Elongation factor 1A1 and the regulation of protein synthesis and cytoskeleton dynamics during lipid-induced stress

延伸因子（EEF）1A 1在肝脏、胰腺和大脑中表达，并通过促进aa-tRNA-核糖体结合参与蛋白质合成。EEF 1A 1还在肌动蛋白细胞骨架调节和响应于应激的凋亡/失巢凋亡中具有“兼职”功能。 EEF 1A 1的经典和非经典功能是否参与细胞对代谢应激的反应尚不清楚。这个项目的目的是阐明蛋白质合成，肌动蛋白细胞骨架的调节和脂质过载引起的细胞应激反应之间的联系。 脂毒性是非脂肪细胞中异位脂质沉积导致细胞功能障碍、细胞死亡和最终组织功能障碍的过程。由于许多脂质代谢过程发生在ER，因此分泌细胞类型中的脂毒性条件伴随着ER应激和未折叠蛋白反应（UPR）的快速诱导。UPR通过停止蛋白质合成来恢复体内平衡，同时促进蛋白质折叠和降解。 然而，严重或长时间的应激会引发内质网应激反应（ERSR），从而导致细胞死亡。 ERSR涉及重新启动蛋白质合成以产生促炎因子和促凋亡因子-一种有害的合成代谢反应。EEF 1A 1在ER应激时被诱导，并且已知是脂毒性细胞死亡的关键介质。我们最近发现，用特异性抑制剂didemnin B阻断EEF 1A 1的蛋白合成活性，可降低肝细胞中的脂毒性细胞死亡。此外，用didemnin B急性干预改善了严重脂肪肝小鼠的肝脏ER应激和相关炎症。但EEF 1A 1参与细胞应激反应和促进肝细胞脂毒性的确切机制尚不清楚。 目前还不清楚EEF 1A 1是否参与其他富含ER的细胞类型（特别是胰腺β细胞）的脂毒性。因此，该项目的短期目标是确定EEF 1A 1在对过量脂质蓄积的细胞应答中的比较功能（1）在培养的肝细胞和胰腺β细胞中，以及（2）在具有饮食诱导的组织脂质蓄积的小鼠的肝脏和胰腺中。 长期计划的目标是了解EEF 1A 1在全身代谢组织（包括肝脏、胰腺、心肌和骨骼肌）中对过量脂质的细胞应激反应中的整体作用机制，部分通过使用独特的转基因Eef 1a 1-EGFP小鼠模型来测试全身Eef 1a 1表达模式，以响应高脂饮食诱导的代谢应激。总的来说，该计划将确定EEF 1A 1的经典和非经典功能如何参与ER应激反应和细胞脂质过度积累诱导的细胞死亡过程，从而进一步加深我们对EEF 1A 1的脂毒性和多种功能的理解。

Elongation factor (EEF) 1A1 is expressed in liver, pancreas, and brain, and participates in protein synthesis by promoting aa-tRNA-ribosome binding. EEF1A1 also has ‘moonlighting' functions in actin cytoskeleton regulation, and in apoptosis/anoikis in response to stress. Whether both the canonical and non-canonical functions of EEF1A1 are involved in cellular responses to metabolic stress is unclear. The goal of this program is to elucidate the links between protein synthesis, regulation of the actin cytoskeleton, and cellular stress responses induced by lipid overload. Lipotoxicity is the process whereby ectopic lipid deposition in non-adipose cells leads to cellular dysfunction, cell death, and eventual tissue dysfunction. Since many lipid metabolic processes occur at the ER, lipotoxic conditions in secretory cell types are accompanied by rapid induction of ER stress and the unfolded protein response (UPR). The UPR restores homeostasis by halting protein synthesis, while promoting protein folding and degradation. However, severe or prolonged stress initiates an ER stress response (ERSR) which can lead to cell death. The ERSR involves re-initiation of protein synthesis to generate pro-inflammatory and pro-apoptotic factors – a deleterious anabolic response. EEF1A1 is induced in upon ER stress, and is known to be a key mediator of lipotoxic cell death. We recently found that blocking the protein synthetic activity of EEF1A1 with a specific inhibitor, didemnin B, decreased lipotoxic cell death in hepatocytes. Moreover, acute intervention with didemnin B improved hepatic ER stress and associated inflammation in mice with severe fatty liver. But the precise mechanisms through which EEF1A1 participates in cellular stress responses and promotes hepatocyte lipotoxicity are unclear. It is also not known whether EEF1A1 participates in lipotoxicity in other ER-rich cell types, particularly pancreatic beta-cells. Thus the short term objectives of this program are to determine the comparative functions of EEF1A1 in cellular responses to excess lipid accumulation (1) in cultured hepatocytes and pancreatic beta-cells, and (2) in liver and pancreas of mice with diet-induced tissue lipid accumulation. The long term program goal is to understand the global mechanism(s) of action of EEF1A1 in cellular stress responses to excess lipid in metabolic tissues throughout the body, including liver, pancreas, myocardium, and skeletal muscle, in part by using a unique transgenic Eef1a1-EGFP mouse model to test whole body Eef1a1 expression patterns in response to metabolic stress induced by high fat diet. Overall, this program will determine how the canonical and non-canonical functions of EEF1A1 are involved in ER stress response and cell death processes induced by excessive cellular lipid accumulation, thereby furthering our understanding of lipotoxicity and the diverse functions of EEF1A1.