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. 2009 Jul 16;4(7):e6255. doi: 10.1371/journal.pone.0006255

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Ling et al.

This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are properly credited.

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(A) Alp14p-GFP does not localize to microtubules in mia1-MutNES4 and mia1Δ cells. Shown are single maximum intensity reconstructions of live cells expressing Alp14p-GFP and Pcp1-mCherry. (B) Mia1p-GFP and Alp14p-TagRFP are spatially separated in mia1-MutNES4 cells. (C) MBP-Mia1p-MutNES4 showed weaker interaction with Alp14p-myc in pull-down assays, as compared to MBP-Mia1p. Alp14p-myc yeast lysates were extracted from mia1Δ cells and detected with anti-myc antibody. MBP-tagged proteins were detected by Coomassie staining. (D) When released from the nucleus using pim1-1 mutation at 36°C, Mia1p-MutNES4 is capable of loading Alp14p-GFP on cytoplasmic microtubules. Localization of Alp14p-GFP in wild type septated cells at 24°C and 36°C is included as a control. Shown are single maximum intensity reconstructions of live cells. (E) Overexpression of Mia1p-MutNES4-mCherry partially restores nuclear accumulation and spindle localization of Alp14p-GFP during mitosis. Shown are single maximum intensity reconstructions of live cells. Scale bars = 5 µm.