Figure 5

Microinjection of a morpholino against 14-3-3η causes absence or deformation of meiotic spindles in cells matured in vitro . Oocytes were injected with 0.1 mM 14-3-3η morpholino, held for 24 hours in prophase I arrest, released from the arrest for 13 hours, fixed, permeabilized and immunolabeled for confocal immunofluorescence with an antibody to α-tubulin (green), the antibody to 14-3-3η protein (red), and counterstained with Hoechst 33342 (blue) to visualize the DNA. The panel on the far right is the merged overlay of immunofluorescence images from all three channels. (A-D and E-H) The upper two rows (cells 1 and 2) are images of two representative cells that have clumped DNA, no spindle and no 14-3-3η accumulation. (I-L, M-P, and Q-T) The lower 3 rows (cells 1–3) are representative images of cells that have deformed spindles, disorganized DNA and no accumulation of 14-3-3η at the spindle region. None of these cells injected with the 14-3-3η morpholino formed a first polar body, indicating that the disrupted spindles shown here are MI spindles. Scale bars represent 10 μm.