Fig. 6. Rac1 promotes mitotic rounding by regulating actomyosin.

(A) Representative images of activated actomyosin (pMLC) in reversed mice. CDs are labeled with DBA (dolichus biflorus agglutinin; green). pMLC is shown with a fire color scheme applied. (B) Quantification of average pMLC intensity with dots representing four mice per group. (C) pMLC (white)– and DNA (cyan)–labeled CD cell monolayers with a metaphase shown in the center. A pMLC fire color conversion is shown. Three repeat experiments. Scale bars, 20 μm (A and C). (D) pMLC intensity in the perimitotic area with dots representing 10 individual measurements. (E) F-actin (white)– and DNA (cyan)–labeled CD cells grown on a Matrigel-coated polydimethylsiloxane (PDMS) substrate with fluorescent nanobeads. Mitotic cell colored in green. Traction force maps are shown (in pascals). Scale bars, 20 μm. (F) Quantification of perimitotic forces. (G) F-actin (white)– and DNA (cyan)–labeled mitotic metaphase CD cells. Bleb, blebbistatin (5 μM). Scale bars, 10 μm. Red box outlines Metaphase F-actin shape shown on the right (scale bar, 5 μm). (H) Circularity quantification of mitotic metaphase F-actin as shown in (G). A minimum of 10 measurements are shown per group. (I) Live confocal mitosis imaging of CD cells in vitro. Bleb, 5 μM. Scale bars, 10 μm. The mitotic cell was colored in green. (J) Relative distribution of mitotic defects in vitro. At least 10 mitoses per group. In vitro experiments are representative of n = 3. (K) In vivo injection protocol of blebbistatin [5 days, 2 mg/kg per day, intraperitoneally (i.p.)]. Created with Biorender.com. (L) Super-resolution confocal imaging of mitotic CD cells during repair (scale bars, 10 μm). The yellow box outlines the insets that are shown in the top row, which depict color-inverted metaphase F-actin (scale bars, 5 μm). (M) Quantification of mitotic metaphase circularity from (L) showing n = 4 mice per group. Bars are means ± SD. *P < 0.05.