Retromer subunit, VPS29, regulates vacuolar‐type ATPase and lysosomal acidification in the aging brain

Background The retromer, including VPS35, VPS26, and VPS29, is an evolutionarily conserved protein complex that retrieves cargoes from the endosome to the trans ‐Golgi network or plasma membrane. Retromer regulates Tau phosphorylation and pathogenic processing of the amyloid precursor protein, both of which are neuropathological hallmarks of Alzheimer’s disease (AD). Previously, we found that loss of VPS29 in Drosophila causes progressive lysosomal dysfunction during aging, but the underlying molecular mechanism remains elusive. Method To investigate the molecular events in Drosophila heads caused by loss of Vps29, we performed quantitative proteomics on Vps29 mutants and age‐matched control animals, and identified 480 significant, differentially expressed proteins. To study the lysosomal acidity in fly Vps29 mutants, we neuronally expressed LAMP1‐GFP , a pH‐sensitive indicator that is localized to the lysosomal lumen. To evaluate the potential translational relevance of our findings, we knocked down retromer subunits in mammalian cell culture and examined levels of vacuolar‐type ATPase (v‐ATPase) subunits. We also interrogated available human postmortem brain proteomic data from the Religious Orders Study and Rush Memory and Aging Project (ROSMAP). Result Unbiased proteomics from VPS29 mutant flies revealed a sharp decline in subunits of the v‐ATPase, a multimeric proton pump that is primarily responsible for lysosomal acidification. Importantly, we obtained consistent evidence from mammalian cell culture. In aged Vps29 mutant fly brains, the LAMP1‐GFP pH sensor indicated impaired lysosomal acidification. Lastly, based on ROSMAP human brain proteomics, we found that VPS29 protein levels were significantly coexpressed with other proteins implicated in energy‐coupled proton transmembrane transport, including multiple subunits of the conserved v‐ATPase. Conclusion Our findings highlight a conserved role for retromer in regulating the v‐ATPase, possibly contributing to lysosomal acidification. Drosophila is a powerful model system for functional dissection of the retromer in the mammalian brains, including in AD pathogenesis.