Centrosome Hyperamplification and Chromosomal Damage after Exposure to Radiation

Abstract

Objective: In order to elucidate the effects of radiation on centrosome hyperamplification (CH), we examined the centrosome duplication cycle in KK47 bladder cancer cells following irradiation. Methods: KK47 cells were irradiated with various doses of radiation and were examined for CH immunostaining for γ-tubulin. Results: Nearly all control cells contained one or two centrosomes, and mitotic cells displayed typical bipolar spindles. The centrosome replication cycle is well regulated in KK47. Twenty-four hours after 5-Gy irradiation, ∼80% of irradiated cells were arrested in G2 phase, and at 48 h after irradiation, 56.9% of cells contained more than two centrosomes. Laser scanning cytometry performed 48 h after irradiation showed the following two pathways: (1) unequal distribution of chromosomes to daughter cells, or (2) failure to undergo cytokinesis, resulting in polyploidy. With mitotic collection, M-phase cells with CH could be divided into G1 cells with micronuclei and polyploidal cells. Fluorescence in situ hybridization analysis showed clear signs of chromosomal instability (CIN) at 48 h after irradiation. The present study had two major findings: (1) continual duplication of centrosomes occurred in the cell cycle-arrested cells upon irradiation, leading to centrosome amplification; (2) cytokinesis failure was due to aberrant mitotic spindle formation caused by the presence of amplified centrosomes. Abnormal mitosis with amplified centrosomes was detected in the accumulating G2/M population after irradiation, showing that this amplification of centrosomes was not caused by failure to undergo cytokinesis, but rather that abnormal mitosis resulting from amplification of centrosomes leads to cytokinesis block. Conclusion: These results suggest that CH is a critical event leading to CIN following exposure to radiation.

Differential role of repair proteins, BRCA1/NBS1 and Ku70/DNA‐PKcs, in radiation‐induced centrosome overduplication

“…(20,31) We next evaluated the frequencies of cells containing centrosome overduplication in radiation sensitive cells on the basis of cell killing, as we described previously. (32) When the frequency of cells containing overduplicated centrosomes was plotted against cell death, which was derived from Figs 3-5 (1-5 Gy) and from Fig. S1 (8-15 Gy), the curves of all wild-type cells were similar, regardless of the radiation dose rate (Fig.…”

Suppression of Centrosome Amplification after DNA Damage Depends on p27 Accumulation

“…4C). Micronuclei and amplified centrosomes are related to radiationinduced mitotic abnormalities (34,35). Notably, p27-silenced cells with micronuclei were substantially increased (0.4-22.1%) compared with control cells (0.4-1.85%; Fig.…”

A Dominant Role for p53-Dependent Cellular Senescence in Radiosensitization of Human Prostate Cancer Cells

“…These cells may be predisposed to an accumulation of chromosomal abnormalities, such as micronuclei formation following IR. 38 To determine whether the loss of G 2 checkpoint control in DU145 was associated with increased chromosomal instability, a cytokinesis-block micronucleus assay 28 was used to examine the relationships between cell survival, G 2 checkpoint control and chromosome fragmentation in CaP cells. We determined the baseline micronuclei frequencies, in order to assess the extent of preexisting chromosomal damage, and induced (24 h after 6 Gy) frequency of micronuclei.…”