Figure 6

NEDD1 depletion causes abnormal centrosomes. An aliquot of cells treated with control or NEDD1 siRNA as in Figure 5 were stained for NEDD1 (green) and GT335 (red) at day 5 when the siRNA was removed and again at days 8 and 11. (a–c) At all time points in the control (scrambled) siRNA-treated MEFs, NEDD1 and GT335 staining show normal centrioles (1–2 pairs per cell). (d) 5 days after the first NEDD1 siRNA transfection, the level of NEDD1 is dramatically reduced at the centrosome, and GT335 staining reveals some cells have more than two centrosomal structures. (e and f) At 8 and 11 days after NEDD1 siRNA transfection, there are more cells with more than two centrosomal structures even though NEDD1 is becoming reexpressed. (g) The number of centrosomal structures in control scrambled siRNA-treated and NEDD1 siRNA-treated cells was quantitated. At days 5, 8 and 11, there is a significant increase in the number of cells containing more than two centrosomal structures in the NEDD1 siRNA-treated cells compared with control cells. In addition, there is a significant increase in the number of cells with more than two centrosomes between day 5 and day 11 in the NEDD1 siRNA-treated cells. Approximately 300 cells were counted for each group. Error bars show S.E.M. from three independent experiments. Scale bars=40 μm