Figure S6.
Positive regulation of neuronal peroxisome abundance by PKC.(A) Western blot of β-3 tubulin in the non-differentiated and 18-day differentiated SH-SY5Y. Quantification shows the ratio of and β-3 tubulin to GAPDH, mean ± SEM, N = 3, ****P < 0.0001, unpaired t test. (B and C) Quantitative PCR of the neuronal markers and PKC isoforms in the (B) 18-day differentiated SH-SY5Y cells in control and Go6983 1 μM conditions; Go6983 was added on days 10–18 of the differentiation protocol. Quantification shows the relative expression of differentiated/non-differentiated markers, expressed as 2−ΔΔCt, mean ± SEM, N = 4, *P < 0.05, Kruskal–Wallis test, and (C) comparison of day 0 and 18 expression, mean ± SEM, N = 4, *P < 0.05, **P < 0.01, ****P < 0.0001, one-way ANOVA. (D) Confocal microscopy of peroxisomes in the non-differentiated and 18-day differentiated SH-SY5Y cells in control and Go6983 1 μM conditions. Go6983 was added on days 10–18 of the differentiation protocol. Peroxisomes were visualized with the PMP70 antibody, nuclei were stained with Hoechst (10 μg/ml), and neuronal differentiation was visualized with β-3 tubulin antibody. Representative images are shown. Scale bar: 5 μm. See zoomed in images in Fig. 5 F. (E) Quantitative PCR of the neuronal markers in the 12-day differentiated NPCs. Quantification shows the relative expression of differentiated/non-differentiated markers, expressed as 2−ΔΔCt, mean ± SEM. (F) Confocal microscopy of peroxisomes in the non-differentiated and 12-day differentiated NPCs in control and Go6983 1 μM conditions. Go6983 was added on days 1–12 of the differentiation protocol. Peroxisomes were visualized with the PMP70 antibody, nuclei were stained with Hoechst (10 μg/ml), and neuronal differentiation was visualized with β-3 tubulin antibody. Representative images are shown. Scale bar: 5 μm; inlet: 1 μm. (G) Confocal microscopy of peroxisomes in the neuronal terminals of the 18-day differentiated SH-SY5Y and 12-day differentiated NPCs in control and Go6983 1 μM conditions. Peroxisomes were visualized with the PMP70 antibody, and neuronal terminals were visualized with β-3 tubulin antibody. Representative images are shown. Scale bar: 5 μm. Source data are available for this figure: SourceData FS6.

Positive regulation of neuronal peroxisome abundance by PKC. (A) Western blot of β-3 tubulin in the non-differentiated and 18-day differentiated SH-SY5Y. Quantification shows the ratio of and β-3 tubulin to GAPDH, mean ± SEM, N = 3, ****P < 0.0001, unpaired t test. (B and C) Quantitative PCR of the neuronal markers and PKC isoforms in the (B) 18-day differentiated SH-SY5Y cells in control and Go6983 1 μM conditions; Go6983 was added on days 10–18 of the differentiation protocol. Quantification shows the relative expression of differentiated/non-differentiated markers, expressed as 2−ΔΔCt, mean ± SEM, N = 4, *P < 0.05, Kruskal–Wallis test, and (C) comparison of day 0 and 18 expression, mean ± SEM, N = 4, *P < 0.05, **P < 0.01, ****P < 0.0001, one-way ANOVA. (D) Confocal microscopy of peroxisomes in the non-differentiated and 18-day differentiated SH-SY5Y cells in control and Go6983 1 μM conditions. Go6983 was added on days 10–18 of the differentiation protocol. Peroxisomes were visualized with the PMP70 antibody, nuclei were stained with Hoechst (10 μg/ml), and neuronal differentiation was visualized with β-3 tubulin antibody. Representative images are shown. Scale bar: 5 μm. See zoomed in images in Fig. 5 F. (E) Quantitative PCR of the neuronal markers in the 12-day differentiated NPCs. Quantification shows the relative expression of differentiated/non-differentiated markers, expressed as 2−ΔΔCt, mean ± SEM. (F) Confocal microscopy of peroxisomes in the non-differentiated and 12-day differentiated NPCs in control and Go6983 1 μM conditions. Go6983 was added on days 1–12 of the differentiation protocol. Peroxisomes were visualized with the PMP70 antibody, nuclei were stained with Hoechst (10 μg/ml), and neuronal differentiation was visualized with β-3 tubulin antibody. Representative images are shown. Scale bar: 5 μm; inlet: 1 μm. (G) Confocal microscopy of peroxisomes in the neuronal terminals of the 18-day differentiated SH-SY5Y and 12-day differentiated NPCs in control and Go6983 1 μM conditions. Peroxisomes were visualized with the PMP70 antibody, and neuronal terminals were visualized with β-3 tubulin antibody. Representative images are shown. Scale bar: 5 μm. Source data are available for this figure: SourceData FS6.

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