Lipid metabolism is highly compartmentalized between cellular organelles that dynamically adapt their compositions and interactions in response to metabolic challenges. Here, we investigate how diet-induced hepatic lipid accumulation, observed in non-alcoholic fatty liver disease (NAFLD), affects protein localization, organelle organization, and protein phosphorylation in vivo. We develop a mass spectrometric workflow for protein and phosphopeptide correlation profiling to monitor levels and cellular distributions of ∼6,000 liver proteins and ∼16,000 phosphopeptides during development of steatosis. Several organelle contact site proteins are targeted to lipid droplets (LDs) in steatotic liver, tethering organelles orchestrating lipid metabolism. Proteins of the secretory pathway dramatically redistribute, including the mis-localization of the COPI complex and sequestration of the Golgi apparatus at LDs. This correlates with reduced hepatic protein secretion. Our systematic in vivo analysis of subcellular rearrangements and organelle-specific phosphorylation reveals how nutrient overload leads to organellar reorganization and cellular dysfunction.

脂质代谢在细胞器之间高度区分开，细胞器可动态适应其组成和相互作用以应对代谢挑战。在这里，我们调查在非酒精性脂肪肝疾病（NAFLD）中观察到的饮食诱导的肝脂质蓄积如何影响体内的蛋白质定位，细胞器组织和蛋白质磷酸化。我们开发了蛋白质和磷酸肽相关性分析的质谱工作流程，以监测脂肪变性发展过程中约6,000种肝蛋白和约16,000种磷酸肽的水平和细胞分布。几种细胞器接触位点蛋白靶向脂肪肝中的脂质滴（LDs），栓系细胞器协调脂质代谢。分泌途径的蛋白质急剧地重新分布，包括COPI复合物的错误定位和LD处高尔基体的隔离。这与肝蛋白分泌减少有关。我们对亚细胞重排和细胞器特异性磷酸化的系统体内分析揭示了营养素超负荷如何导致细胞器重组和细胞功能障碍。