Genomic Convergence and Suppression of Centrosome Hyperamplification in Primary p53−/− Cells in Prolonged Culture

Difference in the centrosome duplication regulatory activity among p53 ‘hot spot’ mutants: potential role of Ser 315 phosphorylation-dependent centrosome binding of p53

“…To test this hypothesis, we ®rst examined the frequency of mitotic defect (formation of multipolar spindles instead of normal bipolar spindles) in p537/7(R175H)-1 cells, p537/7(R249S)-1 cells, and p537/7(Vec) cells (control) by immunostaining with anti-g-tubulin polyclonal antibody (centrosomes) and anti-a+b tubulin monoclonal antibodies (mitotic spindles). Consistent with the previous studies (Chiba et al, 2000), *30% of mitotic p537/7(Vec) cells contained abnormal spindles (data not shown). As expected, defective mitoses were detected at a higher frequency in cells expressing R175H (*35%) than those expressing R249S (*15%) (Figure 8a, representative immunostaining of abnormal and normal bipolar mitotic spindles were shown in Figure 8b).…”

“…To compromise the background noise and to examine statistically signi®cant numbers of metaphase spreads at the same time, we allowed +3 deviation in the chromosome number (n) ranging from n531 to n4107 (Figure 8c). As previously shown for p537/7 cells (Chiba et al, 2000), p537/7(Vec) cells showed extensive deviation in the chromosome number per cell (data not shown). Consistent with the degree of centrosome abnormality and the frequency of defective mitosis, extensive chromosome instability was observed in the p537/7(R175H)-1 cells (upper panel) at a level similar to p537/7(Vec) cells.…”

“…Although human cells are most desirable as host cells, currently available p53-null human cells all have undergone extensive culturing. We have previously shown that the extent of centrosome abnormality decreases during prolonged passaging (Chiba et al, 2000). Since presence of hyperampli®ed centrosomes frequently results in mitotic defects and cell death, the`natural' restoration of centrosome numbers in p537/7 cells during a long term culture is likely due to acquisition of additional mutation(s) that complement the dysregulation of centrosome duplication initially imposed by loss of p53.…”

Overexpression of aurora kinase A in mouse mammary epithelium induces genetic instability preceding mammary tumor formation

“…It is also possible that prolonged effect of Aurora-A overexpression in vivo may induce some changes that repress the ability of Aurora-A in inducing centrosome amplification. An example of such an inhibition effect was reported in both p53 À/À embryonic epithelial cells (Chiba et al, 2000) and fibroblast cells (Oikawa et al, 2005). p53 À/À cells exhibited centrosome amplification in early passages, however, after 30 passages, the population with abnormal number of centrosome declined and eventually disappeared after passage 50.…”

Centrosome Amplification and Chromosomal Instability in Feline Lymphoma Cell Lines

“…Recently, it has been shown that the centrosome amplification is reduced in prolonged cell cultures [3]. Therefore, we established a new feline lymphoma cell line (R96) for use in this study.…”