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. 2024 Jul 15;38(13):e23797.
doi: 10.1096/fj.202400213RR.

Impaired myoblast differentiation and muscle IGF-1 receptor signaling pathway activation after N-glycosylation inhibition

Giosuè Annibalini  1 , Laura Di Patria  1 , Giacomo Valli  2 , Matteo Bocconcelli  1 , Roberta Saltarelli  1 , Lorenzo Ferri  3 , Laura Barberi  4 , Fabiana Fanelli  1 , Amelia Morrone  3   5 , Rita Barone  6   7 , Renzo Guerrini  3   5 , Antonio Musarò  4 , Vilberto Stocchi  8 , Elena Barbieri  1
Affiliations

Impaired myoblast differentiation and muscle IGF-1 receptor signaling pathway activation after N-glycosylation inhibition

Giosuè Annibalini et al. FASEB J. .

Abstract

The role of N-glycosylation in the myogenic process remains poorly understood. Here, we evaluated the impact of N-glycosylation inhibition by Tunicamycin (TUN) or by phosphomannomutase 2 (PMM2) gene knockdown, which encodes an enzyme essential for catalyzing an early step of the N-glycosylation pathway, on C2C12 myoblast differentiation. The effect of chronic treatment with TUN on tibialis anterior (TA) and extensor digitorum longus (EDL) muscles of WT and MLC/mIgf-1 transgenic mice, which overexpress muscle Igf-1Ea mRNA isoform, was also investigated. TUN-treated and PMM2 knockdown C2C12 cells showed reduced ConA, PHA-L, and AAL lectin binding and increased ER-stress-related gene expression (Chop and Hspa5 mRNAs and s/uXbp1 ratio) compared to controls. Myogenic markers (MyoD, myogenin, and Mrf4 mRNAs and MF20 protein) and myotube formation were reduced in both TUN-treated and PMM2 knockdown C2C12 cells. Body and TA weight of WT and MLC/mIgf-1 mice were not modified by TUN treatment, while lectin binding slightly decreased in the TA muscle of WT (ConA and AAL) and MLC/mIgf-1 (ConA) mice. The ER-stress-related gene expression did not change in the TA muscle of WT and MLC/mIgf-1 mice after TUN treatment. TUN treatment decreased myogenin mRNA and increased atrogen-1 mRNA, particularly in the TA muscle of WT mice. Finally, the IGF-1 production and IGF1R signaling pathways activation were reduced due to N-glycosylation inhibition in TA and EDL muscles. Decreased IGF1R expression was found in TUN-treated C2C12 myoblasts which was associated with lower IGF-1-induced IGF1R, AKT, and ERK1/2 phosphorylation compared to CTR cells. Chronic TUN-challenge models can help to elucidate the molecular mechanisms through which diseases associated with aberrant N-glycosylation, such as Congenital Disorders of Glycosylation (CDG), affect muscle and other tissue functions.

Keywords: PMM2; IGF1R signaling pathway; congenital disorders of glycosylation; glycosylation; muscle atrophy; myoblast differentiation.

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References

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