Figure 8. JADE1S phosphorylation and chromatin dissociation is hindered by pharmacological inhibitor of Aurora A kinase. (A) Experimental design: HeLa cells were synchronized by arresting cell cycle at late G₁ with mimosine. At 0 h cell cycle was released by refreshing media supplemented with nocodazole. At 15 h, inhibitor of Aurora A, VX-680 was added into the media, and at 18 h samples were collected for analysis as described in Figure 4, except here the total cell population was collected (no mitotic shake off). (B) Soluble (S) and chromatin-enriched (Ch) fractions were analyzed for endogenous JADE1S, β-Actin (loading control) and nucleolin (nuclear marker) by western blots. Samples 1–4: (1) cells treated with mimosine only, (2) cells treated with mimosine, nocodazole, and VX-680 added at 15 h, (3) cells treated with nocodazole only, (4) asynchronous cells. Soluble fraction after nocodazole (3) and chromatin fraction of asynchronous cells (4) was used as a reference for high and low molecular mass specie of JADE1S, respectively. Note that the band shift and the chromatin dissociation corresponding to JADE1S were inhibited by the addition of VX-680. (C) Cell cycle profiles of the samples 1–4 by FACS analysis.