Gamma secretase orthologs are required for lysosomal activity and autophagic degradation in <i>Dictyostelium discoideum</i>, independent of PSEN (presenilin) proteolytic function

Mutations in the γ-secretase complex are strongly associated with familial Alzheimer disease. Both proteolytic and non-proteolytic functions for the γ-secretase complex have been previously described in mammalian model organisms, but their relative contributions to disease pathology remain unclear. Here, we dissect the roles of orthologs of the γ-secretase components in the model system <i>Dictyostelium</i>, focusing on endocytosis, lysosomal activity and autophagy. In this model, we show that the orthologs of PSEN (psenA and psenB), Ncstn (nicastrin) and Aph-1 (gamma-secretase subunit Aph-1), are necessary for optimal fluid-phase uptake by macropinocytosis and in multicellular development under basic pH conditions. Disruption of either psenA/B or Aph-1 proteins also leads to disrupted phagosomal proteolysis as well as decreased autophagosomal acidification and autophagic flux. This indicates a general defect in lysosomal trafficking and degradation, which we show leads to the accumulation of ubiquitinated protein aggregates in cells lacking psenA/B and Aph-1 proteins. Importantly, we find that all the endocytic defects observed in <i>Dictyostelium</i> PSEN ortholog mutants can be fully rescued by proteolytically inactive <i>Dictyostelium</i> psenB and human PSEN1 proteins. Our data therefore demonstrates an evolutionarily conserved non-proteolytic role for presenilin, and γ-secretase component orthologs, in maintaining <i>Dictyostelium</i> lysosomal trafficking and autophagy. Atg8: autophagy protein 8a; Aph-1: gamma-secretase subunit Aph-1; crtA: calreticulin; ER: endoplasmic reticulum; GFP: green fluorescent protein; GSK3B: glycogen synthase kinase 3 beta; Ncstn: nicastrin; PSEN1: presenilin 1; psenA and psenB: <i>Dictyostelium</i> presenilin A and B; TRITC; tetramethylrhodamine isothiocyanate.