Fig. 4
APP TMD mutations alter GSEC processivity and function, paralleling PSEN1/2 mutation effects. A Schematic representation of APP TMD primary structure highlighting residues affected by selected mutations. Color-coding: red: pathogenic and blue: not classified. Mutation positions in APP are shown, and the corresponding position in the Aβ sequence is shown in brackets. Pathogenicity information was taken from Alzforum database. B Aβ profiles (relative abundance of Aβ37, Aβ38, Aβ40, and Aβ42 peptides relative to total Aβ levels) generated by HEK293T cells expressing WT or mutant APPC99 substrates. Aβ43 levels (very low) were excluded (*); Aβ43 levels were measured in at least 3 independent experiments (Supplementary Figure S4). Data presented as mean ± SD, N ≥ 3 independent experiments. C Efficiency of the 4 th enzymatic turnover of APPC99 (GSEC ‘dysfunction’) quantified by the adapted (*) processivity Aβ(37 + 38 + 40)/42 ratio, normalised to APP WT. Data presented as mean ± SD, N ≥ 3 independent experiments. Statistics: One-way ANOVA with Dunnett's post-hoc test vs WT; ****p < 0.0001, F(18, 77) = 157.5. D Correlation analysis between Aβ(37 + 38 + 40)/42* processivity ratio and clinical AAOs for APP TMD mutations. Significant linear correlation found (equation: Y = 1.5 x—34, R2 = 0.69) and 95% confidence interval shown as blue area. Error bars represent SD for Aβ ratio (x-axis) and AAO (y-axis). E Aβ40/42 ratio data normalised to WT APP. Data presented as mean ± SD, N ≥ 3 independent experiments. Statistics: One-way ANOVA with Dunnett's post-hoc test vs WT; ****p < 0.0001, F(19, 80) = 231.9. F Correlation analysis between Aβ40/42 ratio and clinical AAOs for APP TMD mutations. Significant linear correlation found (equation: Y = 1,8*X – 57, R2 = 0.72) and 95% confidence interval shown as blue area. Error bars represent SD for Aβ ratio (x-axis) and AAO (y-axis). G Product line preference ratio (Aβ(37 + 40)/(38 + 42)) data normalised to WT APP. Data presented as mean ± SD, N ≥ 3 independent experiments. Statistics: One-way ANOVA with Dunnett's post-hoc test vs WT; ****p < 0.0001, F(18, 85) = 286.3. H Correlation analysis between Aβ(37 + 40)/(40 + 42) product line preference ratio and AAOs for APP TMD mutations. Significant linear correlation found (equation: Y = 1.8 x—57, R2 = 0.74) and 95% confidence interval (blue area). Error bars represent SD for Aβ ratio (x-axis) and AAO (y-axis)