Figure 4.

Cell cycle defects in the TIF1 knockout mutant. (A) Growth curves for the wild-type (CU428, dashed line) and homozygous TIF1 knockout (TXk202, solid line) mutant were generated by averaging six hemocytometer cell counts per time point (n = 5 experiments). (B) Delayed cell division in the TIF1 germline knockout. Asynchronous vegetative cultures were visually examined for dividing cells (presence of a cleavage furrow). Knockout: homozygous TIF1::neo germline replacement (TXk202). Heterozygote: heterozygous TIF1::neo germline replacement (TXh102). The percentage of cells in late cytokinesis was determined by averaging the results from six to seven independent experimental analyses. (C) Elongated macronuclear S phase and delayed cytokinesis in TIF1-deficient cells. Wild-type and homozygous TIF1 knockout strains were synchronized with a stationary/starvation/refeeding protocol (Mohammad et al., 2003). Refed cultures were pulse-labeled with BrdU for a 15 min at 30-min intervals. Indirect immunoflourescence was used to quantify the percentage of BrdU-positive cells; dashed black line: wild-type (CU428); solid black line: TIF1 knockout (TXk202). Cytokinesis (dashed gray line: wild-type; solid gray line: TIF1 knockout) was scored by microscopic detection of a cleavage furrow.