Disruption of Centrosome Structure, Chromosome Segregation, and Cytokinesis by Misexpression of Human Cdc14A Phosphatase

In budding yeast, the Cdc14p phosphatase activates mitotic exit by dephosphorylation of specific cyclin-dependent kinase (Cdk) substrates and seems to be regulated by sequestration in the nucleolus until its release in mitosis. Herein, we have analyzed the two human homologs of Cdc14p, hCdc14A and hCdc14B. We demonstrate that the human Cdc14A phosphatase is selective for Cdk substrates in vitro and that although the protein abundance and intrinsic phosphatase activity of hCdc14A and B vary modestly during the cell cycle, their localization is cell cycle regulated. hCdc14A dynamically localizes to interphase but not mitotic centrosomes, and hCdc14B localizes to the interphase nucleolus. These distinct patterns of localization suggest that each isoform of human Cdc14 likely regulates separate cell cycle events. In addition, hCdc14A overexpression induces the loss of the pericentriolar markers pericentrin and γ-tubulin from centrosomes. Overproduction of hCdc14A also causes mitotic spindle and chromosome segregation defects, defective karyokinesis, and a failure to complete cytokinesis. Thus, the hCdc14A phosphatase appears to play a role in the regulation of the centrosome cycle, mitosis, and cytokinesis, thereby influencing chromosome partitioning and genomic stability in human cells

ε-Tubulin Is an Essential Component of the Centriole

Centrioles and basal bodies are cylinders composed of nine triplet microtubule blades that play essential roles in the centrosome and in flagellar assembly. Chlamydomonas cells with the bld2-1 mutation fail to assemble doublet and triplet microtubules and have defects in cleavage furrow placement and meiosis. Using positional cloning, we have walked 720 kb and identified a 13.2-kb fragment that contains ε-tubulin and rescues the Bld2 defects. The bld2-1 allele has a premature stop codon and intragenic revertants replace the stop codon with glutamine, glutamate, or lysine. Polyclonal antibodies to ε-tubulin show peripheral labeling of full-length basal bodies and centrioles. Thus, ε-tubulin is encoded by the BLD2 allele and ε-tubulin plays a role in basal body/centriole morphogenesis