Localization of PanK1α within the nucleolus and with the perichromosomal region during mitosis.

<p>HEK293 cells were transfected with expression plasmids encoding mPanK1α(1–185) fused to ZsGreen1 (panels A and B, a, c, d, f, green). (A) Cells were co-transfected with plasmid pAA076 encoding human fibrillarin fused to mCherry (Fibrillarin, magenta, b, c) or plasmid pAA079 encoding human B23 fused to mCherry (B23, magenta, e, f). Fibrillarin designates the dense fibrillar component and B23 designates the granular component of nucleoli. Cell nuclei were visualized by staining with Hoechst 33342 (blue). Cells were visualized using live-cell confocal imaging. Co-localization in merged images are indicated by white pixels containing both magenta and green pseudocolored contributions. Insets in merged images show details at higher magnification. (B) HEK293 cells were co-transfected with with a plasmid encoding human heterochromatin protein 1α fused to mCherry (HP1a, b, c, e, f, magenta). Cells were visualized using live-cell confocal imaging. During interphase, mPanK1α associated with nucleoli and HP1a associated with relaxed heterochromatin, thus designating the nuclear region. In mitotic cells, the nuclear envelope is absent and mPanK1α associated with condensed chromosomes, while HP1a was dissipated throughout the cytoplasm. Dashed lines delimit cell borders (note that mitotic cells are spherical and detached from the substratum). Results are representative of at least 2 independent experiments. Results obtained using hPanK1α were the same. Scale bar, 10 µm.