Induction of centrosome amplification in p53 siRNA‐treated human fibroblast cells by radiation exposure

Abstract

Centrosome amplification can be detected in the tissues of p53 -/-mice. In contrast, loss of p53 does not induce centrosome amplification in cultured human cells. However, examination of human cancer tissues and cultured cells has revealed a significant correlation between loss or mutational inactivation of p53 and occurrence of centrosome amplification, supporting the notion that p53 mutation alone is insufficient to induce centrosome amplification in human cells, and that additional regulatory mechanisms are involved. It has recently been shown that gamma irradiation of tumor cells induces centrosome amplification. However, the precise mechanism of radiationinduced centrosome amplification is not fully understood. In the present study, CCD32SK diploid normal human fibroblasts were transfected transiently with short interfering RNA (siRNA) specific for human p53 (CCD/p53i). There was a small increase in the frequency of centrosome amplification in CCD/p53i cells (4.0%) without irradiation. In contrast, CCD/p53i cells after 5-Gy irradiation showed a marked increase in abnormal nuclear shapes and pronounced amplification of centrosomes (46.0%). At 12 h after irradiation, irradiated CCD/p53i cells were arrested in G 2 phase. By laser scanning cytometry, abnormal mitosis with amplified centrosomes was observed frequently in the accumulating G 2 /M population at 48 h after irradiation. In the present study, we found that siRNA-mediated silencing of p53 in normal human fibroblasts, together with DNA damage by irradiation, efficiently induced centrosome amplification and nuclear fragmentation, but these phenomena were not observed with either siRNA-mediated silencing of p53 or irradiation alone.

The presence of high‐risk chromosome aberrations in chronic lymphocytic leukaemia does not correlate with centrosome aberrations

“…In addition, also p53 (mapping to 17p13) has been correlated to centrosome regulation. In p53 null mice and human cancer tissues, as well as in cultured cells harbouring loss or mutational inactivation of p53, centrosome amplification could be detected [3]. Finally, CDK-2 (mapping to 12q13) has been shown to have a role in the complex process of centrosome duplication [2].…”

“…Deletions at 17p13 and 11q22-23 are independent prognostic markers, identifying subgroups of patients with rapid disease progression and short survival times in multivariate analysis, whereas 13q14 deletion as unique aberration is associated with favourable outcome [1]. With the exception of the putative pathogenetic 13q14 deletion, all the aberrations found in B-CLL involve loci where genes with a role in the regulation of centrosome duplication have been mapped [2][3][4]. The centrosome is the major microtubule-organizing centre that governs spindle assembly and bipolarity.…”

“…During the cell cycle the centrosome duplicates only once and undergoes characteristic changes [5]. Impairment of the centrosome cycle may lead to chromosome missegregation and aneuploidy [3].…”

Involvement of Centrosome Amplification in Radiation-Induced Mitotic Catastrophe

“…However, this form of mitotic catastrophe requires functional DNA damage checkpoints. 22,39 While p53 mutation alone is insufficient to induce centrosome amplification in human cells, 23,40,41 impaired p53 function contributes to centrosome amplification, 23,36,42 as it is required both to suppress centrosome amplification and to sustain the DNA damage checkpoints that permit centrosome duplication. An explanation of these apparently-contradictory roles may lie in a differential involvement of p53 in G 1 -S and G 2 -M signaling.…”

Roles of cyclins A and E in induction of centrosome amplification in p53-compromised cells

“…Similar to the Aph-mediated early S-phase arrest, the G 2 arrest is believed to allow newly duplicated centrosomes to regain the duplication competency, and reduplicate upon availability of active α-tubulin G0/G1 G0/G1 S/G2/M Cyclins A and E in centrosome amplification K Hanashiro et al CDK2. Indeed, the G 2 arrest induced by exposure to genotoxic insults results in centrosome amplification (Dodson et al, 2004;Kawamura et al, 2006). We thus tested whether it is also the case in cells lacking cyclin E. Cyclins E þ / þ and E À/À MEFs were exposed to DXR for 48 h. As the majority of cells are arrested in G 2 , and cyclin A is known to be expressed at high levels in S and G 2 /M (for a review, see Desdouets et al, 1995), a noticeable increase in cyclin A was detected in both cyclins E þ / þ and E À/À cells (Figure 6b).…”