title: Cyclin B1 is rate limiting but not essential for mitotic entry and progression in mammalian somatic cells.
abstract: Cyclin B1 should have some rate limiting function for cell cycle progression. To test this, we measured the effect of siRNA-mediated depletion of cyclin B1 on mitotic entry and timing. We depleted cyclin B1 in HeLa and hTert-RPE1 cells to levels equivalent or below those achieved in the telophase-to-G(1) window. Average cyclin B1/Cdk1 activity was measured in HeLa cells and depleted by approximately 99%. In both cell lines, this caused approximately 20% increase in the G(2) and approximately 20% increase the M traverse time. However, co-depletion of cyclin B1 and B2 induced a profound increase in G(2) cells, a dramatic reduction in mitotic cells, and an increase in a 4C cycling population. We conclude that any residual levels of cyclin B1 were not sufficient to promote stable mitotic entry and transition in absence of normal levels of cyclin B2. Therefore, we conclude that B cyclin is necessary for mitosis but cyclin B1 is not. Nocodazole treated, cyclin B1-depleted HeLa cells arrested but exited that arrest at higher rates than controls, suggesting that the duration of the spindle checkpoint was affected. In B1 depleted cells, population growth was delayed but evidence of cell death was not consistently observed. A strong phenotype of mitotic chromosomal aberration was observed in HeLa cells depleted for either cyclin but not in RPE cells. In B1 or B2 depleted cells, maloriented chromosomes at metaphase were increased 10 fold and one third of affected metaphase cells entered anaphase without congression. Lagging chromosomes at anaphase were dramatically increased. The aggregate evidence from our study and others suggests that the common effect of cyclin B1 depletion is mild cell cycle perturbation. Lack of uniformity in other phenotypes suggest that these are low penetrance effects that are exacerbated or compensated in some systems by other mechanisms.
authors.affiliation: Environmental Health Sciences, Case Western Reserve University, Cleveland, Ohio, USA.
authors.forename: Deena V
authors.initials: DV
authors.lastname: Soni
authors.forename: R Michael
authors.initials: RM
authors.lastname: Sramkoski
authors.forename: Minh
authors.initials: M
authors.lastname: Lam
authors.forename: Tammy
authors.initials: T
authors.lastname: Stefan
authors.forename: James W
authors.initials: JW
authors.lastname: Jacobberger
mesh_terms: D000705:Anaphase / Q000235:genetics; D016203:CDC2 Protein Kinase / Q000235:genetics / Q000378:metabolism; D018797:Cell Cycle Proteins / Q000235:genetics / Q000378:metabolism*; D049109:Cell Proliferation; D002869:Chromosome Aberrations; D020090:Chromosome Segregation / Q000235:genetics; D019926:Cyclin B / Q000235:genetics / Q000378:metabolism*; D056744:Cyclin B1; D056765:Cyclin B2; D015536:Down-Regulation / Q000235:genetics; D018816:Genes, cdc / Q000502:physiology*; D006367:HeLa Cells; D006801:Humans; D008677:Metaphase / Q000235:genetics; D008938:Mitosis / Q000235:genetics*; D015739:Nocodazole / Q000494:pharmacology; D034622:RNA Interference / Q000502:physiology; D008941:Spindle Apparatus / Q000187:drug effects / Q000378:metabolism; D013997:Time Factors
chemical_list: C534699:CCNB1 protein, human; C503758:CCNB2 protein, human; D018797:Cell Cycle Proteins; D019926:Cyclin B; D056744:Cyclin B1; D056765:Cyclin B2; D016203:CDC2 Protein Kinase; D015739:Nocodazole