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. 2022 Nov 30;15:1069122. doi: 10.3389/fnmol.2022.1069122

FIGURE 2.

FIGURE 2

The effect of CSTB-deficiency on H3cs1 levels and cathepsin L activity. (A) Representative confocal microscopy images of H3cs1 immunohistochemical staining in the cerebellum of P14, P21, and P120 wt and Cstb–/– mice. EGL = external granular cell layer, ML = molecular layer, CGL = cerebellar granular cell layer. Scale bar = 40 μm. (B) Quantitative analysis of H3cs1 immunohistochemical staining in the cerebellum of P7, P14, P21, P30, and P120 wt and Cstb–/– mice (n = 5 mice/timepoint). The charts depict H3cs1 content as mean ± SEM. The pattern of H3cs1 intensity in the examined timepoints is significantly increased in Cstb–/– samples (P < 0.0001). (C) Western blot detection of H3cs1 in cerebellar lysates of P14 and P30 wt and Cstb–/– mice. The bar chart depicts fold changes in H3cs1 levels from P14 to P30 in wt and Cstb–/– mice. Normalized intensity values are plotted as means ± SEM (n = 3 mice/timepoint and genotype). AB = amido black. (D) RT-qPCR analysis of Cstb mRNA expression in postnatal brain of wt mice plotted as means ± SEM (n = 4 mice/timepoint). (E) Bar charts depicting the enzymatic activity of cathepsin L in whole brain lysates and in chromatin-bound protein fractions of wt and Cstb–/– P30 mice. Aminofluorocumarin production values (nmol AFC/min) are normalized to the total protein input and plotted as means ± SEM (n = 6 mice/timepoint and genotype). *P < 0.05, **P < 0.01, and ****P < 0.0001.