Inhibition of PIKfyve maximally swells endolysosomes and restores high [Cl ⁻ ] in Snx10 KO macrophages. (A) WT RAW 264.7 macrophages transiently transfected with ClC-7-GFP and treated either with vehicle alone (DMSO; control) or with 100 nM YM201036 for 30 min prior to imaging. (B) Representative images of WT RAW 264.7 macrophages treated with 100 nM YM201636 for 30 min, then acutely treated with vehicle only (Control) or with 3 mM ATP for 30 s in Na-gluconate medium, then labeled with either FM1–43 or cresyl violet prior to imaging. (C) Endolysosomal [Cl⁻] was measured colorimetrically in WT and Snx10 KO RAW 264.7 macrophages as detailed in Materials and methods. (D) WT and Snx10 KO RAW 264.7 macrophages transiently transfected with ClC-7^Y715C-GFP for 12 h were either untreated or treated with YM201636 for 30 min prior to imaging. (E) HeLa cells were transiently transfected with Rab7-RFP (magenta). Where indicated, (right panel) the cells were also co-transfected to overexpress Snx10-GFP. (F) HeLa cells were transiently transfected with ClC-7-GFP with and without co-transfection (overexpression) of Snx10-mCherry. All images are representative of ≥3 similar experiments of each type.