Fig. 5: Structure-function analysis of CIP2A. | Nature Communications

Fig. 5: Structure-function analysis of CIP2A.

From: CIP2A mediates mitotic recruitment of SLX4/MUS81/XPF to resolve replication stress-induced DNA lesions

Fig. 5

a CIP2A is predicted to form a stable homodimer, involving a globular domain and a long coiled-coil. Two predicted TOPBP1-binding sites and the possibly flexible position of the N-terminal regions onto the coiled-coil shaft are indicated. b Cartoon and surface representation of the predicted CIP2A:CIP2A dimer structure with two copies of TOPBP1 755-860. Predicted TOPBP1:CIP2A binding interfaces are magnified. c Schematic representation of full length (1-905) CIP2A (WT) and schematic representation of V5-tagged CIP2A variants. Quantification of V5 foci and TOPBP1 foci per mitotic cell (d) or micronuclei per cell (e) in RPE1 TP53–/– CIP2A–/– cl#1 cells reconstituted with indicated CIP2A-V5 variants treated with APH (200 nM, 20 h). Individual values, medians, and interquartile range of three (d) or at least three (e) biologically independent experiments are plotted. Ordinary one-way ANOVA with Šidák’s multiple comparisons test (d) or Dunnett’s multiple comparison test (e) on medians per experiment. f–l CIP2A–/– cells reconstituted with CIP2A-WT, CIP2A-S904A or CIP2A-ΔC were treated with APH (200 nM, 20 h). Panel f: representative wide-field images of cells stained for DAPI (blue), V5 (red), SLX4 (green). Scale bar:10 µm. Quantification of foci of SLX4 (g), MUS81 (h) or XPF (i) per mitotic cell in CIP2A–/– cl#1 reconstituted with indicated mutants. Individual values, medians and interquartile range of three biologically independent experiments are plotted. Ordinary one-way ANOVA with Šidák’s multiple comparisons test on the medians per experiment. ‘n’ in panels (d, e, g–i) represents the total number of cells across experiments. Left: confocal overview images. Right: representative STED microscopy images of DAPI (blue), V5 (green) and TOPBP1 (red) structures in CIP2A–/– cells reconstituted with CIP2A-WT (j) or CIP2A-ΔC (k) for indicated structure classes after APH treatment (200 nM, 20 h). Scale bar: 5 µm (confocal) or 500 nm (STED). l Quantification of indicated structures for data from panel (j and k). Bars represent means and SEM, and ‘n’ represents the total number of observed structures, of three biologically independent experiments. Two-tailed unpaired t test was used. Source data are provided as a Source Data file.

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