Ecl development and imaging

Whole-cell lysates for immunoblotting were prepared by dissolving cells in Laemmli Sample Buffer containing 5% 2-mercaptoethanol (Bio-Rad). Protein samples were loaded on 4–15% polyacrylamide gels (BioRad) and transferred onto 0.45 µm pore-size nitrocellulose membranes (Bio-Rad) using the Turboblot (BioRad). After that, blots were blocked with 5% milk for 1 h at room temperature. For phospho-tau, 5% BSA in TBS was used for blocking. Blots were incubated overnight at 4 °C with primary antibodies. And then blots were probed with secondary antibodies for 1 h at room temperature before ECL development and imaging (Bio-Rad). The primary antibodies used for immunoblotting are as follows: Lamin A/C antibody, (Abcam, 1:750); Lamin B1 antibody (Santa Cruz, 1:200); APP antibody (BioLegend, 1:400), total tau antibody (Santa Cruz, 1:200), PHF6 p-tau antibody (Santa Cruz, 1:200), γ-H2AX antibody (Abcam, 1:3000), and β-actin (1:5000, Sigma-Aldrich).

Whole-cell lysates for immunoblotting were prepared by dissolving cells in Laemmli Sample Buffer containing 5% 2-mercaptoethanol (Bio-Rad, Hercules, CA, USA). Nuclear and cytosolic fractions of iPSCs-differentiated cells were acquired using NE-PER™ Nuclear and Cytoplasmic Extraction Reagents (Thermo Fisher Scientific, Waltham, MA, USA #78833) as per the manufacturer’s instructions. Protein samples were loaded on 10–12% polyacrylamide gels and transferred onto 0.45 µm pore-size nitrocellulose membranes (Bio-Rad) using the Turboblot (BioRad). Blots were incubated overnight at 4 °C with primary antibodies and then probed with secondary antibodies before ECL development and imaging (Bio-Rad). The primary antibodies used for immunoblotting are as follows: lamin A/C (1:500, Millipore MAB3211), progerin (1:500, Cao et al., 2011), ∆Np63 (1:500, Biolegend, San Diego, CA, USA #619001), K14 (1:500, Invitrogen MA5-11599), K8 (1:500, Biolegend #904804), K18 (1:1000, Cell Signaling #4548), total β-Catenin (1:1000, Cell Signaling #9562), Non-phospho (Active) β-Catenin (1:1000, Cell Signaling #8814), RCC1 (1:1000, Cell Signaling #3589), S6 Ribosomal Protein (1:1000, Cell Signaling #2217), LEF1 (1:250, Santa Cruz sc-374412), and β-actin (1:1000, Sigma-Aldrich, St. Louis, MO, USA A3854).