ABSTRACT

Lipid accumulation often leads to lipotoxic injuries to hepatocytes, which can cause nonalcoholic steatohepatitis. The association of inflammation with lipid accumulation in liver tissue has been studied for decades; however, key mechanisms have been identified only recently. In particular, it is still unknown how hepatic inflammation regulates lipid metabolism in hepatocytes. Herein, we found that PA treatment or direct stimulation of STING1 promoted, whereas STING1 deficiency impaired, MTORC1 activation, suggesting that STING1 is involved in PA-induced MTORC1 activation. Mechanistic studies revealed that STING1 interacted with several components of the MTORC1 complex and played an important role in the complex formation of MTORC1 under PA treatment. The involvement of STING1 in MTORC1 activation was dependent on SQSTM1, a key regulator of the MTORC1 pathway. In SQSTM1-deficient cells, the interaction of STING1 with the components of MTORC1 was weak. Furthermore, the impaired activity of MTORC1 via rapamycin treatment or STING1 deficiency decreased the numbers of LDs in cells. PA treatment inhibited lipophagy, which was not observed in STING1-deficient cells or rapamycin-treated cells. Restoration of MTORC1 activity via treatment with amino acids blocked lipophagy and LDs degradation. Finally, increased MTORC1 activation concomitant with STING1 activation was observed in liver tissues of nonalcoholic fatty liver disease patients, which provided clinical evidence for the involvement of STING1 in MTORC1 activation. In summary, we identified a novel regulatory loop of STING1-MTORC1 and explain how hepatic inflammation regulates lipid accumulation. Our findings may facilitate the development of new strategies for clinical treatment of hepatic steatosis.

Abbreviations: AA: amino acid; ACTB: actin beta; cGAMP: cyclic GMP-AMP; CGAS: cyclic GMP-AMP synthase; DEPTOR: DEP domain containing MTOR interacting protein; EIF4EBP1: eukaryotic translation initiation factor 4E binding protein 1; FFAs: free fatty acids; GFP: green fluorescent protein; HFD: high-fat diet; HT-DNA: herring testis DNA; IL1B: interleukin 1 beta; LAMP1: lysosomal associated membrane protein 1; LDs: lipid droplets; MAP1LC3: microtubule associated protein 1 light chain 3; MAP1LC3B: microtubule associated protein 1 light chain 3 beta; MEFs: mouse embryonic fibroblasts; MLST8: MTOR associated protein, LST8 homolog; MT-ND1: mitochondrially encoded NADH: ubiquinone oxidoreductase core subunit 1; mtDNA: mitochondrial DNA; MTOR: mechanistic target of rapamycin kinase; MTORC1: MTOR complex 1; NAFL: nonalcoholic fatty liver; NAFLD: nonalcoholic fatty liver disease; NASH: nonalcoholic steatohepatitis; NPCs: non-parenchymal cells; PA: palmitic acid; PLIN2: perilipin 2; RD: regular diet; RELA: RELA proto-oncogene, NF-kB subunit; RPS6: ribosomal protein S6; RPS6KB1: ribosomal protein S6 kinase B1; RPTOR: regulatory associated protein of MTOR complex 1; RRAGA: Ras related GTP binding A; RRAGC: Ras related GTP binding C; SQSTM1: sequestosome 1; STING1: stimulator of interferon response cGAMP interactor 1; TBK1: TANK binding kinase 1; TGs: triglycerides; TREX1: three prime repair exonuclease 1.

摘要

脂质积累通常会导致肝细胞的脂毒性损伤，从而导致非酒精性脂肪性肝炎。炎症与肝组织中脂质积累的关系已经研究了几十年。然而，直到最近才确定了关键机制。特别是，尚不清楚肝脏炎症如何调节肝细胞中的脂质代谢。在这里，我们发现 PA 治疗或 STING1 的直接刺激促进了 MTORC1 的激活，而 STING1 缺乏则损害了 MTORC1 的激活，这表明 STING1 参与了 PA 诱导的 MTORC1 激活。机理研究表明，STING1 与 MTORC1 复合物的几个组分相互作用，并在 PA 处理下 MTORC1 复合物形成中起重要作用。STING1 参与 MTORC1 激活取决于 SQSTM1，MTORC1 通路的关键调节因子。在 SQSTM1 缺陷细胞中，STING1 与 MTORC1 成分的相互作用很弱。此外，通过雷帕霉素治疗或 STING1 缺乏导致 MTORC1 活性受损，减少了​​细胞中 LD 的数量。PA 处理抑制了脂肪吞噬，这在 STING1 缺陷细胞或雷帕霉素处理的细胞中未观察到。通过氨基酸处理恢复 MTORC1 活性可阻止脂肪吞噬和 LDs 降解。最后，在非酒精性脂肪性肝病患者的肝组织中观察到伴随 STING1 激活的 MTORC1 激活增加，这为 STING1 参与 MTORC1 激活提供了临床证据。总之，我们发现了一个新的 STING1-MTORC1 调节环，并解释了肝脏炎症如何调节脂质积累。

缩写：AA：氨基酸；ACTB：肌动蛋白β；cGAMP：环状GMP-AMP；CGAS：环状 GMP-AMP 合酶；DEPTOR：含有 MTOR 相互作用蛋白的 DEP 结构域；EIF4EBP1：真核翻译起始因子 4E 结合蛋白 1；FFA：游离脂肪酸；GFP：绿色荧光蛋白；HFD：高脂肪饮食；HT-DNA：鲱鱼睾丸DNA；IL1B：白细胞介素 1 β；LAMP1：溶酶体相关膜蛋白 1；LDs：脂滴；MAP1LC3：微管相关蛋白 1 轻链 3；MAP1LC3B：微管相关蛋白 1 轻链 3 β；MEFs：小鼠胚胎成纤维细胞；MLST8：MTOR相关蛋白，LST8同源物；MT-ND1：线粒体编码的 NADH：泛醌氧化还原酶核心亚基 1；mtDNA：线粒体DNA；MTOR：雷帕霉素激酶的机制靶点；MTORC1：MTOR复合物1；NAFL：非酒精性脂肪肝；非酒精性脂肪肝：非酒精性脂肪肝；NASH：非酒精性脂肪性肝炎；NPC：非实质细胞；PA：棕榈酸；PLIN2：perilipin 2；RD：规律饮食；RELA：RELA原癌基因，NF-kB亚基；RPS6：核糖体蛋白 S6；RPS6KB1：核糖体蛋白 S6 激酶 B1；RPTOR：MTOR复合物1的调节相关蛋白；RRAGA：Ras 相关 GTP 结合 A；RRAGC：Ras相关的GTP结合C；SQSTM1：隔离体 1；STING1：干扰素反应 cGAMP 相互作用物 1 的刺激物；TBK1：TANK结合激酶1；TGs：甘油三酯；TREX1：三素修复核酸外切酶1。MTOR 复合物 1 的调节相关蛋白；RRAGA：Ras 相关 GTP 结合 A；RRAGC：Ras相关的GTP结合C；SQSTM1：隔离体 1；STING1：干扰素反应 cGAMP 相互作用物 1 的刺激物；TBK1：TANK结合激酶1；TGs：甘油三酯；TREX1：三素修复核酸外切酶1。MTOR 复合物 1 的调节相关蛋白；RRAGA：Ras 相关 GTP 结合 A；RRAGC：Ras相关的GTP结合C；SQSTM1：隔离体 1；STING1：干扰素反应 cGAMP 相互作用物 1 的刺激物；TBK1：TANK结合激酶1；TGs：甘油三酯；TREX1：三素修复核酸外切酶1。