EIF4A3-Induced Circular RNA circSnd1 Promotes Muscle Atrophy and Muscle Ageing by Stabilizing EEF1A1.

CircSnd1 is upregulated in multiple types of muscle atrophy models both in vivo and in vitro.

• circRNA sequencing and quantitative real-time PCR were used to detect changed circRNAs in muscle atrophy models and aged muscle.
• CircSnd1 upregulation was observed across many types of muscle atrophy models in both in vivo and in vitro conditions.
• All p-values for upregulation across models were < 0.01.
• CircSnd1 is derived from the staphylococcal nuclease and Tudor domain containing 1 gene (Snd1) and is highly species-conserved.

CircSnd1 is elevated in aged muscle in humans, mice, and aged myotubes.

• CircSnd1 was upregulated +2.2-fold in aged human muscle (n = 5, p < 0.05).
• CircSnd1 was elevated +43.96% in aged mouse muscle (n = 6, p < 0.05).
• CircSnd1 was elevated +42.21% in aged myotubes (n = 6, p < 0.05).

CircSnd1 promotes muscle atrophy and muscle ageing at both the cellular and mouse levels, and repressing it ameliorates multiple types of muscle atrophy.

• Gain-of-function and loss-of-function experiments were used to investigate the function of circSnd1 in muscle atrophy and muscle ageing.
• Overexpression of circSnd1 promoted muscle atrophy and ageing phenotypes in cell and mouse models.
• Repression of circSnd1 ameliorated multiple types of muscle atrophy (all p < 0.05).
• CircSnd1 is identified as a novel therapeutic target for muscle atrophy.

EIF4A3 binds to the intron flanking sequence of circSnd1 to induce its cyclization and increase circSnd1 expression in muscle atrophy.

• RIP-MS (RNA immunoprecipitation coupled with mass spectrometry) and RIP assays were used to determine upstream and downstream mechanisms of circSnd1.
• The RNA binding protein eukaryotic translation initiation factor 4A3 (EIF4A3) was identified as the upstream regulator.
• EIF4A3 binds to the intron flanking sequence of circSnd1 to promote its back-splicing/cyclization.
• EIF4A3 binding results in increased circSnd1 expression specifically in the context of muscle atrophy.

CircSnd1 stabilizes EEF1A1 protein levels in muscle cells by promoting FAT10 binding to EEF1A1, which competes with ubiquitin and decreases EEF1A1 ubiquitination.

• CircSnd1 promotes the binding between human HLA-F adjacent transcript 10 (FAT10) and eukaryotic translation elongation factor 1 alpha 1 (EEF1A1).
• FAT10 competes with ubiquitin for binding to EEF1A1.
• This competition decreases the ubiquitination of EEF1A1, thereby stabilizing its protein level in muscle cells.
• Stabilization of EEF1A1 was identified as the downstream mechanism by which circSnd1 promotes muscle atrophy.
• RIP-MS and RIP assays were used to identify these downstream interactions.