Endocrinology Graduate Program, Department of Nutritional Science and Toxicology, University of California, Berkeley, CA 94720, USA.
Proc Natl Acad Sci U S A. 2012 Jul 10;109(28):11160-5. doi: 10.1073/pnas.1111334109. Epub 2012 Jun 25.
Glucocorticoids elicit a variety of biological responses in skeletal muscle, including inhibiting protein synthesis and insulin-stimulated glucose uptake and promoting proteolysis. Thus, excess or chronic glucocorticoid exposure leads to muscle atrophy and insulin resistance. Glucocorticoids propagate their signal mainly through glucocorticoid receptors (GR), which, upon binding to ligands, translocate to the nucleus and bind to genomic glucocorticoid response elements to regulate the transcription of nearby genes. Using a combination of chromatin immunoprecipitation sequencing and microarray analysis, we identified 173 genes in mouse C2C12 myotubes. The mouse genome contains GR-binding regions in or near these genes, and gene expression is regulated by glucocorticoids. Eight of these genes encode proteins known to regulate distinct signaling events in insulin/insulin-like growth factor 1 pathways. We found that overexpression of p85α, one of these eight genes, caused a decrease in C2C12 myotube diameters, mimicking the effect of glucocorticoids. Moreover, reducing p85α expression by RNA interference in C2C12 myotubes significantly compromised the ability of glucocorticoids to inhibit Akt and p70 S6 kinase activity and reduced glucocorticoid induction of insulin receptor substrate 1 phosphorylation at serine 307. This phosphorylation is associated with insulin resistance. Furthermore, decreasing p85α expression abolished glucocorticoid inhibition of protein synthesis and compromised glucocorticoid-induced reduction of cell diameters in C2C12 myotubes. Finally, a glucocorticoid response element was identified in the p85α GR-binding regions. In summary, our studies identified GR-regulated transcriptional networks in myotubes and showed that p85α plays a critical role in glucocorticoid-induced insulin resistance and muscle atrophy in C2C12 myotubes.
糖皮质激素在骨骼肌中引发多种生物学反应，包括抑制蛋白质合成和胰岛素刺激的葡萄糖摄取，促进蛋白水解。因此，糖皮质激素过多或慢性暴露会导致肌肉萎缩和胰岛素抵抗。糖皮质激素主要通过糖皮质激素受体 (GR) 传递信号，GR 与配体结合后易位到细胞核，并与基因组糖皮质激素反应元件结合，以调节附近基因的转录。我们使用染色质免疫沉淀测序和微阵列分析相结合的方法，在小鼠 C2C12 肌管中鉴定出 173 个基因。小鼠基因组中包含这些基因的 GR 结合区域或附近的 GR 结合区域，并且基因表达受糖皮质激素调节。其中 8 个基因编码已知调节胰岛素/胰岛素样生长因子 1 途径中不同信号事件的蛋白质。我们发现，这 8 个基因之一的 p85α 的过表达导致 C2C12 肌管直径减小，模拟了糖皮质激素的作用。此外，在 C2C12 肌管中通过 RNA 干扰降低 p85α 的表达显著削弱了糖皮质激素抑制 Akt 和 p70 S6 激酶活性的能力，并减少了糖皮质激素诱导胰岛素受体底物 1 丝氨酸 307 磷酸化。这种磷酸化与胰岛素抵抗有关。此外，降低 p85α 的表达会消除糖皮质激素对蛋白质合成的抑制作用，并削弱糖皮质激素诱导的 C2C12 肌管细胞直径减小。最后，在 p85α 的 GR 结合区域中鉴定出一个糖皮质激素反应元件。总之，我们的研究在肌管中鉴定出 GR 调节的转录网络，并表明 p85α 在糖皮质激素诱导的 C2C12 肌管胰岛素抵抗和肌肉萎缩中发挥关键作用。
