Space and time in the universe of the cell nucleus after ionizing radiation attacks: a comparison of cancer and non-cancer cell response

Abstract

Application of ionizing radiation has an increasing impact on bio-medical research, and cancer diagnosis and treatment. Nevertheless, there are a lot of open questions concerning the understanding of radiation DNA damaging mechanisms and repair processes within the light of radio-sensitivity and thus, individualized medical applications. The three-dimensional architecture of genomes on the micro-, meso- and nano-scale acts in combination with epigenetic modifications as an important player of gene regulation and, consequently, fundamental biological processes such as DNA damage response and repair. So far only little is known about the impact of chromatin architecture on DNA double strand break (DSB) repair pathway selection and progression at individual damage sites. How does a cell nucleus manage DSBs and re-organize the chromatin towards functionally intact repair units? Is there a radiosensitivity-related difference in this reaction? We present investigations of spatial and topological parameters of chromatin and repair foci during a time period of repair to glimpse key aspects related to these questions. Nano-probing of radiation-induced chromatin damage sites and the recruited DNA repair proteins in combination with super-resolution Single Molecule Localization Microscopy (SMLM) are powerful methods for geometric and topological analyses of these structures in single cells and single DSB sites and, thus, to study mechanisms of their formation and repair pathway regulation. We used variable tools for such investigations based on image-free high-precision SMLM, nano-scaled molecule distribution analyses, appropriate metrics following Ripley´s distance frequencies and cluster formation analyses, as well as topological quantifications employing persistence homology. Comparing the topology of repair foci by persistence homology suggests general similarities in repair cluster formation, indicating a well-defined non-random, molecule topology at given time points during repair. However, at the same time, the data reveal a specific nano-architecture of DNA damage foci depending on the chromatin domain and cell type. Showing how chromatin architecture around complex damage sites and repair focus nano-architecture may contribute to ongoing repair process, our studies contribute to the molecular understanding of cellular radiation response and its regulation in cancer and non-cancer cells at sub-light microscopic chromatin levels.