Abstract
The resolution of chromosomes during anaphase is a key step in mitosis. Failure to disjoin chromatids compromises the fidelity of chromosome inheritance and generates aneuploidy and chromosome rearrangements, conditions linked to cancer development. Inactivation of topoisomerase II, condensin, or separase leads to gross chromosome nondisjunction. However, the fate of cells when one or a few chromosomes fail to separate has not been determined. Here, we describe a genetic system to induce mitotic progression in the presence of nondisjunction in yeast chromosome XII right arm (cXIIr), which allows the characterisation of the cellular fate of the progeny. Surprisingly, we find that the execution of karyokinesis and cytokinesis is timely and produces severing of cXIIr on or near the repetitive ribosomal gene array. Consequently, one end of the broken chromatid finishes up in each of the new daughter cells, generating a novel type of one-ended double-strand break. Importantly, both daughter cells enter a new cycle and the damage is not detected until the next G2, when cells arrest in a Rad9-dependent manner. Cytologically, we observed the accumulation of damage foci containing RPA/Rad52 proteins but failed to detect Mre11, indicating that cells attempt to repair both chromosome arms through a MRX-independent recombinational pathway. Finally, we analysed several surviving colonies arising after just one cell cycle with cXIIr nondisjunction. We found that aberrant forms of the chromosome were recovered, especially when RAD52 was deleted. Our results demonstrate that, in yeast cells, the Rad9-DNA damage checkpoint plays an important role responding to compromised genome integrity caused by mitotic nondisjunction.
Highlights
Chromosomes lagging or bridging during anaphase are believed to be one of the main sporadic causes of cytokinesis failure, which leads to tetraploid cells with multicentrosomes, a hallmark of early tumourigenesis [1,2]
As for the ribosomal DNA array (rDNA)-bearing chromosome XII right arm, the telophase block elicited by the cdc14-1 allele prevents sister chromatid resolution, and segregation, of regions that extend from somewhere within the large rDNA locus to the
When we looked at the segregation pattern of the chromosome XII right arm (cXIIr) telomere in these strains, we found that cdc15-2 always segregated it faithfully in the two cell division that took place (94.060.1% [mean 6 standard error of the mean (SEM), n = 3] of foursomes with four nuclei had one the cXIIr telomere is labelled (tetO) in each nucleus) (Figure 2C for a representative micrograph)
Summary
Chromosomes lagging or bridging during anaphase are believed to be one of the main sporadic causes of cytokinesis failure, which leads to tetraploid cells with multicentrosomes, a hallmark of early tumourigenesis [1,2] If these anaphase bridges break apart, chromosomes could enter the so-called breakage-fusionbridge cycle [3,4,5], which has been related to oncogene amplification and intratumour heterogeneity [6,7,8]. Yeast mutants for any of these players show knotted nuclear masses in anaphase with trailing distal chromosome regions which cannot be resolved in otherwise bipolarly attached centromeres [15,16,17,18,19,20] Despite these anaphase problems, all these mutants often perform cytokinesis, leading to a ‘‘cut’’ phenotype characterized by aneuploid daughter cells carrying broken chromosomes [14,17,21]. This has precluded use of those mutants as tools to follow up the short-term consequences in the progeny of anaphase bridges formed by unresolved sister chromatids
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