![Central spindle assembly is not required for successful cytokinesis.(A) Method used for analysis (left; d, cell diameter) and quantification (right) of contractile ring constriction during cytokinesis in embryos expressing mCherry::H2B and GFP::PHPLC1δ1 (top). Time relative to anaphase onset; only hcp-1/2 disrupted embryos that formed a metaphase plate were analyzed (see Materials and methods and Fig. S3 A); error bars represent the SEM; n is indicated in the key [bottom; n = 13 control(RNAi), n = 7 hcp-1(RNAi); hcp-2Δ, n = 7 hcp-1Δ; hcp-2(RNAi), n = 12 hcp-1Δ, n = 8 hcp-2Δ, n = 9 spd-1(oj5ts), n = 9 cls-2(RNAi), and n = 9 zen-4(or153ts)]; see Video 6. Quantification of cytokinesis success (bottom). (B) Quantification of the average peak rate of contractile ring constriction between 80% and 30% of initial embryo diameter for all dividing embryos in A; genotypes indicated below graph. Statistical significance was determined by one-way ANOVA with Tukey’s multiple comparison test; error bars represent the SD. (C) Schematic of line scan analysis of CYK-4::mNG fluorescence intensity at the central spindle and midbody on sum projections of all Z-planes; white dashed line outlines the embryo. (D) Schematic of selected region shown (top) and representative time-lapse pseudokymographs of the division plane in embryos expressing CYK-4::mNG (green), mCherry::H2B, and mCherry::PHPLC1δ1 (magenta; bottom, left); time at top. t = 0 s is metaphase before chromosome segregation; maximum projection is shown; see Video 7. Right: Quantification of CYK-4::mNG fluorescence intensity along a central line scan at 60 s and 240 s after chromosome segregation to measure central spindle and midbody assembly, respectively. Individual (thin lines) and average (bold lines) line scans are shown for each genotype. n is listed to right side of the 240 s graphs [n = 12 control(RNAi), n = 13 hcp-1/2(RNAi), n = 10 hcp-1(RNAi), n = 14 hcp-2(RNAi), and n = 12 spd-1(RNAi)]. Average levels in control(RNAi) embryos are shown on each graph for reference (gray). Error bars represent the SEM; white scale bars, 10 µm; black scale bar in D, 15 s. n.s., P ≥ 0.05; ****, P < 0.0001.](https://cdn.rupress.org/rup/content_public/journal/jcb/221/3/10.1083_jcb.202011085/4/jcb_202011085_fig3.png?Expires=1773591846&Signature=0G~IkhvHluaI0feilmdcuy-3~58ajjRSS04GCUkeiRRiJA0Hg3QnS5hoRnO3zs-jEs5E5T4-De5LSH-XdLUNZunXEdIhns8iEHJ~AXv7co4~3jgqUusT-TwE1ZXfEFxchZJxbEDct7qJTOwycWqyz7J47f8pYKpb3A80OJ539Aun5kBaa6t932jckxaeU5UpkYTGFnxGsDkAd-NAA-Fd7P6qV-pinbMndLiFqN~MYd4~9X3cESpQNJagVVUZiKnPn0oNbHkIudC9IlG7mXMZARorGsxdrSEAIPRCKDgFKNGftvJ7Zuqt0MrYznGh7NshVx8TvM2IHPWARttMV8Q4qg__&Key-Pair-Id=APKAIE5G5CRDK6RD3PGA)
Central spindle assembly is not required for successful cytokinesis. (A) Method used for analysis (left; d, cell diameter) and quantification (right) of contractile ring constriction during cytokinesis in embryos expressing mCherry::H2B and GFP::PHPLC1δ1 (top). Time relative to anaphase onset; only hcp-1/2 disrupted embryos that formed a metaphase plate were analyzed (see Materials and methods and Fig. S3 A); error bars represent the SEM; n is indicated in the key [bottom; n = 13 control(RNAi), n = 7 hcp-1(RNAi); hcp-2Δ, n = 7 hcp-1Δ; hcp-2(RNAi), n = 12 hcp-1Δ, n = 8 hcp-2Δ, n = 9 spd-1(oj5ts), n = 9 cls-2(RNAi), and n = 9 zen-4(or153ts)]; see Video 6. Quantification of cytokinesis success (bottom). (B) Quantification of the average peak rate of contractile ring constriction between 80% and 30% of initial embryo diameter for all dividing embryos in A; genotypes indicated below graph. Statistical significance was determined by one-way ANOVA with Tukey’s multiple comparison test; error bars represent the SD. (C) Schematic of line scan analysis of CYK-4::mNG fluorescence intensity at the central spindle and midbody on sum projections of all Z-planes; white dashed line outlines the embryo. (D) Schematic of selected region shown (top) and representative time-lapse pseudokymographs of the division plane in embryos expressing CYK-4::mNG (green), mCherry::H2B, and mCherry::PHPLC1δ1 (magenta; bottom, left); time at top. t = 0 s is metaphase before chromosome segregation; maximum projection is shown; see Video 7. Right: Quantification of CYK-4::mNG fluorescence intensity along a central line scan at 60 s and 240 s after chromosome segregation to measure central spindle and midbody assembly, respectively. Individual (thin lines) and average (bold lines) line scans are shown for each genotype. n is listed to right side of the 240 s graphs [n = 12 control(RNAi), n = 13 hcp-1/2(RNAi), n = 10 hcp-1(RNAi), n = 14 hcp-2(RNAi), and n = 12 spd-1(RNAi)]. Average levels in control(RNAi) embryos are shown on each graph for reference (gray). Error bars represent the SEM; white scale bars, 10 µm; black scale bar in D, 15 s. n.s., P ≥ 0.05; ****, P < 0.0001.
