目的:PI3K-AKT通路是否以及如何,代谢稳态的中心节点,高脂诱导的非酒精性脂肪性肝炎(NASH)和肝细胞癌(HCC)仍然是一个谜。在这种情况下,AKT调节的特征将为抗击HCC提供新的策略。
方法:代谢文库筛选揭示棕榈酸(PA)可以激活AKT。采用体内和体外棕榈酰化测定法检测AKT棕榈酰化。不同的细胞和小鼠模型,包括AKT1C77S和AKT1C224S敲入细胞的产生,使用Zdhhc17和Zdhhc24敲除小鼠和Akt1C224S敲入小鼠。来自NASH和HCC患者的人类肝组织,流体动力学转染小鼠模型,我们还进一步探索了高脂/高胆固醇饮食(HFHCD)诱导的NASH/HCC小鼠模型和高脂和蛋氨酸/胆碱缺乏饮食(HFMCD)诱导的NASH小鼠模型进行机制研究。
结果:通过筛选代谢物库,PA已被定义为通过促进其棕榈酰修饰来激活AKT,生长因子诱导的AKT激活的必要步骤。生物学,高脂肪饮食可以促进AKT激酶的活性,从而促进NASH和肝癌。机械上,棕榈酰结合以不依赖PIP3的方式将AKT锚定到细胞膜上,部分通过防止AKT组装成非活性聚合物。棕榈酰转移酶ZDHHC17/24被表征为棕榈酰化AKT以发挥致癌作用。有趣的是,抗肥胖药物奥利司他或特异性穿透肽可以通过限制PA合成或抑制AKT修饰来有效减弱AKT棕榈酰化和激活,分别,从而拮抗肝脏肿瘤发生。
结论:我们的发现阐明了PA-ZDHHC17/24介导的棕榈酰化对AKT的新型微调调节,并通过服用PA限制饮食来强调肿瘤治疗策略,限制PA合成,或直接靶向AKT棕榈酰化。
Whether and how the PI3K-AKT pathway, a central node of metabolic homeostasis, is responsible for high-fat-induced non-alcoholic steatohepatitis (NASH) and hepatocellular carcinoma (HCC) remain a mystery. Characterisation of AKT regulation in this setting will provide new strategies to combat HCC.
Metabolite library screening disclosed that palmitic acid (PA) could activate AKT. In vivo and in vitro palmitoylation assay were employed to detect AKT palmitoylation. Diverse cell and mouse models, including generation of AKT1C77S and AKT1C224S knock-in cells, Zdhhc17 and Zdhhc24 knockout mice and Akt1C224S knock-in mice were employed. Human liver tissues from patients with NASH and HCC, hydrodynamic transfection mouse model, high-fat/high-cholesterol diet (HFHCD)-induced NASH/HCC mouse model and high-fat and methionine/choline-deficient diet (HFMCD)-induced NASH mouse model were also further explored for our mechanism studies.
By screening a metabolite library, PA has been defined to activate AKT by promoting its palmitoyl modification, an essential step for growth factor-induced AKT activation. Biologically, a high-fat diet could promote AKT kinase activity, thereby promoting NASH and liver cancer. Mechanistically, palmitoyl binding anchors AKT to the cell membrane in a PIP3-independent manner, in part by preventing AKT from assembling into an inactive polymer. The palmitoyltransferases ZDHHC17/24 were characterised to palmitoylate AKT to exert oncogenic effects. Interestingly, the anti-obesity drug orlistat or specific penetrating peptides can effectively attenuate AKT palmitoylation and activation by restricting PA synthesis or repressing AKT modification, respectively, thereby antagonising liver tumorigenesis.
Our findings elucidate a novel fine-tuned regulation of AKT by PA-ZDHHC17/24-mediated palmitoylation, and highlight tumour therapeutic strategies by taking PA-restricted diets, limiting PA synthesis, or directly targeting AKT palmitoylation.