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First published online 3 August 2004
doi: 10.1242/jcs.01293
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Research Article |
1 Kirchhoff-Institut für Physik, AG Molekulare Biophysik, Ruprecht-Karls-Universität Heidelberg, Im Neuenheimer Feld 227, 69120 Heidelberg, Germany
2 Molekulare Genetik, Deutsches Krebsforschungszentrum, Im Neuenheimer Feld 280, 69120 Heidelberg, Germany
3 Biomedizinische Strukturanalyse, Deutsches Krebsforschungszentrum, Im Neuenheimer Feld 280, 69120 Heidelberg, Germany
4 Cytometrie, Deutsches Krebsforschungszentrum, Im Neuenheimer Feld 280, 69120 Heidelberg, Germany
5 Intelligente Bioinformatiksysteme, Deutsches Krebsforschungszentrum, Im Neuenheimer Feld 280, 69120 Heidelberg, Germany
6 Genregulation und DNA-Topologie, Deutsches Krebsforschungszentrum, Im Neuenheimer Feld 280, 69120 Heidelberg, Germany
* Author for correspondence (e-mail: karsten.rippe{at}kip.uni-heidelberg.de)
Accepted 26 April 2004
The effect of trichostatin A (TSA)-induced histone acetylation on the interphase chromatin structure was visualized in vivo with a HeLa cell line stably expressing histone H2A, which was fused to enhanced yellow fluorescent protein. The globally increased histone acetylation caused a reversible decondensation of dense chromatin regions and led to a more homogeneous distribution. These structural changes were quantified by image correlation spectroscopy and by spatially resolved scaling analysis. The image analysis revealed that a chromatin reorganization on a length scale from 200 nm to >1 µm was induced consistent with the opening of condensed chromatin domains containing several Mb of DNA. The observed conformation changes could be assigned to the folding of chromatin during G1 phase by characterizing the effect of TSA on cell cycle progression and developing a protocol that allowed the identification of G1 phase cells on microscope coverslips. An analysis by flow cytometry showed that the addition of TSA led to a significant arrest of cells in S phase and induced apoptosis. The concentration dependence of both processes was studied.
Key words: TSA, Histone deacetylase, Image correlation spectroscopy, Fractal dimension, Cell cycle
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