FIGURE 6.

Inhibition of endogenous MLL5 phosphorylation impedes mitotic progression and arrests cells at the G₂/M transition. HEK 293T cells were transfected with GFP-CD-4 in the absence (no treatment (NT)) or presence of RO-3306 for 20 h before release into complete medium or nocodazole-containing medium for 5 h. A, cell cycle analysis for GFP-negative control cells and GFP-CD-4-expressing cells. The percentage represents the population that successfully progressed through mitosis and re-entered G₁ phase after 5 h release from G₂ arrest. B, population of GFP-CD-4 positive or GFP-negative cells released from RO-3306 was sorted separately before harvesting for Western blotting. Phosphorylation of endogenous MLL5 at G₂/M phase (G₂ release + nocodazole (Noc)) was inhibited by ectopic overexpression of GFP-CD-4. Arrowhead indicates phospho-MLL5; arrow indicates MLL5.