Effect of Compound K on the expressions of Bcl-2 family proteins for Compound K-induced apoptosis in HL-60 cells. (A) Cells were harvested after being incubated with Compound K (20 μM) for the indicated times. Mitochondrial (M) and cytosolic (C) fractions were prepared as described in Methods. Bid, tBid, Bax, Bcl-2, and Bcl-xL were analyzed by Western blotting. α-tubulin, COX IV and β-actin were used as internal controls. (B) HL-60 cells were treated with 20 μM of Compound K for 4 h and cell lysates were prepared. Bax was immunoprecipitated from total protein lysates and proteins were subjected to Western blotting for anti-Bax 6A7 using specific antibody. (C) Compound K-induced interactions between Bcl-2 family members. tBid and Bax were immunoprecipitated using specific antibodies from total protein lysates and Western blotting was conducted for Bcl-2 and Bcl-xL (IP; immunoprecipitation, IB; immunoblotting).