Fig. 2. Importin-ß localization to spindle poles is temporally regulated during mitosis. (A) Progression from G2, in which the NE is still intact (a) through mitosis (b-f) is revealed by DAPI (top). Centrosomes are visualized by -tubulin staining (red); importin ß is depicted in green. Scale bar, 10 µm. (B) Frequency of mitotic cells with pole-associated importin-ß signals. At least 100 mitotic figures per stage were analysed in three experiments. (C) Western blot of importin ß in extracts from asynchronous cultures (asy) in thymidine-arrested cells (time 0), and at the indicated times after thymidine wash-out, with or without inhibition of protein synthesis by cycloheximide (CHX). For control, cyclin B1 levels peaked 5-10 hours after the block release (G₂- to M-phase progression) in a protein-synthesis-dependent manner.