Super-Resolution Imaging of Higher-Order Chromatin Structures at Different Epigenetic States in Single Mammalian Cells

Abstract

Eukaryotic genome is hierarchically packaged by DNA wrapping around histone proteins into chromatin to fit inside the eukaryotic nucleus. Chemical modification to histone proteins influences the higher-order chromatin structure at individual epigenetic states and chromatin environment to regulate gene expression. However, the genome-wide higher-order chromatin structure shaped by different histone modifications in a single mammalian cell nucleus remains poorly characterized. With stochastic optical reconstruction microscopy (STORM), we perform superresolution imaging on a set of histone acetylation (i.e., H3K9ac, H3K27ac, H3ac, H4ac) and methylation marks (i.e., H3K4me1, H3K4me2, H3K4me3, H3K36me3, H3K27me3, H3K9me3) to characterize the genome-wide higher-order chromatin structures at their epigenetic states and their spatial proximity. We found that in the interphase nuclei of mammalian cells, the higher-order chromatin structure can be categorized into three major types of characteristics: histone acetylation marks form spatially segregated nanoclusters, active histone methylation marks form spatially dispersed larger nanodomains and repressive histone methylation marks form highly condensed large clumps that span a wide range of length scales. We also observed that active histone marks mostly coincide with less compact chromatin, and the repressive histone marks mostly highly coincided with densely packed chromatin. The distinct structural features for histone acetylation and methylation marks are preserved even within the highly condensed chromosomes in the mitotic phase. Furthermore, we found that the active and repressive histone marks are generally spatially exclusive from each other, and also spatially distant from the active form of RNA polymerase II (RNAP II). On the other hand, active histone marks exhibit a significantly higher level of spatial colocalization with each other and with active RNAPII. Taken together, we demonstrate that super-resolution imaging reveals the genome-wide higher-order chromatin structure at their epigenetic states and their environment, which establishes the basis for future investigation of their functional significance at normal and diseased states.