Lysosomal proteomics reveals mechanisms of neuronal apoE4-associated lysosomal dysfunction

ApoE4 is the primary risk factor for Alzheimer’s Disease. While apoE is primarily expressed by astrocytes, AD pathology including endosomal abnormalities and mitochondrial dysfunction first occurs in neurons. Lysosomes are poised at the convergence point between these features. We find that apoE4-expressing cells exhibit lysosomal alkalinization, reduced lysosomal proteolysis, and impaired mitophagy. To identify driving factors for this lysosomal dysfunction, we performed quantitative lysosomal proteome profiling. This revealed that apoE4 expression results in lysosomal depletion of Lgals3bp and accumulation of Tmed5 in both Neuro-2a cells and postmitotic human neurons. Modulating the expression of both proteins affected lysosomal function, with Tmed5 knockdown rescuing lysosomal alkalinization in apoE4 cells, and Lgals3bp knockdown causing lysosomal alkalinization and reduced lysosomal density in apoE3 cells. Taken together, our work reveals that apoE4 exerts gain-of-toxicity by alkalinizing the lysosomal lumen, pinpointing lysosomal Tmed5 accumulation and Lgals3bp depletion as apoE4-associated drivers for this phenotype. ### Competing Interest Statement Yadong Huang is a co-founder and scientific advisory board member of GABAeron. The Krogan Laboratory has received research support from Vir Biotechnology, F. Hoffmann-La Roche, and Rezo Therapeutics. Nevan J. Krogan has previously held financially compensated consulting agreements with the Icahn School of Medicine at Mount Sinai, New York, and Twist Bioscience Corp. He currently has financially compensated consulting agreements with Maze Therapeutics, Interline Therapeutics, Rezo Therapeutics, and GEn1E Lifesciences, Inc. He is on the Board of Directors of Rezo Therapeutics and is a shareholder in Tenaya Therapeutics, Maze Therapeutics, Rezo Therapeutics, and Interline Therapeutics. Danielle L. Swaney has financially compensated consulting agreements with Maze Therapeutics and Rezo Therapeutics. The other authors declare no competing interests.