Figure 7.

Nuclear localization of Dicer and Ago2. (A) Western analysis of Dicer in cytoplasmic and nuclear fractions. The 5% cytoplasmic and 20% nuclear fractions prepared from HeLa cells were loaded in a 4–12% SDS–PAGE gel, transferred to a membrane and proteins were detected using antibodies. Alpha-tubulin and hnRNP A2 were used as controls for cytoplasmic and nuclear proteins, respectively. (B) Ago2 can be found in both cytoplasmic and nuclear fractions. Western analysis was performed using the same samples as in (C), and the membrane was probed using different antibodies. GAPDH and hnRNPA2 were detected and used as controls for cytoplasmic and nuclear proteins, respectively. (C) Localization of Dicer (upper panel) and Ago2 (lower panel) in Hela cells. Cells grown on glass-bottom dishes were fixed, stained with first antibodies against Dicer (1:200, ab14601, from mouse, Abcam) and Nucleolin (1:200, ab22758, from rabbit, Abcam), or against Ago2 (1:150, ab57113, from mouse, Abcam) and Nucleolin, as described in ‘Materials and Methods’ section. Secondary antibodies were anti-mouse antibody (1:200, ab6785, Abcam, conjugated with FITC (green) and anti-rabbit antibody (1:200, Ab6719, conjugated with Texas Red). Nucleolin (red) was detected and served as a nucleolar marker. The nucleus was stained with DAPI (blue). The arrow indicates the positions of nucleoli. The scale bars: 10 µm.