蛋白质肉豆寇酰化修饰及内质网蛋白动态平衡介导脂肪酸对肌小节的影响及机制研究

Fatty acid (FA) metabolism has profound impact on tissue maintenance and animal development. Abnormal FA metabolism has been shown to be critically involved in muscle degenerative diseases, but the underlying mechanisms remain to be understood. Our preliminary results indicated reduction of overall FA levels by blocking FA biosynthesis or decrease in protein myristoylation in the body wall muscle (skeletal muscle in Caenorhabditis elegans) cells leads to age-related defects in sarcomere structure and loss of mobility in the animals. Imbalanced proteostasis of ER was observed in these animals with mobility defect. Moreover, alleviation of such ER stress condition with genetic manipulation can partially suppress both sarcomere defect and paralysis. Finally, our published data demonstrated that reduction of FAs biosynthesis caused decrease of protein myristoylation. Therefore, we hypothesized that FAs impact sarcomere integrity, at least partially through regulation of myristoylation of proteins and ER homeostasis. We will test this hypothesis through elucidating the cellular and molecular mechanism that myristoylation affects ER homeostasis, which in turn influences the sarcomere integrity, by using C elegans and mouse myoblast C2C12 cells. This study unveiled a profound impact of FA on muscle function through affecting sarcomere integrity and will discover the underlying mechanism that links FA metabolism, protein myristoylation and ER homeostasis with muscle maintenance, which should provide valuable insights into understanding the impact of nutrients and ER stress on the onset of muscle degeneration.

脂肪酸代谢在组织维持及动物发育过程中发挥重要作用。脂肪酸代谢异常与肌肉退行性疾病的发生密切相关，但相关分子机理尚不明确。申请人前期研究发现：在秀丽隐杆线虫中，脂肪酸合成减少导致肉豆寇酰化水平下调；脂肪酸合成或蛋白质肉豆寇酰化修饰的下降，可引发肌小节结构缺陷并导致运动能力的渐进性丧失；内质网应激在这些肌小节缺陷线虫中被诱发；缓解内质网应激可部分恢复肌小节结构并逆转瘫痪。鉴于此，我们推测：肉豆寇酰化修饰通过影响内质网蛋白动态平衡，部分介导了脂肪酸对肌小节的作用。本研究拟以秀丽隐杆线虫及小鼠成肌细胞C2C12为研究对象，从分子水平上解析蛋白质肉豆寇酰化修饰影响内质网蛋白动态平衡的机制，及该影响调控肌小节结构的机理。申请人前期工作首次揭示了脂肪酸可通过调控肌小节结构影响肌肉功能，本项目将从肉豆寇酰化修饰和内质网应激的视角探索相关机制，为理解代谢异常引发肌肉退行性疾病的机理提供新思路。

机体维持组织完整性能力的下降与衰老和退行性疾病密切相关。脂肪酸代谢对动物的发育和组织维持有着深远的影响，但我们对相关机制的了解仍然有限。该项目中，我们使用秀丽隐杆线虫研究了脂肪酸水平是否以及如何影响肌肉完整性。我们发现，通过阻断脂肪酸生物合成降低总脂肪酸水平或抑制蛋白肉豆蔻酰化会导致肌小节结构紊乱和成年线虫瘫痪。进一步机制研究表明，两种ARF鸟苷三磷酸酶(GTPases)的肉豆蔻酰化介导了脂肪酸缺乏对肌小节完整性的影响。具体来说，脂肪酸减少下调arf蛋白的肉豆蔻酰化修饰水平，继而通过诱导内质网(ER)应激和ER未折叠蛋白反应(UPRER)磷酸化eIF2a，从而导致肌小节成分PINCH水平的降低和肌球蛋白的紊乱。该研究揭示了脂肪酸信号、蛋白质肉豆蔻酰化、内质网稳态与肌小节完整性的因果联系，为理解营养物质和内质网稳态在肌肉维持中的调节作用提供新的视角。.此外，鉴于蛋白肉豆蔻酰修饰的重要性，我们还建立了高信噪比可系统性分离和鉴定动物体内肉豆蔻酰化修饰蛋白的方法，利用该方法我们鉴定到了许多以前未知的肉豆蔻酰化蛋白为进一步理解肉豆蔻酰修饰的生理功能奠定了基础。更为重要的是，这些方法还可拓展到研究和发现其它类型的脂肪酸修饰蛋白，通过相应优化，目前我们已通过质谱鉴定到了一类全新的以前未知的脂肪酸修饰类型。对这一新型脂肪酸修饰类型的研究将进一步拓宽我们对脂肪酸发挥生理功能的机制理解。