Figure 3. sas-1 is required maternally for centriole integrity.

(A–E) Images from DIC time-lapse recordings of a wild type embryo (A) and of four embryos resulting from the fertilization of a sas-1(t1476) oocyte by control sperm (B–E). See Movies S1 and S6. Note that in some instances tripolar spindles were observed, presumably because centrioles disintegrate during mitosis (see main text). (F) Schematic showing centriole duplication in the first two cell cycles. C0  =  centrioles contributed by sperm (red), C1  =  centrioles formed next to C0 centrioles (yellow), C2  =  centrioles formed next to C1 centrioles (blue). Blue lines in C1 centrioles indicate formation during the second cell cycle. See also (G–H). (G–H) Visualization of the actual division patterns (G, n = 40 movies) and model 2 (H), with 50’000 simulated embryos. Green  =  Bipolar division, red  =  mono- or tripolar division, black  =  not relevant (descendant of failed division), grey  =  not determined. Insets in (H): a mathematical model of centriole disintegration. This model 2 assumes a probability for the disintegration one cell cycle after C1 formation ( = P_D1) and a different probability for disintegration two cell cycles after C1 formation ( = P_D2). The most likely probabilities given the experimental data (see F) are P_D1 = 0.0625 (0.0253–0.1385, 95% CI) and P_D2 = 0.3028 (0.1465–0.4768, 95% CI). The data is from both sas-1(t1476) (N = 16, 6 at 24°C, 10 at 20°C) and sas-1(t1521) (N = 24, 14 at 24°C, 10 at 20°C) mated to either fog-2 or plg-1 males. (I) Progeny test revealing sas-1 maternal requirement. (J) A sas-1 mutant oocyte fertilized by wild type sperm carrying GFP-SAS-6- labeled centrioles, stained for tubulin (cyan), GFP (yellow) and IFA (magenta). DNA is shown in red. Shown are the top 10/20 planes and bottom 10/20 Z-planes; arrows point to the poles of the tripolar figures (note that one MTOC in ABp is present in both bottom and top planes and only indicated once). Note that whereas the tripolar figures are in ABp and EMS, the paternal GFP-SAS-6 positive centrioles are in ABa and P₂; N = 8 embryos at the four-cell stage that exhibit a phenotype in at the least one blastomere. Given that there are 32 blastomeres in total and that 10 of them exhibited abnormal spindle assembly (6 embryos with one abnormal blastomere, 2 embryos with two abnormal blastomeres –ABp and EMS in one case, ABa and EMS in the other) and assuming that paternally contributed centrioles have a 50% chance of ending up in any blastomere, it follows that the likelihood that the absence of paternal centrioles in those blastomeres that exhibit abnormal spindle assembly is purely due to chance is 0.5¹⁰ = 9.7×10⁻⁴.