Loss of the CHC–GTSE1 interaction impacts spindle shape but not k-fiber stability. (A) Immunofluorescence images of mitotic spindles (α-tubulin staining) from U2OS, GTSE1^WT, and GTSE1^5xLID cells, transfected with control (Ctrl) or GTSE1 siRNA. (B–D) The ratio of spindle width over spindle length (B), the inner-spindle total tubulin fluorescence intensity (C), and the spindle volume (D) were measured in cells from A using 3D objects representing the spindles without astral MTs (n > 60 cells per condition; n = 2 experiments pooled for analysis and presented as box and whisker plots; P values from Wilcoxon test adjusted for multiple comparisons [false discovery rate]). n.s., not significant. (E) Immunofluorescence images showing cold-resistant MTs (α-tubulin) in U2OS, GTSE1^WT, and GTSE1^5xLID cells, transfected with control (Ctrl) or GTSE1 siRNA. G2-synchronized cells (RO3306) were released into mitosis for 55 min before cold treatment. (F) Quantification of the remaining total tubulin fluorescence intensity in cells treated as in E (n = 3, n ≥ 50 cells per condition and per experiment, one experiment presented as a box and whisker plot; P values from Wilcoxon test adjusted for multiple comparisons [false discovery rate]).