(Downloading may take up to 30 seconds.
If the slide opens in your browser, select File -> Save As to save it.)
Click on image to view larger version.



Fig. 6. NA14 localization to centrosomes in human cells (A) Left panel: Recombinant DIP13 (lane 1) and NP-40-soluble fraction from COS-7 cells transfected with a HA-tagged version of NA14 (lane 2) were analyzed with anti-NA14 antibody (Probe:anti-NA14;1:100). Positions of molecular weight standards are indicated at the right. Middle and right panels: results of immunoprecipitation experiments performed from a solution containing recombinant DIP13 (lanes 3,7) and from extracts of HA-NA14 overexpressing cells, with 10 µl of anti-DIP13 antibody (lanes 5,9) linked to protein A-Sepharose beads. Negative controls were done from the same HA-NA14-overexpressing cell extracts with 10 µl of preimmune serum linked to protein A-Sepharose beads (lanes 4,8). After washing, bead pellets and supernatants (lanes 6,10) were analyzed by immunoblotting using anti-NA14 antibody (central panel, lanes 3-6) or anti-HA antibody (right panel, lanes 7-10). (B) DIP13/NA14 localizes to centrosomes in HeLa cells and both basal bodies and flagella in human spermatozoa. HeLa cells or human spermatozoa (bottom panels) were double-stained with anti-DIP13 antibody (left panels, green) and anti-{gamma}-tubulin, CTR453 or anti-{alpha}-tubulin (central panels, red). Separate green and red images were collected and merged (right panels). Yellowish staining indicates colocalization of both labelings. Arrows indicate centrosomes or basal bodies. Bars, 10 µm. (C) Triton-soluble and insoluble fractions of KE37 cells and a preparation of isolated centrosomes were resolved by SDS-PAGE, blotted and probed with anti-NA14 antibody.