BACKGROUND Eukaryotic elongation factor-2 kinase (eEF2K) is a Ca 2+ /calmodulin (CaM)-dependent protein kinase that inhibits protein synthesis. However, the role of eEF2K in cancer development was reported paradoxically and remains to be elucidated. METHODS Herein, A549 cells with eEF2K depletion or overexpression by stably transfected lentivirus plasmids were used in vitro and in vivo study. MTT and colony assays were used to detect cell proliferation and growth. Extracellular glucose and lactate concentration were measured using test kit. Immunoblot and co-immunoprecipitation assays were used to examine the molecular biology changes and molecular interaction in these cells. LC-MS/MS analysis and [γ- 32 P] ATP kinase assay were used to identify combining protein and phosphorylation site. Nude mice was utilized to study the correlation of eEF2K and tumor growth in vivo. RESULTS We demonstrated that eEF2K inhibited lung cancer cells proliferation and affected the inhibitory effects of EGFR inhibitor gefitinib. Mechanistically, we showed that eEF2K formed a complex with PKM2 and STAT3, thereby phosphorylated PKM2 at T129, leading to reduced dimerization of PKM2. Subsequently, PKM2 impeded STAT3 phosphorylation and STAT3-dependent c-Myc expression. eEF2K depletion promoted the nuclear translocation of PKM2 and increased aerobic glycolysis reflected by increased lactate secretion and glucose. CONCLUSIONS Our findings define a novel mechanism underlying the regulation of cancer cell proliferation by eEF2K independent of its role in protein synthesis, disclosing the diverse roles of eEF2K in cell biology, which lays foundation for the development of new anticancer therapeutic strategies.

中文翻译：

---

eEF2 激酶诱导的 STAT3 失活通过 PKM2 磷酸化抑制肺癌细胞增殖。


背景真核延伸因子2激酶(eEF2K)是一种抑制蛋白质合成的Ca 2+ /钙调蛋白(CaM)依赖性蛋白激酶。然而，eEF2K 在癌症发展中的作用的报道是矛盾的，并且仍有待阐明。方法本文使用稳定转染慢病毒质粒去除eEF2K或过表达eEF2K的A549细胞进行体外和体内研究。 MTT和集落测定用于检测细胞增殖和生长。使用测试套件测量细胞外葡萄糖和乳酸浓度。使用免疫印迹和免疫共沉淀测定来检查这些细胞中的分子生物学变化和分子相互作用。使用LC-MS/MS分析和[γ- 32 P] ATP激酶测定来鉴定结合蛋白和磷酸化位点。利用裸鼠研究eEF2K与体内肿瘤生长的相关性。结果我们证明eEF2K抑制肺癌细胞增殖并影响EGFR抑制剂吉非替尼的抑制作用。从机制上讲，我们发现 eEF2K 与 PKM2 和 STAT3 形成复合物，从而在 T129 磷酸化 PKM2，导致 PKM2 二聚化减少。随后，PKM2 阻碍 STAT3 磷酸化和 STAT3 依赖性 c-Myc 表达。 eEF2K 耗竭促进了 PKM2 的核转位，并通过乳酸分泌和葡萄糖的增加反映了有氧糖酵解的增加。结论 我们的研究结果定义了 eEF2K 调节癌细胞增殖的新机制，与其在蛋白质合成中的作用无关，揭示了 eEF2K 在细胞生物学中的多种作用，这为开发新的抗癌治疗策略奠定了基础。