Cep152 interacts with Plk4 and is required for centriole duplication

Cep152, the orthologue of Drosophila Asterless, is a Plk4 target that functions with Plk4 in centriole assembly

Identification of the kinetochore generated "Wait- Anaphase" signal of the mitotic checkpoint

To ensure accurate segregation, the major cell cycle control mechanism in mitosis, the mitotic checkpoint, delays anaphase onset until all chromosomes have properly attached to spindle microtubules. This is achieved through the production of a "wait anaphase" inhibitor(s) that blocks recognition of cyclin B and securin by Cdc20- activated APC/C, an E3 ubiquitin ligase which targets them for destruction. Using physiologically relevant levels of Mad2, Bub3, BubR1, and Cdc20, unattached kinetochores on purified chromosomes are demonstrated to catalyze generation of a soluble Cdc20 inhibitor or inhibition of Cdc20 already bound to APC/C. Antibody inhibition of Mad1 and dimerization deficient Mad2 are used to demonstrate that the chromosome-produced inhibitor requires both recruitment of Mad2 by Mad1 stably bound at unattached kinetochores and dimerization competent Mad2. By acting directly on Mad2, but not BubR1, purified chromosomes promote BubR1 binding to Cdc20 and APC/C, supporting a model in which immobilized Mad1/Mad2 at kinetochores provides a template for initial assembly of Mad2 bound to Cdc20 that is then converted to BubR1-Cdc20 as sequentially produced mitotic checkpoint inhibitor

Identification of the kinetochore generated "Wait- Anaphase" signal of the mitotic checkpoint

To ensure accurate segregation, the major cell cycle control mechanism in mitosis, the mitotic checkpoint, delays anaphase onset until all chromosomes have properly attached to spindle microtubules. This is achieved through the production of a "wait anaphase" inhibitor(s) that blocks recognition of cyclin B and securin by Cdc20- activated APC/C, an E3 ubiquitin ligase which targets them for destruction. Using physiologically relevant levels of Mad2, Bub3, BubR1, and Cdc20, unattached kinetochores on purified chromosomes are demonstrated to catalyze generation of a soluble Cdc20 inhibitor or inhibition of Cdc20 already bound to APC/C. Antibody inhibition of Mad1 and dimerization deficient Mad2 are used to demonstrate that the chromosome-produced inhibitor requires both recruitment of Mad2 by Mad1 stably bound at unattached kinetochores and dimerization competent Mad2. By acting directly on Mad2, but not BubR1, purified chromosomes promote BubR1 binding to Cdc20 and APC/C, supporting a model in which immobilized Mad1/Mad2 at kinetochores provides a template for initial assembly of Mad2 bound to Cdc20 that is then converted to BubR1-Cdc20 as sequentially produced mitotic checkpoint inhibitor

Cep152 interacts with Plk4 and is required for centriole duplication.

Centrioles are microtubule-based structures that organize the centrosome and nucleate cilia. Centrioles duplicate once per cell cycle, and duplication requires Plk4, a member of the Polo-like kinase family; however, the mechanism linking Plk4 activity and centriole formation is unknown. In this study, we show in human and frog cells that Plk4 interacts with the centrosome protein Cep152, the orthologue of Drosophila melanogaster Asterless. The interaction requires the N-terminal 217 residues of Cep152 and the crypto Polo-box of Plk4. Cep152 and Plk4 colocalize at the centriole throughout the cell cycle. Overexpression of Cep152 (1-217) mislocalizes Plk4, but both Cep152 and Plk4 are able to localize to the centriole independently of the other. Depletion of Cep152 prevents both normal centriole duplication and Plk4-induced centriole amplification and results in a failure to localize Sas6 to the centriole, an early step in duplication. Cep152 can be phosphorylated by Plk4 in vitro, suggesting that Cep152 acts with Plk4 to initiate centriole formation

Epidermal development, growth control, and homeostasis in the face of centrosome amplification

As nucleators of the mitotic spindle and primary cilium, centrosomes play crucial roles in equal segregation of DNA content to daughter cells, coordination of growth and differentiation, and transduction of homeostatic cues. Whereas the majority of mammalian cells carry no more than two centrosomes per cell, exceptions to this rule apply in certain specialized tissues and in select disease states, including cancer. Centrosome amplification, or the condition of having more than two centrosomes per cell, has been suggested to contribute to instability of chromosomes, imbalance in asymmetric divisions, and reorganization of tissue architecture; however, the degree to which these conditions are a direct cause of or simply a consequence of human disease is poorly understood. Here we addressed this issue by generating a mouse model inducing centrosome amplification in a naturally proliferative epithelial tissue by elevating Polo-like kinase 4 (Plk4) expression in the skin epidermis. By altering centrosome numbers, we observed multiciliated cells, spindle orientation errors, and chromosome segregation defects within developing epidermis. None of these defects was sufficient to impart a proliferative advantage within the tissue, however. Rather, impaired mitoses led to p53-mediated cell death and contributed to defective growth and stratification. Despite these abnormalities, mice remained viable and healthy, although epidermal cells with centrosome amplification were still appreciable. Moreover, these abnormalities were insufficient to disrupt homeostasis and initiate or enhance tumorigenesis, underscoring the powerful surveillance mechanisms in the skin