Fig. 1: DONSON and Cdc6, DNA replication factors, maintain centriole engagement and proper centrosome number.

a List of genes involved in microcephalic primordial dwarfism (MPD) based on previous studies²². The genes involved in MPD are mostly classified as centrosomal proteins, DNA replication components, or a regulator for DNA damage response. b The number of Cep192 foci upon siRNA treatment against the indicated proteins. c, Histograms represent the frequency of interphase cells with >2 Cep192 foci observed in (b). n = 3 independent experiments, 50 cells for each. d Schematic of the DONSON gene, indicating mutations observed in cells from MPD patient 2 (P2)²⁵. The genomic structure is based on the longest ORF, containing ten coding exons (white rectangles) (NCBI NM_017613.3). e The number of Cep192 foci in hTERT-immortalized fibroblasts derived from the MPD patient 2. Fibroblasts were infected with pMSCV-empty-vector or pMSCV-DONSON and immunostained with antibodies against Cep192 (red). Arrowheads indicate centrosomes. f Histograms represent the frequency of interphase cells with the indicated phenotypes observed in (e). n = 3 independent experiments, 50 cells for each. g The number of centrin and Cep192 foci in G1 or S/G2 phase upon DONSON or Cdc6 depletion. HeLa cells were treated with siRNA treatment against the indicated proteins for 48 h and 10 μM EdU for 30 min. CENP-F and EdU double-negative cells were classified as G1 phase cells, and others as S or G2 phase cells. Arrowheads indicate centrioles. h Histograms represent the frequency of cells with the indicated phenotype observed in (g). n = 3 independent experiments, 30 cells for each. i Time-lapse observation of centriole disengagement in DONSON-depleted cells. HeLa GFP-centrin-1 cells were visualized every 20 min for 48 h after 24 h treatment with siRNA and 3 h treatment with 100 nM SiR-DNA. Arrowheads indicate engaged centriole pairs, and left–right arrows indicate precociously disengaged centrioles. j Cumulative scatterplot indicates the duration from nuclear envelope breakdown (NEBD) to centriole disengagement observed in (i). k Schematic model of the phenotypes of DONSON or Cdc6 depletion. All scale bars, 5 μm. Values are mean percentages ± s.d. Tukey’s multiple comparison test was used in c, and a two-tailed, unpaired Student’s t-test was used in (f) to obtain P-value. Throughout all the figures in this study, the single-channel images were generated by displaying only the signal from one channel extracted directly from the original merged representative image, without any adjustment to intensity or contrast. The insets represent magnified views of regions taken directly from either the merged image or the corresponding single-channel image, also without any further modification. The insets are displayed in the following order from top to bottom: merged, green, red, and cyan. All source data are provided as a Source Data file.