Figure 1.
Velocities of poleward-segregating acentrics decrease as acentrics pass through nuclear envelope channels.(A and B) Stills from videos of mitotic neuroblasts expressing H2Av-RFP (magenta) and GFP-NLS (green; A) or H2Av-RFP, GFP-NLS, and I-CreI (B). Nuclear envelope reassembly initiation (arrowheads) began later in the division with late-segregating acentrics (arrows). Despite this delay, the delay in acentric segregation is much more severe, and thus acentrics were still far from the daughter nuclei when the nuclear envelope initiated reassembly. Time is written in seconds after anaphase onset. Scale bars are 2 µm. (C) GFP-NLS intensity on daughter nuclei from control neuroblasts (black line; n = 26) and neuroblasts expressing I-CreI (purple line; n = 13). Lines represent averages. Dark-shaded regions represent ± the standard error. Light-shaded regions represent ± twice the standard error. Dashed lines indicate time of nuclear envelope reassembly initiation. On average, nuclear envelope reassembly initiated later in neuroblasts expressing I-CreI (difference in the times of GFP-NLS accumulation on nuclei between control and I-CreI neuroblasts measured as significant by a two-sided Mann–Whitney–Wilcoxon test: P = 0.008). (D) Distance of acentrics from control daughter nuclei (black line; n = 13) and GFP-NLS intensity on daughter nuclei from neuroblasts expressing I-CreI (purple line; n = 13). Lines represent averages. Dark-shaded regions represent ± the standard error. Light-shaded regions represent ± twice the standard error. Dashed line indicates the distance of acentrics from daughter nuclei at the time of nuclear envelope reassembly initiation. On average, acentrics were >2.5 µm away from daughter nuclei when the nuclear envelope initiated reassembly. (E) Percentage of neuroblast divisions in which acentrics formed micronuclei when segregating equally to each daughter cell (left; n = 14) and when segregating unequally to each daughter cell (right; n = 11). Additionally, stills from videos of mitotic neuroblasts expressing I-CreI, H2Av-RFP (magenta), and Lamin-GFP (green) when acentrics (arrows) segregated equally (left) and unequally (right) and entered daughter nuclei through channels in the nuclear envelope (arrowheads). Scale bar is 2 µm. (F) Stills from a video (Video 1) of a mitotic neuroblast expressing I-CreI, H2Av-RFP (magenta), and Lamin-GFP (green). Acentrics (arrows) moved off the metaphase plate and toward daughter nuclei during the initial poleward phase (0–270 s) before entering nuclear envelope channels (arrowheads) and being surrounded by a complete nuclear envelope (270–396 s) during the channel passage phase. Time is written in seconds after initial acentric poleward movement. Scale bar is 2 µm. Yellow dashed boxes indicate magnified regions. (G) Diagram illustrating how acentric velocities were measured. The time between when acentrics first move poleward and when they enter nuclear envelope channels is deemed the initial poleward phase. The time between when acentrics first enter channels and when the nuclear envelope reassembles completely around them is termed the channel passage phase. Distance was measured between the furthest point on the acentric and the closest point on the daughter nucleus (red brackets). Velocity was calculated as the difference in measured distances at the beginning and ending of each phase divided by the time it took for acentrics to complete each phase. (H) Velocities of acentrics (n = 30) during their initial poleward phase (left) and during their channel passage phase (right). Each dot represents one acentric. Lines connect the measured initial poleward and channel passage velocities of the same acentric. The velocities of 24 of 30 acentrics decreased from their initial poleward phase to their channel passage phase (blue dots). The velocities of 6 of 30 acentrics increased from their initial poleward phase to their channel passage phase (red dots). Boxes represent interquartile ranges and lines represent medians of the measured data. Asterisks indicate statistical significance (P = 0.0003) determined by a two-sided Wilcoxon signed-rank test.

Velocities of poleward-segregating acentrics decrease as acentrics pass through nuclear envelope channels. (A and B) Stills from videos of mitotic neuroblasts expressing H2Av-RFP (magenta) and GFP-NLS (green; A) or H2Av-RFP, GFP-NLS, and I-CreI (B). Nuclear envelope reassembly initiation (arrowheads) began later in the division with late-segregating acentrics (arrows). Despite this delay, the delay in acentric segregation is much more severe, and thus acentrics were still far from the daughter nuclei when the nuclear envelope initiated reassembly. Time is written in seconds after anaphase onset. Scale bars are 2 µm. (C) GFP-NLS intensity on daughter nuclei from control neuroblasts (black line; n = 26) and neuroblasts expressing I-CreI (purple line; n = 13). Lines represent averages. Dark-shaded regions represent ± the standard error. Light-shaded regions represent ± twice the standard error. Dashed lines indicate time of nuclear envelope reassembly initiation. On average, nuclear envelope reassembly initiated later in neuroblasts expressing I-CreI (difference in the times of GFP-NLS accumulation on nuclei between control and I-CreI neuroblasts measured as significant by a two-sided Mann–Whitney–Wilcoxon test: P = 0.008). (D) Distance of acentrics from control daughter nuclei (black line; n = 13) and GFP-NLS intensity on daughter nuclei from neuroblasts expressing I-CreI (purple line; n = 13). Lines represent averages. Dark-shaded regions represent ± the standard error. Light-shaded regions represent ± twice the standard error. Dashed line indicates the distance of acentrics from daughter nuclei at the time of nuclear envelope reassembly initiation. On average, acentrics were >2.5 µm away from daughter nuclei when the nuclear envelope initiated reassembly. (E) Percentage of neuroblast divisions in which acentrics formed micronuclei when segregating equally to each daughter cell (left; n = 14) and when segregating unequally to each daughter cell (right; n = 11). Additionally, stills from videos of mitotic neuroblasts expressing I-CreI, H2Av-RFP (magenta), and Lamin-GFP (green) when acentrics (arrows) segregated equally (left) and unequally (right) and entered daughter nuclei through channels in the nuclear envelope (arrowheads). Scale bar is 2 µm. (F) Stills from a video (Video 1) of a mitotic neuroblast expressing I-CreI, H2Av-RFP (magenta), and Lamin-GFP (green). Acentrics (arrows) moved off the metaphase plate and toward daughter nuclei during the initial poleward phase (0–270 s) before entering nuclear envelope channels (arrowheads) and being surrounded by a complete nuclear envelope (270–396 s) during the channel passage phase. Time is written in seconds after initial acentric poleward movement. Scale bar is 2 µm. Yellow dashed boxes indicate magnified regions. (G) Diagram illustrating how acentric velocities were measured. The time between when acentrics first move poleward and when they enter nuclear envelope channels is deemed the initial poleward phase. The time between when acentrics first enter channels and when the nuclear envelope reassembles completely around them is termed the channel passage phase. Distance was measured between the furthest point on the acentric and the closest point on the daughter nucleus (red brackets). Velocity was calculated as the difference in measured distances at the beginning and ending of each phase divided by the time it took for acentrics to complete each phase. (H) Velocities of acentrics (n = 30) during their initial poleward phase (left) and during their channel passage phase (right). Each dot represents one acentric. Lines connect the measured initial poleward and channel passage velocities of the same acentric. The velocities of 24 of 30 acentrics decreased from their initial poleward phase to their channel passage phase (blue dots). The velocities of 6 of 30 acentrics increased from their initial poleward phase to their channel passage phase (red dots). Boxes represent interquartile ranges and lines represent medians of the measured data. Asterisks indicate statistical significance (P = 0.0003) determined by a two-sided Wilcoxon signed-rank test.

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