Protein extraction and western blotting

Protein extraction was performed using a modified Dignam method, as described previously [44, 45]. Briefly, a cytoplasmic extraction was made with buffer A (10 mM Tris pH 7.8; 10 mM KCl; 1.5 mM MgCl₂) and a nucleoplasmic extraction was made with buffer C (10 mM Tris pH 7.8; 0.42 M NaCl; 1.5 mM MgCl₂; 0.2 mM EDTA; 25% glycerol). Then, both fractions were mixed. For histone extraction, an acidic extraction of the pellet with 0.2M HCl was performed following protein isolation.

For the Western blotting procedure, proteins were separated on SDS-PAGE and transferred onto a nitrocellulose membrane. The membrane was blocked with 5% milk in PBS-0.1% Tween, incubated with a primary antibody diluted in PBS-0.1% Tween as directed by the antibody supplier (Supplementary Table 2), and subsequently incubated with HRP secondary antibody. The image was detected using the Odyssey FC LI·COR machine.