Advanced image-free analysis of the nano-organization of chromatin and other biomolecules by Single Molecule Localization Microscopy (SMLM).

The cell as a system of many components, governed by the laws of physics and chemistry drives molecular functions having an impact on the spatial organization of these systems and vice versa. Since the relationship between structure and function is an almost universal rule not only in biology, appropriate methods are required to parameterize the relationship between the structure and function of biomolecules and their networks, the mechanisms of the processes in which they are involved, and the mechanisms of regulation of these processes. Single molecule localization microscopy (SMLM), which we focus on here, offers a significant advantage for the quantitative parametrization of molecular organization: it provides matrices of coordinates of fluorescently labeled biomolecules that can be directly subjected to advanced mathematical analytical procedures without the need for laborious and sometimes misleading image processing.

Topological Analysis of γH2AX and MRE11 Clusters Detected by Localization Microscopy during X-ray-Induced DNA Double-Strand Break Repair.

DNA double-strand breaks (DSBs), known as the most severe damage in chromatin, were induced in breast cancer cells and normal skin fibroblasts by 2 Gy ionizing photon radiation. In response to DSB induction, phosphorylation of the histone variant H2AX to γH2AX was observed in the form of foci visualized by specific antibodies. By means of super-resolution single-molecule localization microscopy (SMLM), it has been recently shown in a first article about these data that these foci can be separated into clusters of about the same size (diameter ~400 nm).

Elucidation of the Clustered Nano-Architecture of Radiation-Induced DNA Damage Sites and Surrounding Chromatin in Cancer Cells: A Single Molecule Localization Microscopy Approach.

In cancer therapy, the application of (fractionated) harsh radiation treatment is state of the art for many types of tumors. However, ionizing radiation is a "double-edged sword"-it can kill the tumor but can also promote the selection of radioresistant tumor cell clones or even initiate carcinogenesis in the normal irradiated tissue. Individualized radiotherapy would reduce these risks and boost the treatment, but its development requires a deep understanding of DNA damage and repair processes and the corresponding control mechanisms.