Studying Chromatin Epigenetics with Fluorescence Microscopy

Abstract

Epigenetic modifications of histones (methylation, acetylation, phosphorylation, etc.) are of great importance in determining the functional state of chromatin. Changes in epigenome underlay all basic biological processes, such as cell division, differentiation, aging, and cancerous transformation. Post-translational histone modifications are mainly studied by immunoprecipitation with high-throughput sequencing (ChIP-Seq). It enables an accurate profiling of target modifications along the genome, but suffers from the high cost of analysis and the inability to work with living cells. Fluorescence microscopy represents an attractive complementary approach to characterize epigenetics. It can be applied to both live and fixed cells, easily compatible with high-throughput screening, and provide access to rich spatial information down to the single cell level. In this review, we discuss various fluorescent probes for histone modification detection. Various types of live-cell imaging epigenetic sensors suitable for conventional as well as super-resolution fluorescence microscopy are described. We also focus on problems and future perspectives in the development of fluorescent probes for epigenetics.

Visualizing the Nucleome Using the CRISPR–Cas9 System: From in vitro to in vivo

“…At present, new data have been obtained on the visualization of chromatin dynamics. The role of replication stress and the role of functional loss by the key regulators of histone dynamics in the emergence of global epigenetic changes, including the development of precancerous conditions, are being studied [ 157 - 159 ].…”

“…We should emphasize the advantages of imaging using genetically encoded FPs, whose contribution to the studies of molecular interactions in biology and biomedicine has been invaluable [ 167 , 168 ]. A large variety of imaging sensors have been designed based on the FRET pairs of colored FPs [ 159 , 169 ].…”

Fluorescence-based super-resolution-microscopy strategies for chromatin studies

“…Here, we will review the current labelling strategies applicable to SRM. We refer to other reviews for a more extensive discussion on other labelling aspects and a more elaborate list of fluorophores, including small- (Grimm and Lavis 2022 ), photoswitchable- (Chozinski et al 2014 ; Endesfelder et al 2011 ), organic- (Dempsey et al 2011 ), live-cell-compatible- (van de Linde et al 2012 ), STED-specific- (Jeong and Kim 2022 ; Kostiuk et al 2019 ), and epigenetic-state-probing (Stepanov et al 2022 ) fluorophores.…”

“…Chromatin compaction can be assessed by analysing the DNA density directly (Martin et al 2021 ), or by using FRET-FISH probes (Mota et al 2022 ). Often also more indirect methods are applied, using transcription activity (Martin et al 2021 ) or epigenetic histone modifications (Stepanov et al 2022 ) as indicators. Alternatively, an assay that probes transposase-accessible chromatin (ATAC) can be used, as the genome accessibility relates to its compaction (Xie et al 2020 ).…”