Regensburg 2013 – scientific programme

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BP: Fachverband Biologische Physik

BP 15: DNA/RNA and related enzymes

BP 15.5: Talk

Tuesday, March 12, 2013, 13:15–13:30, H43

Identifying chromatin structure during DNA repair by super-resolution microscopy — •Judith Seel and Günther Dollinger — Universität der Bundeswehr München, Neubiberg, Germany

High LET irradiation of living cells using heavy ions generates a high amount of DNA DSB in close vicinity to each other. Various repair proteins and damage markers cluster to the damage sites, such as gamma-H2AX and 53BP1, forming so-called ionizing radiation induced foci of a gross size of about 1um. While structures of this size can be easily resolved using a conventional fluorescence microscope, its substructures cannot be resolved due to the diffraction limit of about 250nm in conventional fluorescence microscopy. For analyzing foci fine-structures systematically, Stimulation Emission Depletion Microscopy (STED) is utilized, which provides a lateral resolution of about 60nm fwhm.

With these improvements the microscopic images clearly indicate a fine-structure when 53BP1 is stained with two colors and the quantitative analysis proves its existence at a scale of a few hundert nm. Using the same methods with images where one color labels 53BP1 and the other gamma-H2AX, it can be shown that there is no total correlation between these two markers on a small scale.

Using these experimental and analytical methods it is possible to determine the way of clustering of one single DNA damage marker to DSB to clarify the structure of a DSB and the structure of chromatin architecture. Secondly, the comparison of two damage markers gives a deeper understanding of the interaction of repair markers and at the end maybe the possibility to decode the structure of DNA repair.