Analysis of localization of cell-cycle regulators in Neurospora crassa

Most fungi are multinucleated organisms. In some fungi, they have asynchronous nuclei in the same cytoplasm. We analyzed a cell-cycle regulation mechanism using a model fungus Neurospora crassa, which can make heterokaryon cells. G1/S cyclin CLN-1 and cyclin-dependent kinase CDC-2 were tagged with different fluorescence in different strains and expressed. By forming a heterokaryon strain of these, two different fluorescence-tagged proteins were expressed in the same cytoplasm. CDC-2 was localized in all nuclei, whereas CLN-1 was not detected in most of the nuclei and was dispersed in the cytoplasm with small granular clusters. This indicates that in multinucleated fungi, cell-cycle regulators, similar to other proteins, are shared around the nuclei regardless of different cell-cycle stages. Moreover, each nucleus can select and use a special cell-cycle regulator only when it is necessary. Fungal nuclei may have a novel pickup mechanism of necessary proteins from their cytoplasm at the point of use.