Independent modulation of the kinase and polo-box activities of Cdc5 protein unravels unique roles in the maintenance of genome stability

• Published in: Proceedings of the National Academy of Sciences - PNAS 2011-10, Vol.108 (43), p.17585-17586
• Main Authors: Ratsima, Hery, Ladouceur, Anne-Marie, Pascariu, Mirela, Sauvé, Véronique, Salloum, Zeina, Maddox, Paul S., D'Amours, Damien
• Format: Article
• Language: English
• Subjects: Biological Sciences; Blotting, Western; Cell cycle; Cell Cycle Proteins - genetics; Cell Cycle Proteins - isolation & purification; Cell Cycle Proteins - metabolism; Cell division; Electrophoresis; Flow Cytometry; Genomic Instability - physiology; Image Processing, Computer-Assisted; Kinases; Microscopy, Fluorescence; Mitosis - genetics; Mitosis - physiology; Mutation; Mutation - genetics; Peptides; Phosphorylation; PNAS Plus; PNAS PLUS (AUTHOR SUMMARIES); Protein Interaction Domains and Motifs - genetics; Protein Kinases - genetics; Protein Kinases - isolation & purification; Protein Kinases - metabolism; Protein-Serine-Threonine Kinases; Saccharomyces cerevisiae; Saccharomyces cerevisiae Proteins - genetics; Saccharomyces cerevisiae Proteins - isolation & purification; Saccharomyces cerevisiae Proteins - metabolism; Spindle Apparatus - metabolism; Spindle Apparatus - physiology
• ISSN: 0027-8424; 1091-6490
• DOI: 10.1073/pnas.1106448108

Polo-like kinases (PLKs) are evolutionarily conserved kinases essential for cell cycle regulation. These kinases are characterized by the presence of a C-terminal phosphopeptide-interaction domain, the polo-box domain (PBD). How the functional domains of PLKs work together to promote cell division is not understood. To address this, we performed a genetic screen to identify mutations that independently modulate the kinase and PBD activities of yeast PLK/Cdc5. This screen identified a mutagenic hotspot in the F-helix region of Cdc5 kinase domain that allows one to control kinase activity in vivo. These mutations can be systematically engineered into other major eukaryotic cell cycle kinases to similarly regulate their activity in live cells. Here, using this approach, we show that the kinase activity of Cdc5 can promote the execution of several stages of mitosis independently of PBD activity. In particular, we observe that the activation of Cdc14 and execution of mitotic exit are uniquely sensitive to the modulation of Cdc5 kinase activity. In contrast, PBD-defective mutants are capable of completing mitosis but are unable to maintain spindle pole body integrity. Consistent with this defect, PBD-deficient cells progressively double the size of their genome and ultimately lose genome integrity. Collectively, these results highlight the specific contributions of Cdc5 functional domains to cell division and reveal unexpected mechanisms controlling spindle pole body behavior and genome stability.