甲状腺素（TMX）是一种选择性雌激素受体调节剂，常用于治疗激素敏感性癌症。然而，TMX在含有雌激素受体的肌肉等合成组织中的作用尚不清楚。我们报道了一种串联质谱标记法，用于研究C2C12细胞中TMX处理后的肌生成，该细胞是一个经充分表征的小鼠体外骨骼肌分化模型。在未经处理的C2C12肌生成中，对超过10,000个蛋白质的纵向分析发现了一组新的，受肌生成调控的1,062个蛋白质。这些蛋白质可分为五个明显的纵向表达趋势，这些趋势与在人类肌细胞中进行类似分析得到的结果具有显著重叠。我们记录到TMX处理后的肌生成中与脂联素信号通路独特相关的功能富集，以及在TMX处理细胞与对照细胞的一个或多个肌生成阶段中差异表达的一组198个蛋白质，其中大多数参与类固醇和脂质代谢。进一步分析将金属硫蛋白1作为TMX处理C2C12肌生成的一个新靶点。最后，我们提供了一个强大的、自我验证的策略，用于分析肌细胞发育过程中体外处理的总蛋白质反应，并可轻松适应于研究其他药物对肌生成的影响。

Tamoxifen (TMX), a selective estrogen receptor modulator, is commonly used in the treatment of hormone-responsive cancers. However, the effects of TMX in anabolic tissues harboring estrogen receptors, such as skeletal muscle, are poorly understood. We report a tandem mass-tag approach to TMX-treated myogenesis in C2C12 cells, a well-characterized model of in vitro murine skeletal muscle differentiation. A longitudinal analysis of >10,000 proteins identified in untreated C2C12 myogenesis revealed a novel subset of 1,062 myogenically regulated proteins. These proteins clustered into five distinct longitudinal expression trends which significantly overlap those obtained in similar analyses performed in human myocytes. We document a specific functional enrichment for adiponectin-signaling unique to TMX-treated myogenesis, as well as a subset of 198 proteins that are differentially expressed in TMX-treated cells relative to controls at one or more stages of myogenesis, the majority of which were involved in steroid and lipid metabolism. Further analysis highlights metallothionein-1 as a novel target of TMX treatment at each stage of C2C12 myogenesis. Finally, we present a powerful, self-validating pipeline for analyzing the total proteomic response to in vitro treatment across every stage of muscle cell development which can be easily adapted to study the effects of other drugs on myogenesis.