Figure 5. Nuclear bleb formation corresponds to nuclear volume expansion and decreased migration speed. (a) Image sequence depicting the growth and subsequent collapse of the nuclear envelope of HT-1080 cells during nuclear blebbing and rupture in confinement. Arrows indicate time frame of rupture events. Scale bar, 10 µm. (b) Time-dependent change of nuclear longitudinal area of an H2B-mCherry labeled HT-1080 cell during migration in confinement. Arrows indicate incidents of nuclear rupture (top). Time-dependent change of nuclear longitudinal area of an H2B-mCherry labeled HT-1080 cell over the course of one nuclear rupture and repair (bottom). (c) Nuclear volume, measured from confocal Z-stacks of HT-1080 H2B-mCherry nuclear bleb–bearing and nuclear bleb–free cells in confinement (n ≥ 42 cells from four independent experiments). (d) Change in nuclear volume over time of migration through confined channels for representative bleb-bearing and bleb-free cell, as measured from confocal Z-stacks of HT-1080 H2B-mCherry cells. (e) Change in volume over time of migration through confined channels of the nuclear bodies and nuclear blebs, as measured from confocal Z-stacks of HT-1080 H2B-mCherry cells (n = 9 cells from two independent experiments). (f) Percentage of nuclear volume increase relative to the total initial nuclear volume for the total nucleus, nuclear bleb, and nuclear body (n = 8 cells from two independent experiments). (g) Nuclear volume, measured from confocal Z-stacks of HT-1080 H2B-mCherry control and Y27632-treated cells and TRE GFP control and Q63L infected cells in confinement (n ≥ 50 cells from three or more independent experiments). (h) Nuclear volume, measured from confocal Z-stacks of HT-1080 H2B-mCherry control and LMB-treated cells on 2D (n ≥ 52 cells from 3 independent experiments). (i) Nuclear volume, measured from confocal Z-stacks of HT-1080 H2B-mCherry control and LMB-treated cells in confinement (n ≥ 53 cells from three independent experiments). (j) Cell migration speed as a function of nuclear volume as predicted by mathematical modeling based on lubrication theory. (k) HT-1080 speed plotted as a function of nuclear longitudinal area (n = 120 cells from four independent experiments). Solid line represents the best fit, and dotted lines represent the 95% CI. (l) Schematic describing the proposed mechanism for nuclear bleb formation. P denotes pressure. *, P < 0.05 relative to vehicle/scramble control; #, P < 0.05 relative to nuclear bleb bearing; §, P < 0.05 relative to TRE GFP control. Scale bar, 10 µm.
Figure 5.

Nuclear bleb formation corresponds to nuclear volume expansion and decreased migration speed. (a) Image sequence depicting the growth and subsequent collapse of the nuclear envelope of HT-1080 cells during nuclear blebbing and rupture in confinement. Arrows indicate time frame of rupture events. Scale bar, 10 µm. (b) Time-dependent change of nuclear longitudinal area of an H2B-mCherry labeled HT-1080 cell during migration in confinement. Arrows indicate incidents of nuclear rupture (top). Time-dependent change of nuclear longitudinal area of an H2B-mCherry labeled HT-1080 cell over the course of one nuclear rupture and repair (bottom). (c) Nuclear volume, measured from confocal Z-stacks of HT-1080 H2B-mCherry nuclear bleb–bearing and nuclear bleb–free cells in confinement (n ≥ 42 cells from four independent experiments). (d) Change in nuclear volume over time of migration through confined channels for representative bleb-bearing and bleb-free cell, as measured from confocal Z-stacks of HT-1080 H2B-mCherry cells. (e) Change in volume over time of migration through confined channels of the nuclear bodies and nuclear blebs, as measured from confocal Z-stacks of HT-1080 H2B-mCherry cells (n = 9 cells from two independent experiments). (f) Percentage of nuclear volume increase relative to the total initial nuclear volume for the total nucleus, nuclear bleb, and nuclear body (n = 8 cells from two independent experiments). (g) Nuclear volume, measured from confocal Z-stacks of HT-1080 H2B-mCherry control and Y27632-treated cells and TRE GFP control and Q63L infected cells in confinement (n ≥ 50 cells from three or more independent experiments). (h) Nuclear volume, measured from confocal Z-stacks of HT-1080 H2B-mCherry control and LMB-treated cells on 2D (n ≥ 52 cells from 3 independent experiments). (i) Nuclear volume, measured from confocal Z-stacks of HT-1080 H2B-mCherry control and LMB-treated cells in confinement (n ≥ 53 cells from three independent experiments). (j) Cell migration speed as a function of nuclear volume as predicted by mathematical modeling based on lubrication theory. (k) HT-1080 speed plotted as a function of nuclear longitudinal area (n = 120 cells from four independent experiments). Solid line represents the best fit, and dotted lines represent the 95% CI. (l) Schematic describing the proposed mechanism for nuclear bleb formation. P denotes pressure. *, P < 0.05 relative to vehicle/scramble control; #, P < 0.05 relative to nuclear bleb bearing; §, P < 0.05 relative to TRE GFP control. Scale bar, 10 µm.

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