溶酶体是胞内物质降解和代谢调控的重要细胞器,其紊乱导致胞内物质(如脂质)堆积和代谢性疾病发生;促进溶酶体生成并增强其降解脂质的功能,可改善代谢性疾病。目前溶酶体生成的机制远未阐明，因此揭示溶酶体生成的调控机制是重要科学问题。申请人实验发现:饥饿(EBSS)激活AMPK从而诱导转录抑制因子ZKSCAN3出核进入胞质; 出核的ZKSCAN3不再抑制溶酶体基因表达，导致溶酶体生成和脂滴降解。另一方面，申请人找到ZKSCAN3蛋白上AMPK的潜在磷酸化位点，过表达突变型ZKSCAN3可阻断EBSS-AMPK诱导的溶酶体生成和脂滴降解。因此我们假说:营养匮乏时细胞通过AMPK抑制ZKSCAN3、进而促进溶酶体生成和脂滴降解。本项目运用细胞生物学、遗传学、化学生物学等方法并结合小鼠模型，阐明AMPK调控ZKSCAN3进而促进溶酶体生成和脂滴降解的新机制，为改善代谢性疾病提供理论基础和治疗靶点。

Lysosomes are acidified organelles regulating degradation and metabolism within cells. Abnormality of lysosomal homeostasis results in cargo accumulation and severe human disorders, including metabolic diseases and neurodegenerative diseases. Whereas, promotion of lysosome biogenesis enhances lysosomal-dependent degradation and alleviates human lysosome-related diseases. The mechanisms underlying lysosome biogenesis are, however, poorly understood. Therefore, scientific questions about novel mechanisms of lysosome biogenesis need to be addressed. Our studies have found that EBSS activated AMPK, which in turn promoted cytosolic translocation of both EGFP-ZKSCAN3 and endogenous ZKSCAN3. Notably, we also found potential AMPK-associated phosphorylation sites on ZKSCAN3. These mutations were resistant to EBSS-AMPK signaling pathway and blocked EBSS-induced cytosolic translocation of ZKSCAN3, lysosome biogenesis, and clearance of lipid.droplets. Thus, our hypothesis is that EBSS-activated AMPK phosphorylates transcription repressor ZKSCAN3 and promotes cytosolic translocation of ZKSCAN3, resulting in lysosome biogenesis and clearance of lipid droplets in cellular and mouse models. In this project, we plan to further investigate how AMPK-ZKSCAN3 axis regulates metabolism in adipose tissue and other organs, using a combination of approaches including genetic screen, protein-protein interactions, live-cell imaging, protein modifications, and ZKSCAN3 KO/cKO mice. This unique research will not only provide important novel insights into lysosomal homeostasis and clearance of lipid droplets, but also reveal pathological mechanisms underlying lysosome-related human metabolic disorders.