dMtmr6 and MTMR8 maintain endolysosomal homeostasis. (A, B, D, and E) Representative maximum intensity projection of z-stack image from serum-fed (A and D) or serum-starved (B and E) siRNA-transfected HeLa cells stained with antibodies to detect TFEB (green). (C) Quantification of nuclear TFEB in serum-fed and serum-starved HeLa cells transfected with the indicated siRNA as represented in A, B, D, and E. (F) Changes in transcription of known target genes of Mitf/TFEB relative to actin-5c in fat body from Luciferase-RNAi (−) or dMtmr6-RNAi (+) fat body–specific knockdown tissue. Please note that Mitf is the Drosophila TFEB orthologue, and Cp1 is the gene for cathepsin L protein. (G) Representative image of fat body from fed L3 Drosophila larvae with WT (GFP-negative) and clonal dMtmr6-knockdown cells (white dashed outline, green) stained with antibody to detect Rab7 (red) and Rab7 only channel depicted in G′. (H and I) Quantification of cellular Rab7 puncta number (H) and size (I) in WT control and dMtmr6-knockdown cells as represented in G. Data are presented as mean ± SD, n = 10 random images per genotype and condition (C); mean ± SD (n = 3 biological replicates; F); and mean ± minimum to maximum (n ≥ 11; H and I). Asterisks denote statistical significance (*, P < 0.05; ****, P < 0.0001) using one-way ANOVA (C), unpaired two-tailed t test in (F), and paired two-tailed t test (H, and I). Scale bars, 20 µm. n.s., not significant.