Histone lysine methylation patterns in human cell types are arranged in distinct three-dimensional nuclear zones

The impact of histone lysine methylation as an essential epigenetic mechanism for gene regulation has been demonstrated by numerous studies where it was functionally and structurally linked to euchromatin and heterochromatin. Most of these data have been obtained by biochemical and two-dimensional (2D)-microscopic techniques providing little information about the global nuclear arrangement of histone modifications. We investigated the 3D architecture and spatial interrelationships of different histone lysine methylation sites (tri-H3K4, mono-H4K20, mono-H3K9, tri-H3K27, tri-H4K20 and tri-H3K9) in various human cell types. Immunofluorescence and confocal microscopy were used together with a quantitative evaluation of 3D images, to reveal spatial relations of specific methylation sites with either centromeres, nascent RNA or with each other. A close association with centromeres was found only for histone methylation sites previously linked to constitutively repressed chromatin. Differences observed in these sites in relation to the cell cycle emphasize the potential relevance of the dynamic properties of heterochromatin for nuclear functions. Nascent RNA was found associated, though to a different degree, with all histone methylation sites, supporting the increasing evidence that transcription occurs across a wide range of the human genome. Finally we demonstrated by simultaneous visualization of different histone lysine methylation sites that methylation patterns are organized in distinct nuclear zones with little apparent intermingling.