College of Life Sciences, Liaoning Normal University, Dalian 116081, China; College of Life Science and Technology, Dalian University, Dalian 116622, China.
College of Life Sciences, Liaoning Normal University, Dalian 116081, China; Lamprey Research Center, Liaoning Normal University, Dalian 116081, China; Collaborative Innovation Center of Seafood Deep Processing, Dalian Polytechnic University, Dalian 116034, China.
Life Sci. 2022 Oct 15;307:120888. doi: 10.1016/j.lfs.2022.120888. Epub 2022 Aug 17.
Excess cholesterol deposition in lysosomes may result in lysosomal stress and dysfunction. Here, we focus on the role of lysosome membrane protein 2 (LIMP2) in relieving the lysosomal stress caused by excess cholesterol and the mechanism that regulate its expression.
Cholesterol enrichment in lamprey liver tissue was evaluated by RNA transcriptome data analysis, RT-qPCR, H&E, and Oil Red O staining. Gene markers of autophagy and cholesterol synthesis were determined by western blot or RT-qPCR. Lysosomal morphology and pH value was measured by confocal observation or flow cytometry. Dual-Luciferase reporter assay was performed to test the expression regulation relationship.
We report that lamprey limp2 (L-limp2) is evolutionarily highly conserved with human LIMP2 (H-LIMP2). The biological function of L-limp2, consistent with H-LIMP2, includes maintaining lysosomal morphology, modulating autophagy, and aiding cholesterol efflux from lysosomes. Furthermore, we find that both L-limp2 and H-limp2 can restore cholesterol-induced elevation of lysosomal pH and impaired autophagic flux. We demonstrate that lamprey transcription factor binding to IGHM enhancer 3 (L-TFE3) can bind with coordinated lysosomal expression and regulation (CLEAR) elements on the L-limp2 promoter and regulate its expression. Moreover, this regulatory relationship is also available in humans. Taken together, the present study demonstrates that the evolutionarily conserved TFE3-LIMP2 axis may have a protective role against the impaired lysosomal function caused by excess cholesterol.
The protective effect of TFE3-LIMP2 axis against cholesterol-triggered lysosomal stress may provide a new target for the treatment of diseases caused by excessive cholesterol accumulation in lysosomes.
溶酶体中过多的胆固醇沉积可能导致溶酶体应激和功能障碍。在这里，我们专注于溶酶体膜蛋白 2（LIMP2）在缓解过量胆固醇引起的溶酶体应激中的作用及其调节其表达的机制。
通过 RNA 转录组数据分析、RT-qPCR、H&E 和油红 O 染色评估七鳃鳗肝组织中的胆固醇富集。通过 Western blot 或 RT-qPCR 测定自噬和胆固醇合成的基因标志物。通过共聚焦观察或流式细胞术测量溶酶体形态和 pH 值。进行双荧光素酶报告基因检测以测试表达调控关系。
我们报告称，七鳃鳗 limp2（L-limp2）与人类 LIMP2（H-LIMP2）在进化上高度保守。L-limp2 的生物学功能与 H-LIMP2 一致，包括维持溶酶体形态、调节自噬和促进胆固醇从溶酶体中流出。此外，我们发现 L-limp2 和 H-limp2 都可以恢复胆固醇诱导的溶酶体 pH 值升高和受损的自噬通量。我们证明，七鳃鳗转录因子结合免疫球蛋白重链增强子 3（L-TFE3）可以与 L-limp2 启动子上的协调溶酶体表达和调节（CLEAR）元件结合，并调节其表达。此外，这种调节关系在人类中也是存在的。总之，本研究表明进化上保守的 TFE3-LIMP2 轴可能对过量胆固醇引起的溶酶体功能障碍具有保护作用。
TFE3-LIMP2 轴对胆固醇触发的溶酶体应激的保护作用可能为治疗溶酶体中过量胆固醇积累引起的疾病提供新的靶点。
