The causes of replication stress in response to whole chromosomal aneuploidy

Zuzana Storchova, Kaiserslautern


Cancer is a disease of the genome, with both structural and numerical chromosomal aberrations, so called aneuploidy, occurring concomitantly in most tumors. Despite this co-occurrence, the advances in the characterization of point mutations and structural and numerical changes failed to provide an insight into a possible functional relationship between structural and whole chromosomal instability. Recently, novel evidence suggests that numerical chromosomal aberrations leads to delayed S phase and accumulation of DNA damage due to elevated replication stress, thereby facilitating emergence of structural chromosomal rearrangements. Our results indicate that numerical chromosomal imbalance due to chromosome gain leads to gene expression changes, ultimately affecting the abundance of factors required for replication. In particular, the replicative helicase MCM2-7 that is essential for origin licensing and progression of replication is affected by chromosomal imbalance. Consequently, aneuploid cells suffer from replication abnormalities, increased levels of DNA damage and de novo formation of chromosomal rearrangements. In the SP8 project, we will characterize in detail the changes of replication dynamics in response to aneuploidy, in particular the number of active replication forks, replication fork rate and replication stalling. The consequences of the aneuploidy-induced replication stress will be further evaluated by next generation sequencing (SP-Z) to determine the types of arising chromosomal rearrangements. To identify molecular mechanisms leading to replication defects, we will collaborate with Markus Räschle (SP6) and Petra Beli (SP1) to analyze the proteome and phosphoproteome of the replicating chromatin and its changes in aneuploids. This approach will reveal the limiting factors of replication and their possible abnormal regulation in response to aneuploidy. Finally, we will determine what causes the downregulation of the replicative factors, in particular the MCM2-7 helicase, in aneuploid cells. We hypothesize that a defect in maintenance of protein homeostasis triggered by gene expression changes in aneuploids impairs the abundance of crucial replicative proteins. By means of depletion and overexpression of key protein folding factors, we will identify those that can rescue the replication defect in aneuploids. This approach will be further facilitated by genome and transcriptome analysis of cancer cells to identify genetic changes that enable adaptation to aneuploidy (with Maik Kschischo, SP3). Downregulation of MCM2-7 helicase appears to be a frequent response to stress conditions, e.g. in aging hematopoietic stem cells. Together with Johanna Flach (SP7), we will address the possibility that deficient maintenance of protein homeostasis is the common cause of reduced MCM2-7 levels in both aging hematopoietic stem cells and in aneuploid cells. Our multifaceted approach will improve the understanding of the molecular mechanisms that link numerical chromosomal aberrations to structural changes and reveal how cancer cells adapt to their abnormal genomes.

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