Overexpression of an AKI-resistant mutant of Aurkb renders Myc-expressing lymphoma cells resistant to Aurk inhibition. (A) Eμ-Myc lymphoma cells were infected with control virus (pBABE) or pBABE-AurkbG160V (Aurkb-KIR) encoding retrovirus. The cells were then selected with puromycin and treated with 25nM AS703569 (AKI) for 24 hours. The representative histogram shows the DNA content assessed by PI staining. (B) Quantification of aneuploidy and apoptosis on AKI treatment in control cells and cells expressing of Aurk-KIR. Cells were treated with carrier only (untreated) or 25nM AS703569 (AKI) and subjected to DNA content analysis using PI staining and flow cytometry. The PI-stained cells were analyzed using the FL2 channel in a linear scale. Apoptosis measurements were based on the percentage of cells that carried less than diploid DNA content (Sub-G1) in the FL3 channel in a logarithmic scale. (C) Control cells (pBABE) or cells expressing mutant Aurkb (Aurkb-KIR) were treated for the indicated time with 25nM AS703569 (AKI) and assessed for Aurkb and phosphorylated S10-HH3 levels by immunoblotting. Note that Aurkb-KIR was Flag-tagged and migrates slightly slower than endogenous Aurkb.
