The lateral distribution of LGN and NuMA is regulated by Gαi-GDP and not by aPKC. (a) GDP-bound Gαi-subunits recruit LGN-GFP to the cell cortex. Apical (top) and Z (bottom) views of cells expressing LGN-GFP fusion protein alone or in combination with Gαi1, Gαi1-GDP, or Gαolf-GDP. LGN-GFP recruitment is increased and expanded along the apical–basal axis upon Gαi and Gαi-GDP expression. (b) NuMA is recruited to the cell cortex by LGN. Cells expressing LGN RNAi hairpin or dominant-negative LGN do not show cortical NuMA in metaphase and anaphase. Note that NuMA is still present at the spindle poles (arrows). (c) Two metaphase cells expressing a NuMA RNAi hairpin: NuMA knockdown does not prevent LGN cortical recruitment. (d) Overexpression of a ubiquitous cortical, activated form of aPKC does not inhibit LGN cortical recruitment compared with an inactive mutant version (K281W) of myristylated aPKC. (e) Inhibition of aPKC with a myristylated pseudo-substrate does not block apical exclusion of GFP-LGN. (f) Replacement of the aPKC phosphorylation site (serine 401) by an alanine residue does not block the apical exclusion of GFP-tagged mouse LGN. All images are apical views, except panel e and the bottom rows in panels a and f, which are Z-views showing the apico–basal distribution of LGN. Lightning arrows indicate the electroporated product(s). Bar, 5 µm.