Fig. 4. Nuclear import and localisation of T7-tagged WT1; CT (C-terminus +KTS, amino acids 233-449); CTF1 (+KTS); and EGR1, 18 hours after injecting expression plasmids into the GVs of stage IV oocytes. (A) Western blot. Sets of 20 nuclei and cytoplasms were isolated under oil, and protein extracts equivalent to four nuclei (N) and one cytoplasm (C) were loaded. (B-M) Nuclear spreads were immunostained with anti-T7 tag and counterstained with DAPI. Although CT (B-D) immunostained lampbrush chromosome loops similarly to full-length WT1 (Fig. 3), expression of CTF1 appeared to cause chromosome compaction, loop retraction and more intense labelling of Cajal bodies (arrows in E and G). Expression of EGR1 also resulted in chromosome compaction with immunostaining (H) close to the DNA axes (I). As seen in phase contrast (J), lateral loops were less obvious and Cajal bodies (arrows in J) were not immunostained (H). Inclusion of WT1-derived zinc finger one into EGR1 (EGR+F1), however, mimicked full-length WT1 (–KTS) (K-M). Bar, 10 µm.

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