Fig. 1

In vivo SILAC labelling reveals elevated lysosomal hydrolase expression induced by lack of AEP. a Transmission electron microscopy of WT (upper panel) and AEP^−/− (lower panel) kidney sections. Arrows indicate lysosomes. (Scale bar = 2 μm) b Immunofluorescence showing more prominent lysosomes (CtsD, green) in AEP^−/− kidney sections (lower panel) compared to WT kidney sections (upper panel). (Scale bar = 20 μm). c Western blot analysis confirming the accumulation in AEP^−/− kidneys of proteins identified by mass spectrometry. d Volcano plot showing proteins over-represented (blue dots, lysosomal proteins highlighted in yellow) or under-represented (red dots) in AEP^−/− lysosomal kidney fractions against the −log10 p-value for three independent experiments. Dotted line on y axis indicates p value < 0.05. e Acute AEP inhibition with MVO26630 recapitulates hydrolase induction in WT MEF and AEP reconstitution reverses it in AEP^−/− MEF. f Induction of CtsD/E and CtsB/L activities in AEP^−/− MEF compared to WT MEF. Data are the average ± SD of n = 4 biologically independent samples for CtsD/E or n = 3 biologically independent samples for CtsB/L. Statistical significance was calculated using a two-sided unpaired t-test. g Semi-quantitative RT-PCR analysis of mRNA levels for different cathepsins in WT MEF, WT MEF treated overnight with 50 μM MVO26630 and AEP^−/− MEF. Data are the average ± SD of n = 3 biologically independent samples. Statistical significance was calculated using a Dunnett’s multiple comparison test