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Fig. 3. Addition of recombinant Xenopus NEDD1 (XNEDD1) to egg extracts disrupts microtubule structures. (A) Micrographs of microtubule asters formed in egg extracts in the presence of buffer, 300 nM full-length XNEDD1, XNEDD1-N or XNEDD1-C (all expressed and purified as GST fusion proteins; the GST portion was proteolytically removed prior to the experiment), as indicated. Microtubules (red in the overlays) and DNA (blue) are visualized as described for Fig. 2. Scale bar, 10 µm. (B) Micrographs of spindles formed in the egg extracts in the presence of buffer, 300 nM full-length XNEDD1 or XNEDD1-C, as indicated on the left. Microtubules (red in the overlays) and DNA (blue) are visualized as described for Fig. 2. Scale bar, 10 µm. (C) Quantification of aster intensity for the experiment shown in A, expressed as percent of mock-depleted control. Error bars give the standard error (± s.e.). (D) Micrographs of sperm heads incubated in egg extracts with buffer (top panels) or 300 nM XNEDD1-C (bottom panels) and labeled with antibodies against -tubulin (green in the overlays) or acetylated tubulin (to mark the position of the sperm centrioles; red in the overlays), and stained to visualize the DNA (blue in the overlays). Scale bar, 1 µm. (E) Quantification of the amount of -tubulin immunofluorescence associated with sperm heads for the experiment shown in D, expressed as percent of mock-depleted control. Error bars give the standard error (± s.e.).

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