Figure 2.
Figure 2. Depletion of Mcm21R reduces the number of stable k-fibers. (A) Successive frames every 30 or 40 s after photoactivation of stable PA-GFP–α-tubulin/hTERT-RPE1 cells treated either with siControl, siMcm21R, or siMCAK + siKif2A RNAs. Green arrowheads mark the initial position of a photoactivated mark, and red arrowheads show the final position of the mark. (B) Quantification of poleward MT flux rate in cells treated with siControl (n = 9), siMcm21R (n = 9), or siMCAK + siKif2A (n = 5) RNAs. (C–E) Quantification of stable k-fibers in cold-treated cells (for methodology, see Fig. S2, available). Bar graphs indicate the percentage of kinetochores lacking attached MTs (−), having a weak attachment (+), a medium attachment (++), or a strong attachment (+++) in siControl (n = 354)-, siNuf2R (n = 204)-, and siMcm21R (n = 273)-treated cells with bipolar spindles (C), in siMcm21R (n = 215)- and siEg5 (n = 212)-treated cells with monopolar spindles (D), and in siControl (n = 354)-, siMcm21R (n = 273)-, siMCAK (n = 238)-, and siMcm21R + siMCAK (n = 219)–treated cells with bipolar spindles (E). (F) Representative images from C and E of cold-treated cells stained with α-tubulin antibodies (MTs; green) and CREST antisera (kinetochores; red). Error bars represent SEM. Bars, 10 µm.

Depletion of Mcm21R reduces the number of stable k-fibers. (A) Successive frames every 30 or 40 s after photoactivation of stable PA-GFP–α-tubulin/hTERT-RPE1 cells treated either with siControl, siMcm21R, or siMCAK + siKif2A RNAs. Green arrowheads mark the initial position of a photoactivated mark, and red arrowheads show the final position of the mark. (B) Quantification of poleward MT flux rate in cells treated with siControl (n = 9), siMcm21R (n = 9), or siMCAK + siKif2A (n = 5) RNAs. (C–E) Quantification of stable k-fibers in cold-treated cells (for methodology, see Fig. S2, available). Bar graphs indicate the percentage of kinetochores lacking attached MTs (−), having a weak attachment (+), a medium attachment (++), or a strong attachment (+++) in siControl (n = 354)-, siNuf2R (n = 204)-, and siMcm21R (n = 273)-treated cells with bipolar spindles (C), in siMcm21R (n = 215)- and siEg5 (n = 212)-treated cells with monopolar spindles (D), and in siControl (n = 354)-, siMcm21R (n = 273)-, siMCAK (n = 238)-, and siMcm21R + siMCAK (n = 219)–treated cells with bipolar spindles (E). (F) Representative images from C and E of cold-treated cells stained with α-tubulin antibodies (MTs; green) and CREST antisera (kinetochores; red). Error bars represent SEM. Bars, 10 µm.

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