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. 2014 Nov 27;5(11):e1547. doi: 10.1038/cddis.2014.492

Figure 6.

Figure 6

PKC activation promotes ADAM10 delivery to the postsynaptic compartment. (a) Representative WB of ADAM10 and SAP97 localization in HOMO, TIF and P3 fraction upon PKC activation in hippocampal slices (PDBu, 100 nM, 30 min). PKC activation modifies ADAM10 and SAP97 cellular distribution. (b) Quantification of experiments in (a) (n=7, ADAM10, *P=0.008, TIF, PDBu versus control (CTRL); P=0.008, P3, PDBu versus CTRL; SAP97, P=0.02, P3, PDBu versus CTRL, paired t-test). In this and all subsequent fractionation experiments, data are expressed as the percentage of CTRL. (c) Representative WB of ADAM10 and SAP97 localization in hippocampal slices treated with BFA before PKC activation. BFA affects ADAM10 but not SAP97 trafficking. (d and e) Quantification of experiments in (c) (n=5, *P<0.05 versus CTRL; #P<0.05 versus PDBu, one-way analysis of variance (ANOVA), Bonferroni's post hoc test). (f) Staining of ADAM10/dsRed-ER and ADAM10/GM130 in dendrites and soma of neurons upon PDBu treatment. PKC activation decreases ADAM10 levels in dendritic and somatic ER and Golgi. In all experiments, primary neurons were kept in culture 14 DIV before treatment and staining experiments. Scale bar, 10 μm. (g) Quantification of experiments in (f) (24 neurons per condition from three independent experiments, dsRED-ER: dendrites, *P=0.001; soma, *P=0.009; GM130: dendrites, *P=0.0017; soma, *P=0.0006, PDBu versus CTRL unpaired t-test). (h) ADAM10/PSD-95 staining showing that PKC activation increases ADAM10 levels in the PSD. Scale bar, 10 μm. (i) Quantification of experiments in (h) (24 neurons per condition from three independent experiments, *P=0.00001, PDBu versus CTRL, unpaired t-test). (j) Representative staining of SAP97/GM130 in dendrites and soma of neurons after PDBu treatment; PKC activation reduces SAP97 levels in dendritic and somatic Golgi. Scale bar, 10 μm. (k) Quantification of experiments in (j) (24 neurons per condition from three independent experiments, dendrites, *P=0.0001; soma, *P=0.0006, PDBu versus CTRL, unpaired t-test)