Figure 4. VIF nuclear cage impacts nuclear shape and volume. (a) Confocal immunofluorescence images of the VIF network in cells fixed and stained with anti-vimentin (top, green) and nucleus (DAPI, white); and in orthogonal view for the 3D renderings of the nuclei of the same cells (bottom) in vim+/+ and vim−/− mEFs. Images show loss of vimentin changes nuclear morphology. Scale bar is 20 µm. The nuclei of cells lacking vimentin are rounder and less smooth compared with those in vim+/+ mEFs. (b) Vim+/+ and rescued mEFs have more compressed, elongated nuclear shapes compared with the vim−/− mEFs (n = 83–219 per condition from at least n = 2 independent experiments). (c) Vim−/− mEFs have significantly smaller spread areas compared with the vim+/+ or rescued mEFs (n = 62–87 per condition from n = 2 independent experiments). (d) Vim+/+ and vim−/− mEFs are elongated to the same extent as determined by measuring cell aspect ratio (n = 150–200 per condition from n = 4 independent experiments). Data are presented as means ± standard error, as noted in the statistical analysis section. NS, non significant. (e and f) Lack of vimentin decreases nuclear (e; n = 83–219 per condition from at least n = 2 independent experiments) and cell volume (f; n = 62–87 per condition from n = 2 independent experiments). The lower and upper parts of the box represent the 25th and 75th percentiles. The ends of the notch indicate 1.57 times the interquartile range divided by the square root of the number of samples. The whiskers indicate the range of the circular variance up to 1.5 times the interquartile range away from the quartiles.(g) Cell volume linearly correlates with nuclear volume but with different slopes (P < 0.035) for vim+/+ (9.1) and vim−/− mEFs (7.2); the shaded regions correspond to 95% confidence levels based on linear regression analysis. (h) Quantification of nuclear shape by determinations using Fourier amplitudes, u(q) (n = 40–43) following staining for immunofluorescence with anti-nuclear lamin A to to trace nuclear contour in vim+/+ and vim−/− mEFs. The normalized amplitude decreases as q2 with a prefactor based on undulated membrane theory that is inversely proportional to the nuclear stress (n = 25 cells per condition from n = 2 independent experiments). The shaded blue and red regions of h represent ± 1 SEM for vim+/+ and vim−/− cells, respectively. Scale bar is 5 µm.
Figure 4.

VIF nuclear cage impacts nuclear shape and volume . (a) Confocal immunofluorescence images of the VIF network in cells fixed and stained with anti-vimentin (top, green) and nucleus (DAPI, white); and in orthogonal view for the 3D renderings of the nuclei of the same cells (bottom) in vim+/+ and vim−/− mEFs. Images show loss of vimentin changes nuclear morphology. Scale bar is 20 µm. The nuclei of cells lacking vimentin are rounder and less smooth compared with those in vim+/+ mEFs. (b) Vim+/+ and rescued mEFs have more compressed, elongated nuclear shapes compared with the vim−/− mEFs (n = 83–219 per condition from at least n = 2 independent experiments). (c) Vim−/− mEFs have significantly smaller spread areas compared with the vim+/+ or rescued mEFs (n = 62–87 per condition from n = 2 independent experiments). (d) Vim+/+ and vim−/− mEFs are elongated to the same extent as determined by measuring cell aspect ratio (n = 150–200 per condition from n = 4 independent experiments). Data are presented as means ± standard error, as noted in the statistical analysis section. NS, non significant. (e and f) Lack of vimentin decreases nuclear (e; n = 83–219 per condition from at least n = 2 independent experiments) and cell volume (f; n = 62–87 per condition from n = 2 independent experiments). The lower and upper parts of the box represent the 25th and 75th percentiles. The ends of the notch indicate 1.57 times the interquartile range divided by the square root of the number of samples. The whiskers indicate the range of the circular variance up to 1.5 times the interquartile range away from the quartiles.(g) Cell volume linearly correlates with nuclear volume but with different slopes (P < 0.035) for vim+/+ (9.1) and vim−/− mEFs (7.2); the shaded regions correspond to 95% confidence levels based on linear regression analysis. (h) Quantification of nuclear shape by determinations using Fourier amplitudes, u(q) (n = 40–43) following staining for immunofluorescence with anti-nuclear lamin A to to trace nuclear contour in vim+/+ and vim−/− mEFs. The normalized amplitude decreases as q2 with a prefactor based on undulated membrane theory that is inversely proportional to the nuclear stress (n = 25 cells per condition from n = 2 independent experiments). The shaded blue and red regions of h represent ± 1 SEM for vim+/+ and vim−/− cells, respectively. Scale bar is 5 µm.

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