title: Centrosome amplification in CHO and DT40 cells by inactivation of cyclin-dependent kinases.
abstract: To study the mechanism of centrosome duplication in cycling cells, we established a novel system of multiple centrosome formation in two types of cells: CHO cells treated with RO3306, a Cyclin-dependent kinase 1 (Cdk1) inhibitor and DT40 cells, in which Cdks were knocked out by chemical genetics. Cdk1-inactivated cells initiated DNA replication and centrosome duplication at the onset of S phase. They became arrested at the end of G2, but the centrosome cycle continued to produce supernumerary centrioles/centrosomes without DNA endoreplication in those cells. Centrosomes were amplified in a highly synchronous and reproducible manner: all of them were located next to the nucleus and spread widely apart from each other with several μm in distance. Double knockout of Cdk1 and Cdk2 caused cell cycle arrest at G1/S and centrosomes were no longer duplicated. However, cells continued to grow and increased their volume over 10-fold during 48 hr of culture. Centrosome components, including γ-tubulin and Cep135, were synthesized and accumulated during the arrest, allowing rapid centrosome multiplication upon recovery from the cell cycle arrest or expression of exogenous Plk4 in G1/S cells. Thus centrosome amplification results from the discoordination of the centrosome cycle from the progression of other cell cycle events, which is controlled by different levels of Cdk activities.
authors.affiliation: Department of Genetics, Cell Biology, and Development, University of Minnesota, Minneapolis, Minnesota 55455, USA.
authors.forename: Nathan
authors.initials: N
authors.lastname: Steere
authors.forename: Michael
authors.initials: M
authors.lastname: Wagner
authors.forename: Stephen
authors.initials: S
authors.lastname: Beishir
authors.forename: Ewan
authors.initials: E
authors.lastname: Smith
authors.forename: Loretta
authors.initials: L
authors.lastname: Breslin
authors.forename: Ciaran G
authors.initials: CG
authors.lastname: Morrison
authors.forename: Helfrid
authors.initials: H
authors.lastname: Hochegger
authors.forename: Ryoko
authors.initials: R
authors.lastname: Kuriyama
mesh_terms: D000818:Animals; D016466:CHO Cells; D002453:Cell Cycle / Q000502:physiology; D002460:Cell Line; D018385:Centrosome / Q000201:enzymology / Q000502:physiology*; D006224:Cricetinae; D003412:Cricetulus; D018844:Cyclin-Dependent Kinases / Q000378:metabolism*; D006801:Humans; D008858:Microscopy, Phase-Contrast
chemical_list: D018844:Cyclin-Dependent Kinases