Deciphering the tumor progression-dependent changes in mitotic bookmarking of triple-negative breast cancer cells

• Applicant: Sven Beyes, Ph.D.
• Subject Area: Cell Biology ; General Genetics and Functional Genome Biology
• Term: from 2020 to 2023
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 446959723

Cancer is considered both a genetic and an epigenetic disease whose outcome is influenced by environmental cues. Recent genomic studies showed that the adaptation capacity of disseminating tumor cells to foreign microenvironments could rely on reversible epigenetic alterations and reprogramming. These alterations often result in a loss of cell cycle control and subsequent increased, uncontrolled proliferation. To maintain and transmit the aggressive cancer phenotype, the cancer cell identity has somehow to be preserved during mitosis, as the daughter cells show the same epigenetic alterations and gene expression patterns as does the mother cell. Recent evidence highlighted that this preservation of cell identity is achieved by mitotic bookmarking. It has been shown that both, transcription factors (TFs) and epigenetic marks can serve as mitotic bookmarks for the propagation of cell identity traits by allowing a rapid restoration of enhancer-promoter loops and gene expression patterns after completion of mitosis. But up to date it is not fully understood which TFs are involved in mitotic bookmarking and whether the pattern of mitotic bookmarking changes during tumor progression. With this project, I aim to define the role of tumor-associated TFs in mitotic bookmarking of cancer cells during tumor progression towards metastasis formation of triple-negative breast cancer (TNBC).Moreover, I intend to determine whether a cancer-associated permissive chromatin state could favor mitotic bookmarking and cancer cell identity propagation through mitosis. In order to achieve this goal, I will work out an improved synchronization protocol for the capture of cells at the onset of mitosis, precisely between the G2/M border and prometaphase. To identify the alterations in mitotic bookmarking in stage-specific cancer cells, I will profile the accessible chromatin regions by employing ATAC-seq on mitotically arrested cells. By performing computational analyses on these datasets, I will retrieve information about DNA footprinting of TF, which might be involved in mitotic bookmarking. The relevance of these finding will be corroborated by profiling genomewide DNA binding of the identified TFs by Cut&Run analysis. Finally, I will functionally validate the relevance of tumor-associated TFs in mitotic bookmarking by employing CRISPR/Cas9-mediated genome editing to delete the identified TFs. This project will help to understand how mitotic bookmarking in TNBC cells allows for the preservation of the cellular identity when challenged during mitosis and which TFs might play a cancer-type specific role.

• DFG Programme: WBP Fellowship
• International Connection: Italy
• Host: Dr. Alessio Zippo