The luminal pH of endolysosomal vesicles acidifies as endosomes mature and ultimately fuse with lysosomes. Maintenance of this pH gradient depends on recruitment and assembly of the vacuolar H+-ATPase (V-ATPase). Lysosomal pH is dysregulated in many disease states, but how changes in lysosomal pH impact endosomal trafficking and maturation is still unclear. Upstream of endolysosomal fusion, the small GTPase Rab7 influences maturation of late endosomes (LEs) as part of a ternary complex with Rab-interacting lysosomal protein (RILP) and the V-ATPase subunit V1G1. In this study (Mulligan et al., 2024), Bettina Winckler and colleagues test the effects of disrupting lysosomal pH on LEs and the Rab7–RILP–V1G1 complex. Using both pharmacological and genetic approaches, they find that lysosomal pH neutralisation changes the size and morphology of LEs and increases LE pH without impacting membrane permeability. This leads to increased assembly of V-ATPase at LEs, which correlates with activation and stabilisation of LE-associated Rab7. Dominant-negative Rab7 is still recruited to pH-neutralised LEs positive for V1G1, suggesting that Rab7 is recruited downstream of V-ATPase assembly. The presence of RILP further stabilises Rab7 at LE membranes. Taken together, these data suggest that disruption of lysosomal pH induces an endolysosomal stress response that triggers V-ATPase assembly and Rab7 hyperactivation at LEs, which serves to alter dynamics of membrane trafficking receptors.
