Supplementary Figure 9: Individual western blotting experiments.

(a–h) Individual quantification of the western blot signals for each PDE3A mutant in the three independent experiments shown in Figure 4a,b. The signals were densitometrically quantified in relation to the loading control β-tubulin. Because of experimental variation and quality differences between the antibodies to phosphorylated protein, we observed variations in the investigated phosphorylation of Ser428 and Ser438 in PDE3A1 and PDE3A2. The antibody to PDE3A (Bethyl) recognizes motifs beginning at position 450. Perhaps the G449V substitution alters the neighboring protein domain and alters antibody binding, leading to the weak signals in the upper blots. However, the pooled results (Fig. 4a,b) of all mutants showed statistical significance compared to wild-type PDE3A (WT) in non-parametrical Mann-Whitney rank-sum testing.