Figure 1.
Figure 1. Accelerated mitosis in MAD2L2-deficient cells. (A) MAD2L2 DT40 cells release more rapidly from nocodazole block into G1 than wild-type cells. “n” represents the number of independent experiments. Error bars = SEM; p, unpaired t test. (B) Mitotic aberrations in mad2l2(rev7), rev3, and rev1 DT40 cells. Lagging chromosomes, white segments; anaphase bridges, gray segments. The total number of metaphases scores is indicated. These data were derived from two independent experiments in each of which at least 85 metaphases were counted. Error bars = 1 SD for the independently determined percentages from the two experiments; p, unpaired t test. (C) Confirmation of the effectiveness of the MAD2L2 siRNAs used in this study, showing silencing of only MAD2L2 but not MAD2. The right-hand panel shows the effect of the siRNA against the 3′UTR of human MAD2L2 that does not deplete ectopically expressed human MAD2L2 cDNA carrying a C-terminal myc tag. (D) MAD2L2-depleted U2OS cells release more rapidly than controls from nocodazole block into G1. The percentage of cells in G1 was assessed 60 min after nocodazole release. “n” represents the number of independent experiments. Error bars = SEM; p, unpaired t test. (E) Frames from time-lapse movies of control and siMAD2L2 U2OS cells. NEBD, nuclear envelope breakdown. Note the lagging chromosome at 30 min in the siMAD2L2 frames. Example movies can be found online (Videos 1–4). The white bar in the first frame represents 5 µm. (F) Quantification of the time taken for control (Control siRNA; solid black line), MAD2L2-depleted (siMAD2L2; solid red line), complemented (siMAD2L2 + hMAD2L2; dashed black line), and REV3-depleted (siREV3; solid blue line) U2OS cells expressing mCherry-H2B to complete mitosis, assessed by time-lapse video microscopy (1 frame every 5 min). The plot shows the cumulative percentage of cells that completed mitosis, measured from NEBD to cytokinesis. “n” represents the number of cells examined, collected from at least three independent experiments. The P-value to test whether the distribution of times in siMAD2L2-treated cells is distinct from controls was calculated by the Kolmogorov-Smirnov test.

Accelerated mitosis in MAD2L2-deficient cells. (A) MAD2L2 DT40 cells release more rapidly from nocodazole block into G1 than wild-type cells. “n” represents the number of independent experiments. Error bars = SEM; p, unpaired t test. (B) Mitotic aberrations in mad2l2(rev7), rev3, and rev1 DT40 cells. Lagging chromosomes, white segments; anaphase bridges, gray segments. The total number of metaphases scores is indicated. These data were derived from two independent experiments in each of which at least 85 metaphases were counted. Error bars = 1 SD for the independently determined percentages from the two experiments; p, unpaired t test. (C) Confirmation of the effectiveness of the MAD2L2 siRNAs used in this study, showing silencing of only MAD2L2 but not MAD2. The right-hand panel shows the effect of the siRNA against the 3′UTR of human MAD2L2 that does not deplete ectopically expressed human MAD2L2 cDNA carrying a C-terminal myc tag. (D) MAD2L2-depleted U2OS cells release more rapidly than controls from nocodazole block into G1. The percentage of cells in G1 was assessed 60 min after nocodazole release. “n” represents the number of independent experiments. Error bars = SEM; p, unpaired t test. (E) Frames from time-lapse movies of control and siMAD2L2 U2OS cells. NEBD, nuclear envelope breakdown. Note the lagging chromosome at 30 min in the siMAD2L2 frames. Example movies can be found online (Videos 1–4). The white bar in the first frame represents 5 µm. (F) Quantification of the time taken for control (Control siRNA; solid black line), MAD2L2-depleted (siMAD2L2; solid red line), complemented (siMAD2L2 + hMAD2L2; dashed black line), and REV3-depleted (siREV3; solid blue line) U2OS cells expressing mCherry-H2B to complete mitosis, assessed by time-lapse video microscopy (1 frame every 5 min). The plot shows the cumulative percentage of cells that completed mitosis, measured from NEBD to cytokinesis. “n” represents the number of cells examined, collected from at least three independent experiments. The P-value to test whether the distribution of times in siMAD2L2-treated cells is distinct from controls was calculated by the Kolmogorov-Smirnov test.

This Feature Is Available To Subscribers Only

or Create an Account

Close Modal
Close Modal