Early replication stress leads to abnormal mitosis and genome rearrangement

Abstract

Yeast cell cycle genetics suggests that cells under replication stress generate damage signals that are sufficient to activate checkpoints and restrain mitosis. Yet in mammalian cancer cells, replication stress is not necessarily sufficient to block division. This can lead to abnormal mitotic structures including chromosome bridges, lagging chromosomes, ultrafine anaphase bridges, and micronuclei. This abnormal mitosis is a source for increased mutations including dramatic chromosome rearrangements including chromothripsis, resulting in characteristic genome instability. We have examined the response to different forms of replication stress in the fission yeast, S. pombe, employing visual methods and single‐cell live‐cell analysis.First, we observe distinct patterns of single‐strand binding protein RPA and homologous recombination protein Rad52 depending on the timing and type of stress we induce. This creates a fingerprint of different S phase‐specific stress responses which we can correlate to distinct outcomes.Second, we identify a novel stress‐associated phenotype in a temperature sensitive mutation of the Mcm4 helicase subunit. Despite under‐replicated DNA, the cells fail to arrest during temperature shift, and following release. Biochemical analysis suggests that they evade the checkpoint. Our dynamic pedigrees reveal surprising examples of division under stress including all the mitotic structures described above, including ultrafine anaphase bridges and apparent micronuclei. These abnormal mitoses are associated with genome instability, mutations, and chromosome rearrangement in the surviving cells.Our study shows that a subset of cells within a whole population can continue to divide, despite significant negative effects on genome integrity, and establishes a genetic model to study mechanisms that allow division to occur despite stress.Support or Funding InformationNIH R01 GM081418 &GM111040