Supplemental Materials
This article contains the following supporting material:
- Movie01 - Movie S1 Confocal timelapse imaging of transgenically expressed Chromator-GFP and tubulin-mCherry in a syncytial Drosophila embryo (Leica TCS SP5 tandem scanning microscope). The image sequence illustrates the dynamic relationship of the spindle matrix protein Chromator (in green) relative to microtubule spindle formation (in red) throughout mitosis. The results suggest that the spindle matrix is present within the nucleus prior to invasion of microtubules into the nuclear space. The timelapse covers a period of 6 min 17 s. (Quicktime; 8.3 MB).
- Movie02 - Movie S2 Confocal timelapse imaging of transgenically expressed Jupiter-GFP and tubulin-mCherry in a syncytial Drosophila embryo (Leica TCS SP5 tandem scanning microscope). The image sequence illustrates the dynamic relationship of the MAP Jupiter (in green) relative to microtubule spindle formation (in red) illustrating their co-localization throughtout mitosis. The timelapse covers a period of 7 min 40 s. (Quicktime; 7.7 MB).
- Movie03 - Movie S3 Timelapse imaging of transgenically expressed Chromator-GFP in a syncytial Drosophila embryo using spinning disk confocal microscopy (Ultraview, Perkin Elmer). While Chromator is present throughout the spindle region its poleward boundary does not extend all the way to the centrosome. The timelapse covers a period of 8 min 36 s. (Quicktime; 9.0 MB).
- Movie04 - Movie S4 Confocal timelapse imaging of transgenically expressed Chromator-GFP (in green) and tubulin-mCherry (in red) in a syncytial Drosophila embryo after injection of colchicine at interphase (Leica TCS SP5 tandem scanning microscope). Under these conditions Chromator still relocates from the chromosomes to the matrix; however, in the absence of microtubules the Chromator-defined matrix did not undergo any dynamic changes but instead statically embedded the condensed chromosomes for extended periods. The timelapse covers a period of 8 min 27 s. (Quicktime; 9.7 MB).
- Movie05 - Movie S5 Confocal timelapse imaging of transgenically expressed Chromator-GFP (in green) and histone H2Av-RFP (in red) in a syncytial Drosophila embryo after injection of colchicine at interphase (Leica TCS SP5 tandem scanning microscope). Under these conditions Chromator still relocates from the chromosomes to the matrix; however, in the absence of microtubules the Chromator-defined matrix did not undergo any dynamic changes but instead statically embedded the condensed chromosomes for extended periods. The timelapse covers a period of 7 min. (Quicktime; 8.4 MB).
- Movie06 - Movie S6 Confocal timelapse imaging of transgenically expressed Chromator-GFP (in green) and tubulin-mCherry (in red) in a syncytial Drosophila control embryo after injection of PEM buffer with 1% DMSO at interphase (Leica TCS SP5 tandem scanning microscope). The control embryo underwent normal mitosis indistinguishable from wild-type preparations. The timelapse covers a period of 6 min 44 s. (Quicktime; 9.5 MB).
- Movie07 - Movie S7 Confocal timelapse imaging of transgenically expressed tubulin-mCherry (in red) in a syncytial Drosophila embryo after injection of 70 kDa fluorescine-labeled dextran (in green) (Leica TCS SP5 tandem scanning microscope). The image sequence shows that 70 kDa dextran get enriched in the nuclear space prior to microtubule polymerization, and its dynamics during mitosis until the end of telophase where it gets excluded from the forming daugther nuclei (Movie S7) closely resembles that of the spindle matrix proteins Chromator and Megator (Movie S1 and S8) The timelapse covers a period of 5 min 33 s. (Quicktime; 7.5 MB).
- Movie08 - Movie S8 Confocal timelapse imaging of transgenically expressed full-length Megator-YFP (in green) and histone H2Av-RFP (in red) in a syncytial Drosophila embryo (Ultraview spinning disk confocal microscope, Perkin Elmer). The image sequence illustrates that Megator localizes to the nuclear interior as well as the nuclear rim at interphase and to the spindle matrix at metaphase. The timelapse covers a period of 7 min 24 s. (Quicktime; 8.6 MB).
- Movie09 - Movie S9 Confocal timelapse imaging of transgenically expressed Megator-CTD (in green) and histone H2Av-RFP (in red) in a syncytial Drosophila embryo (Leica TCS SP5 tandem scanning microscope). The image sequence illustrates that Megator-CTD is diffusively present in the nucleoplasm without detectable nuclear rim localization at interphase and is absent from the spindle region at metaphase The timelapse covers a period of 9 min 15 s. (Quicktime; 6.7 MB).
- Movie10 - Movie S10 Confocal timelapse imaging of transgenically expressed full-length Megator-YFP (in green) and histone H2Av-RFP (in red) in a syncytial Drosophila embryo after injection of colchicine at interphase (Leica TCS SP5 tandem scanning microscope). Under these conditions Megator still relocates to the matrix; however, in the absence of microtubules the Megator-defined matrix did not undergo any dynamic changes but instead statically embedded the condensed chromosomes for extended periods. The timelapse covers a period of 13 min. (Quicktime; 7.1 MB)
- Movie11 - Movie S11 Confocal timelapse imaging of transgenically expressed Megator-CTD (in green) and histone H2Av-RFP (in red) in a syncytial Drosophila embryo after injection of colchicine at interphase (Leica TCS SP5 tandem scanning microscope). Under these conditions Megator-CTD disperses on a rapid time-scale in less than 2 min after NE breakdown. The timelapse covers a period of 4 min 18 s. (Quicktime; 7.7 MB)
- Movie12 - Movie S12 Confocal timelapse imaging of transgenically expressed Chromator-GFP (in green) and tubulin-mCherry (in red) in a syncytial Drosophila embryo after injection of colchicine at metaphase (Leica TCS SP5 tandem scanning microscope). The image sequence shows that as the microtubules undergo depolymerization the Chromator-defined matrix contracts and coalesces around the chromosomes. The timelapse covers a period of 5 min 30 s. (Quicktime; 8.4 MB).
- Movie13 - Movie S13 Confocal timelapse imaging of transgenically expressed full-length Megator-YFP (in green) and tubulin-mCherry (in red) in a syncytial Drosophila embryo after injection of taxol at metaphase (Leica TCS SP5 tandem scanning microscope). The image sequence shows that under these conditions both the spindle matrix and microtubules do not undergo any dynamic changes but mantain their metaphase fusiform spindle morphology. The timelapse covers a period of 14 min. (Quicktime; 9.6 MB).