Blue loading buffer

The soluble protein samples were heated for 5 min at 95° C in Blue Loading Buffer (New England Biolabs) with 40 mM of dithiothreitol and a total of 10 μg (β-actin), 20 μg (γH2AX), 25 μg (PS1), 35 μg (LRP), 40 μg (APP and H2AX), 50 μg (mTERT) and 60 μg (BACE1) of protein was then separated on AnykD™ Criterion™ TGX Stain-Free™ Protein Gels (Biorad). A prestained molecular weight marker (PageRuler Prestained Protein Ladder, Thermo Fisher) was loaded onto each gel. After transferring the proteins onto polyvinylidine fluoride membrane (Pall) with 1 × transfer buffer (20% methanol in 25 mM Tris and 19.2 mM glycine) the membranes were blocked in 3% BSA (Amresco) in PBS and 0.1% Tween 20 (PBST). The primary and secondary antibodies used for western blots are detailed in Supplementary Table 1. The proteins were visualized with Clarity™ Western ECL Blotting Substrate (Biorad) and the ChemiDoc™ Imaging System (Biorad). Densitometric analysis was performed with Image Lab 5.1 software (Biorad).

Total protein extraction was performed with the same number of cells for each condition. The pellets were incubated in Blue Loading Buffer (New England Biolabs, B7703S, New England Biolabs, Ipswich, MA, USA) with 0.1 unit/mL Benzonase (Merck, 70664-3, Merck, Darmstadt, Germany) for 1 h at room temperature. The extracted proteins were denatured at 95 °C for 5 min.