People worldwide live longer than ever. By 2030, 1 in 6 people will be aged 60 years or over and by 2050, the number of persons aged 80 years and older will triple compared to 2020 (World Health Organization, WHO). While the proportion of older persons in the population is increasing, the number of years one can expect to enjoy good health is changing relatively little, implying that the years gained in lifespan are in poor health. As the WHO declared: “adding more years to life can be a mixed blessing if it is not accompanied by adding more life to years” 1.

The process of becoming older, known as aging, refers to a progressive decline in physical and mental capacity. Aging is considered a risk factor for many diseases, with muscle function being an important aspect 2,3. A well-functioning muscular system is fundamental to healthy aging. Thus, understanding how muscles change with age is crucial to design treatments to counteract muscle aging and loss of muscle function. 

A variety of interventions have been shown to counteract aging-associated changes and extend healthspan. Perhaps the best known ones are calorie restriction (CR) and the administration of rapamycin (RM), a drug that inhibits cellular growth by acting on a protein complex known as TORC1 (mTORC1 in mammals) 4. In the past, we showed that long-term CR and RM treatments have beneficial and partially overlapping effects on the transcriptomes of the aging muscles 5,6. However, how the long-term treatments end up changing the gene expression programs of various organs, including of various types of muscles, remained unclear. 

In this study 7, we investigated this aspect by determining the effect of long-term CR and RM treatments on protein phosphorylation-dependent signaling pathways. More specifically, we obtained phosphoproteome data from four different muscles (soleus, tibialis anterior, gastrocnemius, and triceps brachii) of four distinct cohorts of mice: 10 months old adult mice and 30-month-old mice that received from 15 months of age either a control, calorie restricted or RM containing-diet (Figure 1A, B). We were able to identify about 7’000 phosphorylation sites, approximately 1,400 of which are not yet found in standard repositories of the field. Our analysis revealed that both CR and RM partially counteract aging-induced changes in protein phosphorylation, with distinct quantitative effects. While RM-associated changes are focused on the mTORC1 pathway, a broader spectrum of cellular processes respond to CR. A further analysis on kinase signatures revealed that aging and the treatments differ in their effects on individual kinase activities and in individual muscles (Figure 1C), emphasizing the complexity of counteracting aging-associated changes at the organism level, especially with small molecules. 

Altogether, our analysis provides a broad view of the impact of aging and long-term anti-aging interventions on the phosphoproteome landscape of mice muscles. Moreover, the comprehensive muscle-specific dataset we provide may be of interest to a large community of scientists who study aging processes and aim to improve muscle functionality in aging. 

References

1. World Health Organization. Decade of Healthy Ageing: Baseline Report. (World Health Organization, 2021).
2. Niccoli, T. & Partridge, L. Ageing as a risk factor for disease. Curr. Biol. 22, R741–52 (2012).
3. Siparsky, P. N., Kirkendall, D. T. & Garrett, W. E., Jr. Muscle changes in aging: understanding sarcopenia. Sports Health 6, 36–40 (2014).
4. Battaglioni, S., Benjamin, D., Wälchli, M., Maier, T. & Hall, M. N. mTOR substrate phosphorylation in growth control. Cell 185, 1814–1836 (2022).
5. Ham, D. J. et al. Distinct and additive effects of calorie restriction and rapamycin in aging skeletal muscle. Nat. Commun. 13, 2025 (2022).
6. Ham, D. J. et al. The neuromuscular junction is a focal point of mTORC1 signaling in sarcopenia. Nat. Commun. 11, 4510 (2020).
7. Ataman, M. et al. Calorie restriction and rapamycin distinctly mitigate aging-associated protein phosphorylation changes in mouse muscles. Commun Biol 7, 974 (2024).