Fig. 11

Syt1 modulates BACE1 maturation and stability. a Levels of total and immature BACE1 in protein lysates from parental and Syt1 KD PC12 cells were determined by western blotting using anti-BACE1 antibodies directed against the C-terminus or prodomain of BACE1, respectively (n = 5). The specificity of the antibodies was confirmed using overexpression of human BACE1 in CHO cells (top gel) and brain lysates from BACE1 KO and control littermate mice (bottom gel). The intensities of all BACE1 bands were quantified by densitometry and normalized to the levels of β actin. Data are presented as mean ± SEM. Statistical significance was determined using unpaired student t-test. ** p < 0.01, *** p < 0.001. b Analysis of BACE1 degradation. Parental and Syt1 KD PC12 cells were treated with cycloheximide and harvested 0, 2, 4, 8, 24, and 48 hours after the treatment (n = 5). Western blot presents BACE1 expression over time. Anti-BACE1 C-terminal antibody was used for detection. The quantitative analysis of BACE1 levels revealed a significantly reduced half-life of BACE1 in Syt1 KD PC12 cells compared to the parental PC12 control (marked with black dashed lines). Statistical significance was determined using 2-way ANOVA with Bonferroni post-test, *** p < 0.001