Figure 1. CycB drives premature NEB and overrides CHX-induced interphase arrest. (A) Injection of CycB induces premature NEB. GFP-H2Av (cyan) embryos were injected with rhodamine-tubulin (red) and with either GST or CycB at the onset of cycle 13 interphase. The schematic describes the injection and imaging sequence. Insets are enlarged images of the noninjected areas (white boxes) and the CycB-injected areas (blue boxes). The GST and CycB-injected embryos are representative of four injected embryos for each protein. The nuclear cell cycle stages of the noninjected and CycB-injected areas (white outlines) are indicated at the top left and the bottom of the images, respectively. NEB is detected when rhodamine-tubulin invades the nucleoplasm (compare left inset with right inset, second row). Time is given in minutes/seconds. Arrowheads mark the site of injection. Bars, 20 μm. (B) Injection of CycB does not promote premature chromosome condensation before NEB. GFP-H2Av (cyan) embryos were injected with rhodamine-tubulin (red) followed by GST or CycB injection at the beginning of cycle 13 interphase. The top two rows show the state of chromosome condensation at NEB, which was determined by the nucleoplasm being filled with rhodamine-tubulin. The bottom two rows show the state of chromosome condensation at metaphase. Bars, 5 μm. (C) CycB injections overcome the CHX-induced cytoplasmic and nuclear interphase arrest. GFP-H2Av (cyan) embryos were injected with rhodamine-tubulin (red) followed by CHX at mitosis. GST or CycB was injected at the onset of the following interphase. The schematic describes the injection and imaging sequence. Arrowheads mark the site of injection. CycB induced NEB, chromosome condensation, and mitotic spindle formation (white outlines; see Video 1, available). The areas outlined in gray in the third row are enlarged in the bottom row. Time is given in minutes/seconds. Bars, 20 μm. (D) Single embryo Western blot of uninjected and CycB-injected embryos arrested at mitosis of cycle 13 with colchicine. Anti-CycB antibodies were used to detect endogenous and injected CycB. Anti-GFP was used to detect a marker for loading controls. The injected embryo extract reveals an additional 78-kD molecular mass GST-CycB band.
Figure 1.

CycB drives premature NEB and overrides CHX-induced interphase arrest. (A) Injection of CycB induces premature NEB. GFP-H2Av (cyan) embryos were injected with rhodamine-tubulin (red) and with either GST or CycB at the onset of cycle 13 interphase. The schematic describes the injection and imaging sequence. Insets are enlarged images of the noninjected areas (white boxes) and the CycB-injected areas (blue boxes). The GST and CycB-injected embryos are representative of four injected embryos for each protein. The nuclear cell cycle stages of the noninjected and CycB-injected areas (white outlines) are indicated at the top left and the bottom of the images, respectively. NEB is detected when rhodamine-tubulin invades the nucleoplasm (compare left inset with right inset, second row). Time is given in minutes/seconds. Arrowheads mark the site of injection. Bars, 20 μm. (B) Injection of CycB does not promote premature chromosome condensation before NEB. GFP-H2Av (cyan) embryos were injected with rhodamine-tubulin (red) followed by GST or CycB injection at the beginning of cycle 13 interphase. The top two rows show the state of chromosome condensation at NEB, which was determined by the nucleoplasm being filled with rhodamine-tubulin. The bottom two rows show the state of chromosome condensation at metaphase. Bars, 5 μm. (C) CycB injections overcome the CHX-induced cytoplasmic and nuclear interphase arrest. GFP-H2Av (cyan) embryos were injected with rhodamine-tubulin (red) followed by CHX at mitosis. GST or CycB was injected at the onset of the following interphase. The schematic describes the injection and imaging sequence. Arrowheads mark the site of injection. CycB induced NEB, chromosome condensation, and mitotic spindle formation (white outlines; see Video 1). The areas outlined in gray in the third row are enlarged in the bottom row. Time is given in minutes/seconds. Bars, 20 μm. (D) Single embryo Western blot of uninjected and CycB-injected embryos arrested at mitosis of cycle 13 with colchicine. Anti-CycB antibodies were used to detect endogenous and injected CycB. Anti-GFP was used to detect a marker for loading controls. The injected embryo extract reveals an additional 78-kD molecular mass GST-CycB band.

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