Fig 2: SAM binds and stabilizes Sugt1 in myoblasts.a Left: Schematic of RNA pull-down assay. Right: The band (framed in the yellow box) was extracted for mass spectrometry (MS) analysis. b The partial list of proteins identified by MS. c Western blot (WB) confirmed the association of SAM with Sugt1 but not Hnrnpl, Dnmt3a, and Dnmt3b proteins. 30 µg (or 1.5%) of cell lysate was used as input. d Left: Schematic of native RNA immunoprecipitation (RIP) assay. Right. WB analysis of Sugt1 protein after IP with Sugt1 antibody. qRT-PCR detection of the retrieved RNAs. e The co-localization of Flag-Sugt1 and SAM by FISH coupled with IF. f Expression of Sugt1 in FISCs, ASCs, and DSCs. g Sugt1 was knocked down in ASCs by two different siRNA oligos. h EdU labeling in the above cells. The percentage of EdU+ cells was quantified. i EdU labeling in ASC transfected with a Vector or Sugt1 expressing plasmid. The percentage of EdU+ SCs was quantified. j Expression of Sugt1 in ASCs from WT vs. KO mice. k Sugt1 levels in WT and KO ASCs treated with cycloheximide (CHX) for 10 h. l Sugt1 in myoblasts transfected with Ctrl or shSAM oligos and treated with CHX for the indicated time. The degradation rates of Sugt1 are shown on the right. m Sugt1 levels in cells treated with CHX or/and MG132 for 12 h. n WB analysis of Sugt1 ubiquitination after IP with Sugt1 antibody in cells transfected with HA-Ub and Sugt1 expressing plasmids. o EdU labeling in WT and KO ASC transfected with a plasmid expressing SUGT1 wild type (WT), or stabilized SUGT1-4A, or unstable SUGT1-4E mutant. The percentage of EdU+ cells was quantified. The data are presented as mean ± SD in d, f–j, and o. The p values by two-tailed unpaired t test are indicated in d, f–j, and o, ns not significant. The total number of biologically independent samples are indicated in d, f–j, and o. Scale bars: 10 µm e, 100 µm h, i, and o. Source data are provided as a Source Data file.

Fig 3: Schematic model depicting the functional role of SAM during SC activation/proliferation.In WT mice, SAM regulates SCs proliferation by binding with co-chaperon protein Sugt1 to facilitate the kinetochore assembly during mitosis, thereby governing the fidelity of cell division. In KO mice, loss of SAM induces degradation of Sugt1 by ubiquitination then disrupts kinetochore assembly in mitotic cells due to mislocalization of Dsn1 and Hec1, delays proliferation and impairs muscle regeneration after acute or chronic muscle injuries.