The nuclear periphery (NP) plays a substantial role in chromatin organization. Heterochromatin at the NP is interspersed with active chromatin surrounding nuclear pore complexes (NPCs); however, details of the peripheral chromatin organization are missing. To discern the distribution of epigenetic marks at the NP of HeLa nuclei, we used structured illumination microscopy combined with a new MATLAB software tool for automatic NP and NPC detection, measurements of fluorescent intensity and statistical analysis of measured data. Our results show that marks for both active and non-active chromatin associate differentially with NPCs. The incidence of heterochromatin marks, such as H3K27me2 and H3K9me2, was significantly lower around NPCs. In contrast, the presence of marks of active chromatin such as H3K4me2 was only decreased very slightly around the NPCs or not at all (H3K9Ac). Interestingly, the histone demethylases LSD1 (also known as KDM1A) and KDM2A were enriched within the NPCs, suggesting that there was a chromatin-modifying mechanism at the NPCs. Inhibition of transcription resulted in a larger drop in the distribution of H1, H3K9me2 and H3K23me2, which implies that transcription has a role in the organization of heterochromatin at the NP.

Department of Biology of the Cell Nucleus Institute of Molecular Genetics CAS v v i Vídeňská 1083 Prague 142 00 Czech Republic

Department of Cybernetics Faculty of Electrical Engineering Czech Technical University Prague Prague 121 35 Czech Republic

Division BIOCEV Institute of Molecular Genetics CAS v v i Průmyslová 595 Vestec Prague 252 50 Czech Republic

Microscopy Centre LM and EM Institute of Molecular Genetics CAS v v i Vídeňská 1083 Prague 142 00 Czech Republic

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