miRNA-9* and miRNA-206 target HDAC4/HDAC5 and Myst3, respectively. (A) BCWM.1 cells (scramble probe–, pre-miRNA-9*–transfected, and untransfected) were harvested at 12 hours after transfection. Nuclear lysates were subjected to Western blot with the use of anti-HDAC4, -HDAC5, -Myst3, and -nucleolin antibodies. (B) BCWM.1 cells (scramble probe–, anti–miRNA-206–transfected, and untransfected) were harvested at 12 hours after transfection. Nuclear lysates were subjected to Western blot with the use of anti-Myst3 and -nucleolin antibodies. (C) BCWM.1 cells (scramble probe–, pre-miRNA-9*–, anti–miRNA-206–transfected, and untransfected) were harvested at 12 hours after transfection. Whole-cell lysates were subjected to Western blotting with the use of anti–acetyl-histone H3, –acetyl-histone H4, –acetylated-tubulin, –acetylated-lysine, and -actin antibodies. (D) BCWM.1 cells (scramble probe–, pre-miRNA-9*–, anti–miRNA-206–transfected, and untransfected) were harvested at 12 hours after transfection. HDAC activity was assessed in vitro with the use of nuclear extracts by Colorimetric HDAC Activity Assay Kit (*P < .05). (E) BCWM.1, RL, and MEC-1 cells were cultured with LBH589 (20-60nM) for 16 hours or with control medium. Whole-cell lysates were subjected to Western blotting with the use of anti–acetyl-histone H3, –acetyl-histone H4, –acetylated-tubulin, –acetylated-lysine, and –α-tubulin antibodies. (F) BCWM.1 cells were cultured in the presence or absence of LBH589 (0-80nM; 8 hours). HDAC activity was assessed in vitro with the use of nuclear extracts by Colorimetric HDAC Activity Assay Kit (all P ≤ .05).