贝肯1
自噬
C2C12型
细胞生物学
肌发生
细胞分化
死孢子体1
程序性细胞死亡
生物
细胞凋亡
心肌细胞
分子生物学
化学
生物化学
基因
作者
Elliott M. McMillan,Joe Quadrilatero
摘要
Several degradative systems assist in formation of multinucleated terminally differentiated myotubes. However, the role of autophagy in this process has not been examined. GFP-LC3B (light chain 3 beta) puncta, LC3B-II protein and LysoTracker fluorescence increased during C2C12 cell differentiation. Importantly, accumulation of LC3B-II protein occurred in CQ (chloroquine)-treated cells throughout differentiation. Furthermore, BECN1 (beclin 1), ATG7 (autophagy-related 7) and ATG12-5 protein increased, whereas SQSTM1/p62 (sequestosome 1) protein was rapidly reduced during differentiation. A transient decrease in BECN1-BCL2 association was observed from day 0.5 to 2 of differentiation. Chemical inhibition of JNK (c-Jun N-terminal kinase) during differentiation reduced LC3B-II protein and GFP-LC3B puncta and maintained BECN1-BCL2 association. Inhibition of autophagy by 3MA (3-methyladenine) or shRNA against Atg7 (shAtg7) resulted in lower myosin heavy chain expression, as well as impaired myoblast fusion and differentiation. Interestingly, 3MA treatment during differentiation increased transient CASP3 (caspase 3) activation, DNA fragmentation and the percentage of apoptotic nuclei. Similarly, shAtg7 cells had increased DNA fragmentation during differentiation compared with the controls. Collectively, these data demonstrate that autophagy increases and is required during myoblast differentiation. Moreover, autophagy protects differentiating myoblasts from apoptotic cell death.
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