Centromere-localized breaks indicate the generation of DNA damage by the mitotic spindle

Abstract

Most carcinomas present some form of chromosome instability in combination with spindle defects. Numerical instability is likely caused by spindle aberrations, but the origin of breaks and translocations remains elusive. To determine whether one mechanism can bring about both types of instability, we studied the relationship between DNA damage and spindle defects. Although lacking apparent repair defects, primary Dido mutant cells formed micronuclei containing damaged DNA. The presence of centromeres showed that micronuclei were caused by spindle defects, and cell cycle markers showed that DNA damage was generated during mitosis. Although the micronuclei themselves persisted, the DNA damage within was repaired during S and G2 phases. DNA breaks in Dido mutant cells regularly colocalized with centromeres, which were occasionally distorted. Comparable defects were found in APC mutant cell lines, an independent system for spindle defects. On the basis of these results, we propose a model for break formation in which spindle defects lead to centromere shearing.centrosome | chromosome instability | double-strand break | H2A.X

Are aneuploidy and chromosome breakage caused by a CINgle mechanism?

“…The recent discovery of centromeric DNA damage in cells with spindle defects has provided evidence for a single mechanism explaining both aneuploidy and breakage. 7 Here we will discuss some recent data pointing to the mitotic spindle as the main, and possibly only, culprit in CIN.…”

“…Spectral karyotyping showed that exactly this instability of chromosome arms occurs together with aneuploidy in Dido mutant cells. 7 Genetic mapping of breakpoints in a wide range of cell lines shows a comparable arm preference, 33 and clinical samples show frequent gains and losses of whole arms. 10,11,[43][44][45] Whereas telomeric end-to-end fusion cannot fully explain this type of instability, a pericentromeric break that leaves the telomere intact agrees with the available data.…”

“…Disruption of the Dido gene causes merotelic attachments, centromere distortion, and lagging anaphase chromosomes. 7 Like Mad2 -/-p53 -/-fibroblasts and nocodazoleblocked cells, Dido mutant cells accumulate DSB in mitosis. 7 Remarkably, the breaks generated by spindle defects localize on or adjacent to centromeres, although overall DSB repair defects can be excluded in the Dido mutant and centromeric DSB are repaired just as efficiently as elsewhere on the chromosome.…”

“…40 Nuclear regions of DNA damage have also been found after induction of mitotic slippage by treatment with a minimally effective concentration of spindle poisons, 41,42 suppression of Cenp-E expression by interfering RNA, 41 or mutations in non-checkpoint genes such as death inducer obliterator (Dido) and adenomatous polyposis coli (Apc). 7 Mutations in genes coding for these accessory spindle proteins produce a mild phenotype compared to mutations in checkpoint genes, show little reduction in cell viability, and allow for a more detailed analysis of the associated breaks.…”

Cancer cells that survive checkpoint adaptation contain micronuclei that harbor damaged DNA

“…Micronuclei form from lagging whole chromosomes 14,15 or chromosome fragments 16 that are not incorporated into the main nucleus following anaphase. 17 This can occur if the centromere is either damaged 18 or missing 19 as is the case for acentric chromosome fragments. In addition, damaged chromosomes may fuse and create a dicentric chromosome 17 that becomes attached to both spindle poles.…”