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. 2008 Jul 28;182(2):381–393. doi: 10.1083/jcb.200712066

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© 2008 Ghosh et al.

This article is distributed under the terms of an Attribution–Noncommercial–Share Alike–No Mirror Sites license for the first six months after the publication date (see http://www.jcb.org/misc/terms.shtml). After six months it is available under a Creative Commons License (Attribution–Noncommercial–Share Alike 3.0 Unported license, as described at http://creativecommons.org/licenses/by-nc-sa/3.0/).

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Figure 3. — Gαi3 and GIV localize at the centrosome/MTOC and Gαi3 is necessary for centrosome repositioning. (A) Deconvolved images through the centrosomes demonstrating the localization of Gαi3 and GIV on centrosomes. HeLa and Cos7 cells were permeabilized, fixed, and costained with a centrosome marker (γ-tubulin or pericentrin) and Gαi3 (a–f), GIV (j–l), or overexpressed V5-GIV (g–i). Bar,10 μm. (B) Schematic illustration (bottom) of the centrosome repositioning assay performed on HeLa cells expressing GFP-Centrin1. Cells located at the edge of a wound are scored positive in which the centrosome (GFP-Centrin1, green) and Golgi (Man II, red) are positioned in front of the nucleus (DAPI, blue) within the 120° trident facing the direction of the wound (arrow). The white box indicates the area magnified below. Bar, 10 μm. (C) Bar graph showing the percentage of cells at the wound edge that achieved centrosome repositioning by 8 h after wounding. Gαi3-depleted cells failed to reposition their centrosomes and transfection of Gαi3wt restored this defect. GIV or Gαs-depleted cells were similar to controls (64%). Results (200–400 cells per experiment; n = 3) are shown as mean ± SEM. *, P < 0.001; #, P > 0.001 (compared with scr siRNA cells).