DEV019075 Supplementary Material

Files in this Data Supplement:

• Supplemental Tables 1-3 (Adobe PDF) - Adobe PDF

• Supplemental Figure S1 -

  Fig. S1. PLK-1 asymmetry regulates the time difference independently of ATL-1. (A-D) Images from time-lapse fluorescent microscopy of wild-type (A), mild plk-1(RNAi) (B), atl-1(tm853) (C) or atl-1(tm853) embryos treated with mild plk-1(RNAi) (D) and expressing GFP-Histone 2B (see corresponding Movies 11-14). Arrowheads with an unbroken outline indicate condensed chromosomes or non-separated sister chromatids, arrowheads with a broken outline indicate separated sister chromatids. The metaphase to anaphase transition in P₀ is defined as t=0, and the time after that is indicated in seconds. In B,D, chromosomes in AB are slightly out of focus in the central panels (see Movies 12 and 14 for fuller picture). (E) Average ratios±s.e.m. of the duration of the entire cell cycle in P₁ over the duration of the entire cell cycle in AB in embryos of the indicated genotype (I+M) _P1/(I+M) _AB. The star indicates that the difference from wild type is statistically significant. See Table S3 for numerical values and statistical analysis.

• Supplemental Figure S2 -

  Fig. S2. NCC-1 is uniformly distributed, but differentially activated in two-cell stage embryos. (A-C) Wild-type two-cell stage embryos in interphase (A), early prophase (B) or AB metaphase (C) stained for NCC-1 (shown alone in the left panel and in red in the merged panels), α-tubulin (green) and DNA (blue). NCC-1 is equally distributed in the nucleus and the cytoplasm from two blastomeres before NEBD. (D-F) Wild-type two-cell stage embryos in interphase (D), early prophase (E) or AB prometaphase (F) stained for phospho-tyrosine 15 (shown alone in the left panel and in red in the merged panels), α-tubulin (green) and DNA (blue). To clearly show the difference in nuclear signal between AB and P₁, maximal intensity projections are shown. Active NCC-1 is inferred from the disappearance of the pT15 signal. In early prophase, the pT15 signal is already weaker in AB than in P₁ (E), whereas later it has disappeared from AB, but not from P₁ (F), indicating that NCC-1 is activated earlier in AB.

• Movie 1
• Movie 2 -

  Movies 1 and 2. Time lapse DIC microscopy of wild-type (Movie 1) and par-2(RNAi) (Movie 2) filmed side by side to compare the timing of entry into mitosis in AB and P₁ of the wild-type embryo with that of the two blastomeres of par-2(RNAi) embryos. In these and other movies, embryos are oriented with anterior towards the left and posterior towards the right. Movies were captured every 5 seconds and are played at 10 frames per second. Elapsed time since cleavage furrow ingression in P₀ is shown in seconds.

• Movie 3
• Movie 4 -

  Movie 3 and 4. Time lapse DIC microscopy of wild-type (Movie 3) and par-3(it71) (Movie 4) filmed side by side to compare the timing of entry into mitosis in AB and P₁ of the wild-type embryo with that of the two blastomeres of par-3(it71) embryos.

• Movie 5
• Movie 6
• Movie 7
• Movie 8
• Movie 9
• Movie 10 -

  Movies 5-10. Time lapse DIC microscopy of wild-type (Movie 5), mild plk-1(RNAi) (Movie 6), ncc-1(RNAi) (Movie 7), atp-2(RNAi) (Movie 8), atl-1(tm853) (Movie 9) or atl-1(tm853) embryos treated with mild plk-1(RNAi) (Movie 10).

• Movie 11
• Movie 12
• Movie 13
• Movie 14 -

  Movies 11-14. Time lapse fluorescent microscopy of wild-type (Movie 11), mild plk-1(RNAi) (Movie 12), atl-1(tm853) (Movie 13), and atl-1(tm853) embryos treated with mild plk-1(RNAi) (Movie 14). Movies were captured every 10 seconds and played at 10 frames per second. Elapsed time since metaphase to anaphase transition in P₀ is shown in seconds. The shutter was closed from shortly after this transition until shortly before NEBD of AB to minimize exposure to the fluorescent source. The movies 4 have been rotated, resulting in the presence of black triangles on the edges of the recordings.