title: Radiation induced chromatin conformation changes analysed by fluorescent localization microscopy, statistical physics, and graph theory.
abstract: It has been well established that the architecture of chromatin in cell nuclei is not random but functionally correlated. Chromatin damage caused by ionizing radiation raises complex repair machineries. This is accompanied by local chromatin rearrangements and structural changes which may for instance improve the accessibility of damaged sites for repair protein complexes. Using stably transfected HeLa cells expressing either green fluorescent protein (GFP) labelled histone H2B or yellow fluorescent protein (YFP) labelled histone H2A, we investigated the positioning of individual histone proteins in cell nuclei by means of high resolution localization microscopy (Spectral Position Determination Microscopy = SPDM). The cells were exposed to ionizing radiation of different doses and aliquots were fixed after different repair times for SPDM imaging. In addition to the repair dependent histone protein pattern, the positioning of antibodies specific for heterochromatin and euchromatin was separately recorded by SPDM. The present paper aims to provide a quantitative description of structural changes of chromatin after irradiation and during repair. It introduces a novel approach to analyse SPDM images by means of statistical physics and graph theory. The method is based on the calculation of the radial distribution functions as well as edge length distributions for graphs defined by a triangulation of the marker positions. The obtained results show that through the cell nucleus the different chromatin re-arrangements as detected by the fluorescent nucleosomal pattern average themselves. In contrast heterochromatic regions alone indicate a relaxation after radiation exposure and re-condensation during repair whereas euchromatin seemed to be unaffected or behave contrarily. SPDM in combination with the analysis techniques applied allows the systematic elucidation of chromatin re-arrangements after irradiation and during repair, if selected sub-regions of nuclei are investigated.
authors.affiliation: Institute for Theoretical Physics, Heidelberg University, Philosophenweg 19, 69120, Heidelberg, Germany.
authors.forename: Yang
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: Institute for Theoretical Physics, Heidelberg University, Philosophenweg 19, 69120, Heidelberg, Germany.
authors.forename: Gabriell
authors.initials: G
authors.lastname: Máté
authors.affiliation: Kirchhoff-Institute for Physics, Heidelberg University, Im Neuenheimer Feld 227, 69120, Heidelberg, Germany.
authors.forename: Patrick
authors.initials: P
authors.lastname: Müller
authors.affiliation: Kirchhoff-Institute for Physics, Heidelberg University, Im Neuenheimer Feld 227, 69120, Heidelberg, Germany.
authors.forename: Sabina
authors.initials: S
authors.lastname: Hillebrandt
authors.affiliation: Kirchhoff-Institute for Physics, Heidelberg University, Im Neuenheimer Feld 227, 69120, Heidelberg, Germany.
authors.forename: Matthias
authors.initials: M
authors.lastname: Krufczik
authors.affiliation: Kirchhoff-Institute for Physics, Heidelberg University, Im Neuenheimer Feld 227, 69120, Heidelberg, Germany.
authors.forename: Margund
authors.initials: M
authors.lastname: Bach
authors.affiliation: Kirchhoff-Institute for Physics, Heidelberg University, Im Neuenheimer Feld 227, 69120, Heidelberg, Germany.
authors.forename: Rainer
authors.initials: R
authors.lastname: Kaufmann
authors.affiliation: Kirchhoff-Institute for Physics, Heidelberg University, Im Neuenheimer Feld 227, 69120, Heidelberg, Germany.
authors.forename: Michael
authors.initials: M
authors.lastname: Hausmann
authors.affiliation: Institute for Theoretical Physics, Heidelberg University, Philosophenweg 19, 69120, Heidelberg, Germany.
authors.forename: Dieter W
authors.initials: DW
authors.lastname: Heermann
mesh_terms: D002843:Chromatin / Q000737:chemistry* / Q000528:radiation effects*; D016000:Cluster Analysis; D022041:Euchromatin; D005720:Gamma Rays; D015894:Genome, Human; D006367:HeLa Cells; D006570:Heterochromatin; D006801:Humans; D008856:Microscopy, Fluorescence / Q000379:methods*; D009690:Nucleic Acid Conformation; D011336:Probability; D013223:Statistics as Topic
chemical_list: D002843:Chromatin; D022041:Euchromatin; D006570:Heterochromatin