Antagonistic control of myofiber size and muscle protein quality control by the ubiquitin ligase UBR4 during aging

Abstract

Sarcopenia is a degenerative condition that consists in age-induced atrophy and functional decline of skeletal muscle cells (myofibers). A common hypothesis is that inducing myofiber hypertrophy should also reinstate myofiber contractile function but such model has not been extensively tested. Here, we find that the levels of the ubiquitin ligase UBR4 increase in skeletal muscle with aging, and that UBR4 increases the proteolytic activity of the proteasome. Importantly, muscle-specific UBR4 loss rescues age-associated myofiber atrophy in mice. However, UBR4 loss reduces the muscle specific force and accelerates the decline in muscle protein quality that occurs with aging in mice. Similarly, hypertrophic signaling induced via muscle-specific loss of UBR4/poe and of ESCRT members (HGS/Hrs, STAM, USP8) that degrade ubiquitinated membrane proteins compromises muscle function and shortens lifespan in Drosophila by reducing protein quality control. Altogether, these findings indicate that these ubiquitin ligases antithetically regulate myofiber size and muscle protein quality control.

Analysis of proteostasis during aging with western blot of detergent-soluble and insoluble protein fractions

“…In Figure 5 , we present an example of changes in proteostasis that occur with aging in mouse skeletal muscles. Specifically, with aging there is an increase in poly-ubiquitinated proteins found in the insoluble fractions of skeletal muscles from 24-month-old mice as compared to muscle from young 6-month-old mice ( Hunt et al., 2021 ), consistent with other studies ( Sakuma et al., 2016 ; White et al., 2016 ).

Figure 5 Decline in protein quality control in skeletal muscles during aging Western blotting of detergent-soluble and insoluble proteins fractions from tibialis anterior and soleus muscles from wild-type mice at 6 months (young) and 24 months (old) of age.

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“…Proteostasis decline with aging and defects in protein quality control are the underlying cause of multiple age-related diseases ( Douglas and Dillin, 2010 ; Labbadia and Morimoto, 2015 ). We have recently found that proteostasis declines during skeletal muscle aging (sarcopenia) in mice ( Demontis et al., 2013 ; Hunt et al., 2021 ; Jiao and Demontis, 2017 ), and that protein quality control is compromised across tissues and organ systems in the fruit fly Drosophila during aging ( Demontis and Perrimon, 2010 ; Rai et al., 2021 ). Moreover, by using thermal stress as a proxy for age-induced challenge to proteostasis, we have investigated the role of genetic and chemical interventions on proteostasis of human brain organoids and HEK293 cells ( Rai et al., 2021 ).…”

“…Tissues from Drosophila and mice of the desired genotypes and ages, and treatment of HKE293 cells and human brain organoids should be done according to the specific experimental requirements and following procedures as described in our and other recent studies ( Hunt et al., 2019a ; Hunt et al., 2021 ; Hunt et al., 2019b ; Hunt et al., 2015 ; Rai et al., 2021 ). Once obtained, these tissues can be examined following the procedures described below.…”

“…For complete details on the use and execution of this protocol, please refer to Rai et al. (2021) and Hunt el al. (2021) .…”

Influence of Age on Skeletal Muscle Hypertrophy and Atrophy Signaling: Established Paradigms and Unexpected Links

“…Atrogenes whose expression is increased in atrophying muscles include ubiquitin, core components of the 26S proteosome, eIF4E inhibitor 4E-BP1 ( Eif4ebp1 ), transcription factors Atf4 , Foxo1 , and Foxo3 [ 194 , 195 ], the muscle specific E3 ubiquitin ligases MurF1 ( Trim63 ) and Atrogin-1/MAFbx ( Fbxo32 ) [ 196 , 197 ], and autophagosome components LC3 ( Map1lc3a ) and Gabarap [ 194 , 198 , 199 ]. Importantly, animal models indicate that both insufficient [ 200 , 201 , 202 , 203 ] and excessive [ 198 , 199 , 204 ] proteolysis, both proteasomal and autophagic, negatively impact the muscle mass and functionality, suggesting an inflection point between a level of proteolysis that supports muscle homeostasis and that which promotes muscle wasting.…”