Figure S5.
Stable transgene expression in RPE1 cells and fate of misaligned chromosomes in RPE1 cells stably expressing GFP-Sec61β. (A–D) Western blots to examine expression of proteins in parental RPE1 cells or clonal cells stably expressing GFP-Sec61β alone or with Histone3.2-mCherry, LBR-mCherry, or mCherry-BAF, as indicated. Membranes were probed for GFP, Sec61β, mCherry, LBR, BAF. Actin or tubulin is shown as a loading control. Green or red arrowheads indicate the expected position of GFP- or mCherry-tagged protein; black arrowheads indicate the untagged protein. (E) Mitotic timing of RPE1 cells stably expressing transgenes. Cumulative frequencies for NEB to metaphase, metaphase to anaphase, and NEB to anaphase are shown. Parental, n = 69; GFP-Sec61β alone, n = 52; GFP-Sec61β and LBR-mCherry, n = 66; GFP-Sec61β and mCherry-BAF, n = 51. (F) Sankey diagram to show the fate (right) of RPE1 cells in each of the three metaphase classes (left). Fates include normal division, micronuclei formation, death, and other defects (lagging chromosome, cytokinesis failure). Note that the fate of cells (and not chromosomes) is tracked. LBR-mCherry/GFP-Sec61β, n = 51; mCherry-BAF/GFP-Sec61β, n = 67; pooled from three experiments. (G) Sankey diagram to show the fate (right) of chromosomes in each of the three metaphase classes (left) after GSK923295 pretreatment. Fates include rescue, micronuclei formation, death, and other defects (lagging chromosome, cytokinesis failure). Number of chromosomes: free, 146; ensheathed, 207; lagging, 9. The same dataset was analyzed for the outcome of cells (classified by the final misaligned chromosome) in Fig. 4. Note that ensheathed chromosomes at metaphase that were rescued all became “free” chromosomes before rescue. Source data are available for this figure: SourceData FS5.

Stable transgene expression in RPE1 cells and fate of misaligned chromosomes in RPE1 cells stably expressing GFP-Sec61β. (A–D) Western blots to examine expression of proteins in parental RPE1 cells or clonal cells stably expressing GFP-Sec61β alone or with Histone3.2-mCherry, LBR-mCherry, or mCherry-BAF, as indicated. Membranes were probed for GFP, Sec61β, mCherry, LBR, BAF. Actin or tubulin is shown as a loading control. Green or red arrowheads indicate the expected position of GFP- or mCherry-tagged protein; black arrowheads indicate the untagged protein. (E) Mitotic timing of RPE1 cells stably expressing transgenes. Cumulative frequencies for NEB to metaphase, metaphase to anaphase, and NEB to anaphase are shown. Parental, n = 69; GFP-Sec61β alone, n = 52; GFP-Sec61β and LBR-mCherry, n = 66; GFP-Sec61β and mCherry-BAF, n = 51. (F) Sankey diagram to show the fate (right) of RPE1 cells in each of the three metaphase classes (left). Fates include normal division, micronuclei formation, death, and other defects (lagging chromosome, cytokinesis failure). Note that the fate of cells (and not chromosomes) is tracked. LBR-mCherry/GFP-Sec61β, n = 51; mCherry-BAF/GFP-Sec61β, n = 67; pooled from three experiments. (G) Sankey diagram to show the fate (right) of chromosomes in each of the three metaphase classes (left) after GSK923295 pretreatment. Fates include rescue, micronuclei formation, death, and other defects (lagging chromosome, cytokinesis failure). Number of chromosomes: free, 146; ensheathed, 207; lagging, 9. The same dataset was analyzed for the outcome of cells (classified by the final misaligned chromosome) in Fig. 4. Note that ensheathed chromosomes at metaphase that were rescued all became “free” chromosomes before rescue. Source data are available for this figure: SourceData FS5.

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