Nuclear localization of Topo IIα during interphase is not necessary for localization to chromosomes in mitosis. (A, top) Sequence of the extreme CTR of Topo IIα. Red residues indicate putative constituents of the bipartite NLS. Blue residues indicate the ChT domain described herein. (A, bottom) Schematic showing GFP- and mCherry-tagged WT Topo IIα and mutants and truncation constructs used herein. GFP–Topo IIα K3R comprised of K1489, K1490, and K1492 mutated to Arg. (B) M. muntjak cells transfected with GFP–Topo IIα or GFP–Topo IIα K3R imaged in live interphase cells. The WT protein is predominantly nuclear; the K3R mutant is predominantly cytoplasmic. Bars, 10 µm. (C) GFP–Topo IIα K3R localizes similarly to WT GFP–Topo IIα in live mitotic M. muntjak cells. Lines indicate locations of line profile plots in Fig 2 D. See also Video 2. Bars, 10 µm. (D) Quantification of WT Topo IIα and Topo IIα K3R abundance at mitotic chromosome cores in live M. muntjak cells. Both localize to the chromosome core with equal intensity. Shown are the mean of maximum intensities across n = 73 (K3R) and n = 53 (WT) core regions after background subtraction. P-value indicates two-tailed unpaired samples t test. Error bars indicate SD.