Figure 4. PSAP knockdown does not affect TFEB localization and increases extracellular PGRN more robustly than other lysosomal proteins.

(a–c) GFP immunofluorescence images of HeLa cells that stably express TFEB-GFP. Images were acquired after cells were transfected with 20 nM of either a control (a) or PSAP siRNA (b), or post treatment with chloroquine (CQ) (c). (d) Quantification of the effects of siRNA transfection or CQ treatment on the nuclear/cytoplasmic ratio of TFEB-GFP (n=21 fields from three independent experiments, totalling ≥70 cells per group). (e) Immunoblot of HeLa cell media from 20 nM control or PSAP siRNA transfections. (f) Quantification of IFI30, cathepsin L (CTSL), cathepsin D (CTSD) and PGRN protein levels in HeLa cell media post PSAP knockdown (n=4 per group). Graphed values represent the mean±s.e.m. *Differs from control-transfected cells, P<0.001; **P<0.0005 by one-way ANOVA (d) or Student's t-tests (f). Scale bar, 10 μm.