Fig. 5.

Formation of large TIN2 domains correlates with exit from the cell cycle. (A) Percentage of S1 cells with large TIN2 domain(s) and Ki-67 staining. S1 cells were cultured in 3D for 3 days to obtain a mixed population of cycling Ki-67 positive (Ki-67+) and growth-arrested Ki-67 negative (Ki-67−) cells, and then fixed and dual immunostained for TIN2 and Ki-67. Shown is the percentage of cells containing large TIN2 domains (TIN2 clusters) and Ki-67 positive (filled bars) or negative (open bars) staining. Error bars show the s.e.m. for three different experiments. *P<0.001 when compared with Ki-67+ cells. (B) Dual immunostaining for TIN2 (green) and Ki-67 (red) in S1 cells after 3D culture for 3 days. The different phases of the cell cycle were identified by the pattern of Ki-67 staining. The percentage of cells showing large TIN2 domains (TIN2 clusters) in each phase of the cell cycle is given below each panel, and is the mean±s.e.m. of three different experiments. Arrowheads indicate large TIN2 domains and dashed lines delineate the nuclear periphery. (C) Histogram of the percentage of synchronized 184 HMECs with large TIN2 domains (TIN2 clusters) as a function of the cell cycle. Nuclei showing large TIN2 domains were counted as a percentage of total nuclei (revealed by DAPI counterstaining) during exponential (EXP), G0, S and G2-M phases of the cell cycle. Percentages of Ki-67-positive nuclei in each phase are shown. Bar, 2.5 μm.