686603

bioRxiv · 10.1101/2025.11.04.686603

Proteomic profiling of skeletal muscle ribosomes from higher versus lower responders to 10 weeks of resistance training

Michel, J. M.·Norton, S. C.·Bergamasco, J.·Scarpelli, M.·Chaves, T.·da Silva, D.·Bittencourt, D.·Ugochukwu, C.·Boersma, M.·Mobley, C. B.·Godwin, J.·Nader, G.·Libardi, C.·Roberts, M.

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Abstract

Ribosome biogenesis is a key driver of resistance training (RT)-induced skeletal muscle hypertrophy in humans. However, high resolution insight into RT-induced compositional alterations in ribosomes remain unexplored. Therefore, the purpose of this study was twofold: 1) develop a protocol sufficient to enrich and examine the ribosomal proteome from a small amount of human muscle tissue, and 2) determine if ribosomal protein composition differs between higher and lower responders to RT. Fourteen participants completed 10 weeks of RT (23 sessions) and were stratified into higher (n=7) and lower (n=7) responders based on changes in vastus lateralis muscle cross-sectional area (VL mCSA) and mean myofiber cross-sectional area (fCSA) from baseline (PRE) to after the 23rd RT session (POST). Participants then performed a twenty fourth RT session and biopsies were collected 24 hours post-RT (POST-24h) to examine acute bout (POST to POST-24h) ribosomal proteome alterations to RT. Ribosome enrichment and analysis included ultracentrifugation (100,000 g, 3h, 2{degrees}C) through 20% sucrose gradients followed by shotgun proteomics to quantify ribosomal protein composition. Our protocol produced exceptional ribosome enrichment, with 74 distinct ribosomal proteins being detected (92% coverage of the putative 80 cytosolic ribosomal proteins) as well as [~]164-fold and [~]71-fold increases in large (RPL) and small (RPS) protein subunit abundances, respectively, compared to conventional tissue homogenization. Despite robust phenotypic differences in hypertrophy between responder groups to the 10-week RT protocol (mCSA: +31% versus +3%; fCSA: +29% versus -2%, P<0.05 for both outcomes), ribosomal protein composition was not significantly different between groups (higher vs. lower responders), nor was there a significant time (POST to POST-24h) or a group*time interaction (P>0.028 for all comparisons). In conclusion, we present a highly effective ribosome enrichment protocol requiring minimal human muscle tissue. Though preliminary analyses indicate that ribosomal protein composition was similar between hypertrophic phenotypes in response to an acute RT session after a 10-week RT intervention, future research leveraging our techniques with more sampling time points are needed to provide more definitive conclusions.

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Michel, J. M., Norton, S. C., Bergamasco, J., Scarpelli, M., Chaves, T., da Silva, D., Bittencourt, D., Ugochukwu, C., Boersma, M., Mobley, C. B., Godwin, J., Nader, G., Libardi, C., Roberts, M.. 2025-11-05. Proteomic profiling of skeletal muscle ribosomes from higher versus lower responders to 10 weeks of resistance training. https://doi.org/10.1101/2025.11.04.686603

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