Figure 2. Compromising Patronin function... | Rockefeller University Press

Figure 2.

$Compromising Patronin function disrupts ncMT pools and severely impedes nuclear dispersion. (A) Still frames showing MT depletion and detachment from nuclei in Patronin shRNA (Patronin) embryos compared to control. (B) Quantitation highlighting the depletion of perinuclear MT intensities in Patronin embryos compared to control at GBE onset; n = 60 perinuclear regions for each background at 0 min and n = 61 and 59 perinuclear regions for control and Patronin measurements, respectively, at 20 min, k = 3 embryos each. (C) Apical–basal view of MTs and nuclei in control (top) and Patronin (bottom) embryos at 0 and 20 min, showing decreased apical MT networks after Patronin disruption. Arrows point to apical MT. (D) Still frame showing MT enrichment at centrosomal regions in Patronin embryos as compared to control embryos. (E) MT intensities at centrosomes in control and Patronin embryos at 0 and 20 min, indicating enhanced centrosomal MT at both time points; n = 200 and 150 centrosomal regions for control and Patronin, respectively, at 0 min, and n = 200 and 149 centrosomal regions for control and Patronin, respectively, at 20 min, k = 3 embryos for each background. (F) Maximum-intensity projections of nuclei with color-codes based on distance from cell apices in control and Patronin embryos at onset and 20 min into GBE. (G) Comparison of the fraction of nuclei present in the apical 10 μm of cells in control and Patronin embryos indicating inhibited nuclear dispersion on disruption of ncMT; n = 263 and 605 nuclei from k = 3 embryos for control and Patronin, respectively. (H) Fraction of nuclei present in the apical-most 2 μm of the cell (apical exclusion zone) in control and Patronin embryos; n = 455 and 833 nuclei in control and Patronin, respectively at 0 min and n = 483 and 605 nuclei in control and Patronin, respectively, at 20 min, k = 3 embryos for each background. (I) Mean nuclear speed in control and Patronin embryos. (J) Percent of active nuclei as detected by the mean squared displacement (MSD) metric. (I and J)n = 546 and 291 measured nuclei for control and Patronin, respectively, k = 3 embryos each background. Scale bar = 10 μm for (F), 5 μm for (A, C, and D). Fig. 2, A, C, D, and F; Fig. 5 E; Fig. 3, C, D, and F; Fig. 5 H; and Fig. 7 C, control images/plots reproduced for comparison purposes. All scatter plots show the mean ± SD. Statistical significance was calculated using the Mann–Whitney U-test. ns, not significant. ****P < 0.0001.$

Compromising Patronin function disrupts ncMT pools and severely impedes nuclear dispersion. (A) Still frames showing MT depletion and detachment from nuclei in Patronin shRNA (Patronin) embryos compared to control. (B) Quantitation highlighting the depletion of perinuclear MT intensities in Patronin embryos compared to control at GBE onset; n = 60 perinuclear regions for each background at 0 min and n = 61 and 59 perinuclear regions for control and Patronin measurements, respectively, at 20 min, k = 3 embryos each. (C) Apical–basal view of MTs and nuclei in control (top) and Patronin (bottom) embryos at 0 and 20 min, showing decreased apical MT networks after Patronin disruption. Arrows point to apical MT. (D) Still frame showing MT enrichment at centrosomal regions in Patronin embryos as compared to control embryos. (E) MT intensities at centrosomes in control and Patronin embryos at 0 and 20 min, indicating enhanced centrosomal MT at both time points; n = 200 and 150 centrosomal regions for control and Patronin, respectively, at 0 min, and n = 200 and 149 centrosomal regions for control and Patronin, respectively, at 20 min, k = 3 embryos for each background. (F) Maximum-intensity projections of nuclei with color-codes based on distance from cell apices in control and Patronin embryos at onset and 20 min into GBE. (G) Comparison of the fraction of nuclei present in the apical 10 μm of cells in control and Patronin embryos indicating inhibited nuclear dispersion on disruption of ncMT; n = 263 and 605 nuclei from k = 3 embryos for control and Patronin, respectively. (H) Fraction of nuclei present in the apical-most 2 μm of the cell (apical exclusion zone) in control and Patronin embryos; n = 455 and 833 nuclei in control and Patronin, respectively at 0 min and n = 483 and 605 nuclei in control and Patronin, respectively, at 20 min, k = 3 embryos for each background. (I) Mean nuclear speed in control and Patronin embryos. (J) Percent of active nuclei as detected by the mean squared displacement (MSD) metric. (I and J)n = 546 and 291 measured nuclei for control and Patronin, respectively, k = 3 embryos each background. Scale bar = 10 μm for (F), 5 μm for (A, C, and D). Fig. 2, A, C, D, and F; Fig. 5 E; Fig. 3, C, D, and F; Fig. 5 H; and Fig. 7 C, control images/plots reproduced for comparison purposes. All scatter plots show the mean $\pm$ SD. Statistical significance was calculated using the Mann–Whitney U-test. ns, not significant. ****P < 0.0001.

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