Normal cells of a given type possess nuclei of a particular size and shape and maintain a constant nuclear:cell volume ratio. It has been known for many years that abnormalities in nuclear size and morphology are hallmarks of cancer and aging. However, our understanding of the factors that determine nuclear size and shape remains poor. In higher eukaryotes, the nuclear envelope (NE) must expand after it is reformed at the end of mitosis to allow for chromosome decondensation. In yeast, which undergo closed mitosis, the NE must expand to allow chromosome segregation and the formation of two daughter nuclei. The mechanisms by which NE expansion is achieved and controlled are unknown.
Work in our lab has shown that when S. cerevisiae is delayed in mitosis the NE continues to expand despite the fact that chromosome segregation is paused, and a nuclear extension, or “flare”, is formed adjacent to the site of the nucleolus. We hypothesize that flare formation allows maintenance of the nuclear:cell volume ratio in the face of continued NE expansion during an arrest of cell growth. Through a genetic screen, we have discovered that reduced activity of Cdc5, a key cell cycle regulator, causes a no-flare phenotype, where the nucleus remains round during a mitotic arrest. Our previous experiments have shown that Cdc5 is not affecting nuclear morphology through rDNA condensation or any of its other known roles in mitosis. We have also shown that the nuclei of cdc5-nf mutants expand isometrically during a mitotic arrest, indicating that the no-flare phenotype is not a result of reduced NE expansion. In our current work, we found that cdc5-nf cells grow significantly more than WT during a mitotic arrest, indicating that Cdc5 plays a role in inhibiting cell growth. We tested whether the increased cell growth in cdc5-nf is the cause of the no-flare phenotype, reasoning that in WT a flare forms because cell growth is inhibited while NE expansion continues, whereas in cdc5-nf, continued cell growth could allow isometric nuclear expansion without disturbing the nuclear:cell volume ratio. We inhibited cell growth in cdc5-nf using cycloheximide or mutations in the Sec pathway. However, inhibition of cell growth in cdc5-nf did not restore the flared nuclear phenotype. This result demonstrates that Cdc5 plays a role in the inhibition of cell growth and regulation of nuclear morphology through different pathways during a mitotic arrest, and that Cdc5 is required for specifying the site of the nucleolus for nuclear expansion during a mitotic delay.