The role of TP53 in cellular response to monosomy and trisomy

• Applicant: Professorin Dr. Zuzana Storchova
• Subject Area: Cell Biology ; General Genetics and Functional Genome Biology
• Term: since 2018
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 404082216

Somatic human cells are diploid, and contain 22 pairs of somatic chromosomes and one pair of sex chromosomes. Unbalanced changes in chromosome numbers, so called aneuploidy, have largely detrimental consequences, yet, nearly 90 % of cancer cells carry aneuploid karyotype. The reasons for the defects triggered by aneuploidy, as well as the mechanisms that enable cancer cells to adapt to aberrant karyotypes are poorly understood. Additionally, it remains unclear what the function of aneuploidy is in cancer. We have previously engineered human cells to carry extra chromosomes and determined the consequences of chromosome gains by systematic functional analysis. We have now succeeded to establish human cells lacking a single chromosome (monosomy) and characterized their phenotypes. We found that monosomic cells proliferate slower than their diploid isogenic control, accumulate DNA damage, show impaired translation and ribosome biogenesis, and activate the p53 pathway. These observations raise three main questions: A) What are the molecular causes of these phenotypes? B) What is the upstream trigger of the p53 activation? and, finally, C) How cancer cells with monosomy bypass the detrimental consequences of chromosome loss? In the following project, we will identify the molecular mechanisms underlying the accumulation of DNA damage. We will also validate our hypothesis that haploinsufficiency of ribosome protein coding genes is responsible for the impaired translation and ribosome biogenesis. We will test two not mutually exclusive possibilities that either the DNA damage accumulated due to monosomy contributes to the activation of p53, or that the ribosome biogenesis stress activates the p53 pathway. Additionally, we will determine how human cells adapt to monosomy. To this end, we will perform an in vitro evolution of monosomic cells and by next generation sequencing identify the genetic changes that enabled improved proliferation of monosomic cells. Second, we will perform CRISPR/Cas9-based genome wide screens for genes whose activation or inhibition (CRISPRa and CRISPRi, respectively) affect proliferation of monosomic cells. The screen will be performed in both p53-positive and -negative cell lines. Similar approach will be used also for cells carrying an extra chromosome. Selected candidates that had showed a significant, opposite effect in both CRISPRa and CRISPRi screen will be subsequently functionally validated in aneuploid cells. The results from these experiments will be also compared with the available cancer genome databases. Our project will not only improve our understanding of consequences of chromosome aberrations, but will shed light on how monosomy and trisomy contribute to tumorigenesis and may provide new opportunities for therapies of aneuploid cancers.

• DFG Programme: Research Grants