Fig. 6: NE rupture causes premitotic gene expression.

a HeLa cells were transfected with pEF1α-RFP plasmid. NE was ruptured with a microinjection needle by microinjecting FITC-dextran into the nucleus. Left, schematics of microinjected cells (shaded in green) in upper left panel in b. Regions colored in dark green represent the position of the nucleus with higher FITC-dextran signals. Right, bright-field image of the same cells as in b. An arrow indicates a microinjection needle used for microinjection into the nucleus to achieve NE rupture. Bar, 10 μm. b Time-lapse images of the microinjected cells shown in a. Immediately after microinjection (NE rupture), time-lapse images were acquired every 10 min (2 μm × 5 z-stacks for each time point). Images projected by the maximum intensity projection for z-stacks are shown. The cell in the red-boxed region in the top panel (time 0:00) is shown in the lower panels (time 4:30–9:00). From left to right, FITC-dextran, RFP, and Hoechst, merged, bright-field images. Colors in merged images, FITC-dextran (green), RFP (red), and Hoechst 33342 (blue). The numbers represent time (h:min) from the start of imaging. Red arrow indicates the start of RFP expression. G1, S, G2, M represent the cell-cycle phases. Bars, 10 μm. c Western blotting of HeLa cells treated with siRNA targeting EMD (iemerin) and the control (iLuc), showing that emerin expression in the cells treated with siRNA was reduced to 17% of control levels. d Time-lapse images of the cells treated with iemerin. HeLa cells were treated with siRNA targeting EMD (iemerin). The cells were transfected with pEF1α-RFP plasmid. DNA was stained with Hoechst 33342 (white). Time-lapse images were collected every 10 min (2 μm × 5 z-stacks for each time point) after transfection. Images projected by the maximum intensity projection for z-stacks are shown. Time 0 represents the onset of chromosome segregation. RFP expression (magenta) from transfected plasmid appeared in premitotic stage. Magenta arrow indicates the start of RFP expression. S, G2, and M represent the S, G2, and M phases of the cell cycle. Bar, 10 μm.