Abstract
Inside the nucleus of an intact cell, DNA is folded around histone proteins into nucleosomes and compacted into a multi-layered three-dimensional chromatin network. The nanometre spacing between nucleosomes positioned throughout this structural framework is known to locally modulate local DNA template access and regulate genome function. However, given that this structural feature occurs on a spatial scale well below the diffraction limit of optical microscopy, real time observation of nucleosome proximity in live cells has proven technically difficult, despite recent advances in live cell super resolution imaging. Thus, in recent work, we have been exploring a powerful alternative solution that is based on fluorescence lifetime and anisotropy imaging microscopy (FLIM and FAIM) of Förster resonance energy transfer (FRET) between fluorescently labelled histones - the core protein of a nucleosome. Here we present recent findings from application of this technology to the study of live cell chromatin network dynamics, which demonstrate local ensemble nucleosome proximity, spatiotemporally oscillates between different spacings that are ‘invisible’ to diffraction limited microscopy.
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