Fig 1: SFI1 is important for centriole architecture but not for duplication ARepresentative confocal images of expanded duplicating centrioles from siCT- and siSFI1-treated U2OS cells. Cells were stained for SFI1 (green) and a/ß-tubulin (aßTub, magenta). The inset shows a distal position of the mother centriole where SFI1 signal is visible. The white arrowhead indicates the position of SFI1 distal dot in the procentriole of the control cell, which is lost in SFI1-depleted cells (yellow arrowhead). Scale bars: 200 nm.BQuantification of the percentage of SFI1-negative procentrioles.CQuantification of the percentage of duplicating centrioles.DRepresentative confocal images of expanded duplicating centrioles from siCT and siSFI1 U2OS treated cells. Cells were stained for HsSAS-6 (yellow) and a/ß-tubulin (aßTub, magenta). Quantification shows no difference in the percentage of HsSAS-6-positive centrioles in SFI1-depleted cells compared to control cells.ERepresentative confocal images of expanded duplicating centrioles from siCT and siSFI1 U2OS treated cells. Cells were stained for STIL (yellow) and a/ß-tubulin (aßTub, magenta). Quantification shows no difference in the percentage of STIL-positive centrioles in SFI1-depleted cells compared to control cells.FTop views of expanded U2OS centrioles treated with siCT or siSFI1 stained for a/ß-tubulin (aßTub, magenta). Scale bars: 200 nm.GRoundness index of centrioles from siCT- and siSFI1-treated cells.HLength (circle) and diameter (square) of expanded centrioles in siCT- or siSFI1-treated cells.IRepresentative confocal images of expanded U2OS centrioles from siCT- and siSFI1-treated cells stained for a/ß-tubulin (aßTub, magenta). White stars point to a broken microtubule wall. Scale bars: 200 nm.JPercentage of abnormal centrioles in the indicated conditions.KTop views of expanded procentrioles from U2OS cells treated with siCT or siSFI1 and stained for a/ß-tubulin (aßTub, magenta). Scale bars: 200 nm.LRoundness index of procentrioles from siCT- and siSFI1-treated cells.MRepresentative confocal images of expanded U2OS centrioles from siCT- and siSFI1-treated cells stained for a/ß-tubulin (aßTub, magenta) and Centrin (Cetn2/3, gray). White dashed lines delimitate the proximal, central, and distal regions. White star points to the broken microtubule wall. Scale bar: 200 nm.NCentrin coverage (% of the total tubulin length) along the centriole in the indicated conditions.ORepresentative images of expanded U2OS expressing GFP alone or GFP + SFI1-RR and treated with siCT or siSFI1. Cells were stained for a/ß-tubulin (aßTub, magenta) and SFI1 (green) or Centrin 2/3 (gray). The red arrowhead indicates an abnormal centriole. Scale bar: 250 nm.PPercentage of abnormal centrioles in the indicated conditions. Data information: Average ± SD, N, statistical analysis: (B) siCT = 0% ± 0, siSFI1 = 71% ± 11. N = 4 independent experiment (50 centrioles per experiment), Mann–Whitney test (*P = 0.028). (C) siCT = 43% ± 6, siSFI1 = 40% ± 4. N = 7 independent experiment (50 centrioles per experiment), Unpaired t-test (P = 0.3492). (D) siCT = 100% ± 0, siSFI1 = 100% ± 0. N = 3 independent experiments, Mann–Whitney test (P > 0.999). (E) siCT = 100% ± 0, siSFI1 = 100% ± 0. N = 3 independent experiments, Mann–Whitney test (P > 0.999). (G) siCT = 0.88 ± 0.05, siSFI1 = 0.79 ± 0.12. N = 37 and 50 for siCT and siSFI1 respectively from 4 independent experiments, Unpaired t-test (***P = 0.0002). (H) siCT = 417 ± 45 nm (length) and 188 ± 10 nm (diameter), siSFI1 = 423 ± 71 nm (length) and 193.5 ± 13 nm (diameter). N = 50–90 for length and 30–40 for diameter from 4 independent experiments, Mann–Whitney test (P = 0.8440 (length), P = 0.079 (diameter)). (J) siCT = 0% ± 0, siSFI1 = 35.7% ± 11. N = 4 and 6 independent experiments for siCT and siSFI1 respectively, Mann–Whitney test (**P = 0.0095). (L) siCT = 0.89 ± 0.05, siSFI1 = 0.87 ± 0.06. N = 34 and 33 for siCT and siSFI1 respectively from 4 independent experiments, Unpaired t-test (P = 0.376). (N) siCT: 69% ± 13, siSFI1: 47% ± 13, siSFI1 abnormal: 24% ± 20. N = 25, 34, 29 centrioles for siCT, siSFI1, and siSFI1 abnormal respectively from two independent experiments. One-way ANOVA followed by Tukey's post-hoc test (siCT vs. siSFI ***P < 0.0001, siCT vs. siSFI1 abnormal ***P < 0.0001). (P) siCT-GFP: 1.4% ± 1.2, siCT-RR: 1% ± 1.7, siSFI1-GFP: 23.1% ± 5.8, siSFI1-RR: 7.2% ± 2. N = 3 independent experiments (> 50 centrioles/experiments). One-way ANOVA followed by Tukey's multiple comparison (siCT-GFP vs. siSFI1-GFP, ***P = 0.0003; siSFI1-GFP vs. siSFI1-RR, **P = 0.0026).