Identification and characterization of Fbxl22, a novel skeletal muscle atrophy-promoting E3 ubiquitin ligase

Abstract

Muscle-specific E3 ubiquitin ligases have been identified in muscle atrophy-inducing conditions. The purpose of the current study was to explore the functional role of Fbxl22, and a newly identified splice variant (Fbxl22-193), in skeletal muscle homeostasis and neurogenic muscle atrophy. In mouse C2C12 muscle cells, promoter fragments of the Fbxl22 gene were cloned and fused with the secreted alkaline phosphatase reporter gene to assess the transcriptional regulation of Fbxl22. The tibialis anterior muscles of male C57/BL6 mice (12-16 weeks old) were electroporated with expression plasmids containing the cDNA of two Fbxl22 splice variants and tissues collected after 7, 14 and 28 days. Gastrocnemius muscles of wild type and MuRF1 knockout mice were electroporated with an Fbxl22 RNAi or empty plasmid, denervated three days post-transfection, and tissues collected 7 days post-denervation. The full-length gene and novel splice variant are transcriptionally induced early (after 3 days) during neurogenic muscle atrophy. In vivo overexpression of Fbxl22 isoforms in mouse skeletal muscle lead to evidence of myopathy/atrophy suggesting that both are involved in the process of neurogenic muscle atrophy. Knockdown of Fbxl22 in MuRF1 KO muscles resulted in significant additive muscle sparing at 7 days of denervation. Targeting two E3 ubiquitin ligases appears to have a strong additive effect on protecting muscle mass loss with denervation and these findings have important implications in the development of therapeutic strategies to treat muscle atrophy.

Identification of the MuRF1 Skeletal Muscle Ubiquitylome Through Quantitative Proteomics

“…However, recently we showed that transfection of unchallenged TA muscle with Fbxl22, an F-box E3 ligase, resulted in muscle fiber degeneration demonstrating that certain F-box proteins can induce a phenotype when transfected into muscle. 22 Clearly, MAFbx plays an important role in skeletal muscle atrophy, which is distinct from MuRF1. Additional studies are needed to identify the in vivo substrates of MAFbx and to find how ubiquitination of those substrates alters the function and/or structure of skeletal muscle.…”

The Role of Cullin-RING Ligases in Striated Muscle Development, Function, and Disease

“…Recently, the temporal and spatial regulation of murine Fbxl22 was elucidated by Hughes and co-workers for skeletal muscles [126]. Intriguingly, the authors uncovered a novel Fbxl22 splice isoform in muscles.…”

Pectoralis major muscle atrophy is associated with mitochondrial energy wasting in cachectic patients with gastrointestinal cancer

“… 35 It would be interesting to further investigate the proteasome‐mediated degradation pathway through other relevant ubiquitin ligases, as it seems likely that many others, with different substrate‐specificity, are involved in cancer cachexia. Fbxl22 36 and Musa1, 37 for example, would be valuable targets as they have been recently identified in the context of muscle atrophy. Analysis of myostatin‐related and IGF‐related signalling pathways would also be interesting to further improve our understanding on the interplay of molecular mechanisms underlying skeletal muscle wasting during cancer cachexia (refer to review 38 ).…”