The loss and gain of genomic material is a defining feature of cancer cells. A major mechanism for large scale genomic change is the missegregation of DNA into micronuclei during cell division. Micronuclei can instigate complex genomic rearrangements known as chromothripsis, and they lead to chromosome loss when they are expelled from the genome. Despite the importance of micronuclei in remodelling cancer genomes, little is known about the identity of the DNA within micronuclei.
We have developed unique flow cytometry methods to quantify and purify large numbers of micronuclei. This has enabled us to perform whole genome sequencing of micronuclei to identify that micronuclei are biased towards certain chromosomes; chromosomes 4, 7, 8 and 11. Frequently encapsulated chromosomes are almost 2-fold larger (average size: 1.60 x 108 base pairs) than the poorly encapsulated chromosomes (average size: 8.78 x 107 base pairs), although some large chromosomes (1-3, 5, 6 and X) are not overrepresented. We validated these findings using fluorescent in situ hybridisation to show high frequency of chromosome 11 in micronuclei.
Loss of p53 loss is strongly implicated in chromothripsis which we confirm through analysis of TCGA datasets to show a significant association between p53 -/- and chromothripsis in bladder cancer. As yet there is no direct link between micronuclei, which reintegrate during chromothripsis, and the loss of p53, and indeed these may affect different aspects of chromothripsis. However, we identified that p53-/- cells produce more micronuclei, that these micronuclei are larger, and that these micronuclei also have a higher frequency of centromeres. In addition we show that p53 -/- cells fail to undergo mitotic slippage, possibly leading to greater opportunity to form micronuclei. We are currently sequencing p53-/- micronuclei to identify whether these have a different genomic profile to wildtype micronuclei. These data will shed light on whether p53 loss could influence the genomic profile of DNA that is later reintegrated into the genome during chromothripsis.