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. 2019 Jul 4;18(16):1976–1994. doi: 10.1080/15384101.2019.1637201

Figure 6.

Figure 6.

The localization of Kiz-pT379 on chromatin at metaphase is perturbed in HeLa S3 cells expressing Myc-CHC_T606D. (A, B, C) Typical IF images of Tet-ON inducible HeLa S3 cells expressing Myc-vector, Myc-CHC_WT, Myc-CHC_T606A, or Myc-CHC_T606D at interphase and during M phase. Cells were labeled with anti-Kiz-pT379 (A), anti-γ-tubulin (A, C), anti-PLK1 (B), and anti-Myc (C) Abs. DNA was counter-stained with Hoechst33258 to detect chromatin. Yellow arrowheads, turquoise arrows, and orange arrows indicate Kiz-pT379 signals at the centrosome, chromatin, and midbody, respectively. Pink arrows indicate co-localized Kiz-pT379 and chromatin signals. White arrows in Myc-CHC_T606D-expressing cells indicate the absence of Kiz-pT379. Scale bars, 10 µm. (D) Representative images of Kiz-pT379 immunofluorescence in GL2- and siGAK-treated cells. The scattered γ-tubulin foci (more than two) were merged with the Kiz-pT379 foci (white arrowheads). Scale bars, 10 µm. (E) Comparison of Kiz-pT379 signal intensity at the centrosome of GL2- and siGAK-treated cells using Metaview software. Integrated intensities of GL2-treated Kiz-pT379 dot foci (n = 50) and siGAK-treated Kiz-pT379 dot foci (n = 210) were scored. The dot graphs in (E) show the average + SE values of these measurements. All measurements are statistically significant when GL2 and siGAK are compared (P < 0.01).