Myoblast Phosphoproteomics as a Tool to Investigate Global Signaling Events During Myogenesis

Jones, Fiona K.; Hardman, Gemma; Ferries, Samantha; Eyers, Patrick A.; Pisconti, Addolorata

doi:10.1007/978-1-4939-8897-6_18

Cited by 3 publications

(2 citation statements)

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“…We profiled phosphopeptide abundance in Ctrl and S3 kd myoblasts and studied how they change in response to SDC3 depletion and/or serum stimulation, with the aim of identifying candidate signaling pathways to then validate in primary MuSC progeny isolated from wild type and Sdc3 -/- mice. Following phosphopeptide enrichment on titanium dioxide (TiO 2 ) columns and label free LC-MS/MS (Figure 2A-C and Jones et al (2019) we identified and quantified changes in serine, threonine and tyrosine phosphorylation dependent on either SDC3 depletion (comparisons 1 and 2 in Figure 2C), serum stimulation (comparisons 3 and 4 in Figure 2C) or both combined in a two-dimensional analysis (comparison 5 in Figure 2C). We identified a total of 11,777 peptides of which 7,190 were phosphopeptides, corresponding to 1,871 phosphoproteins.…”

Section: Resultsmentioning

confidence: 99%

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SDC3 acts as a timekeeper of myogenic differentiation by regulating the insulin/AKT/mTOR axis in muscle stem cell progeny

Jones

Pisconti

2020

Preprint

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Muscle stem cells (MuSCs) are indispensable for muscle regeneration. A multitude of extracellular stimuli affect MuSCs from quiescent progenitors to differentiated myocytes, the activity of which is modulated by coreceptors such as syndecan-3 (SDC3). Here we investigated the global landscape of SDC3-mediated regulation of myogenesis using a phosphoproteomics approach which revealed, with the precision level of individual phosphosites, the large-scale extent of SDC3-mediated regulation of MuSC signal transduction. We then focused on INSR/AKT/mTOR as a key pathway regulated by SDC3 during myogenesis and mechanistically dissected SDC3-mediated inhibition of insulin signalling in MuSCs. SDC3 interacts with INSR limiting insulin signal transduction via INSR/AKT/mTOR. Both knockdown of INSR and inhibition of AKT rescue Sdc3-/- MuSC differentiation to wild type levels. Since SDC3 is rapidly downregulated at the onset of differentiation, our study suggests that SDC3 acts a timekeeper to maintain proliferating MuSC response to insulin to low levels and prevent premature differentiation.

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Section: Resultsmentioning

confidence: 99%

“…Lysates were prepared as previously described (Jones et al, 2019). Ctrl and S3 kd myoblasts were cultured in growth medium for two days and for each condition four biological replicates were used.…”

Section: Phosphoproteomics Sample Preparation and Lc-ms/msmentioning

confidence: 99%