Study of mitotic chromatin supports a model of bookmarking by histone modifications and reveals nucleosome deposition patterns

Abstract

Mitosis encompasses key molecular changes including chromatin condensation, nuclear envelope breakdown, and reduced transcription levels. Immediately after mitosis, the interphase chromatin structure is reestablished and transcription resumes. The reestablishment of the interphase chromatin is probably achieved by "bookmarking," i.e., the retention of at least partial information during mitosis. To gain a deeper understanding of the contribution of histone modifications to the mitotic bookmarking process, we merged proteomics, immunofluorescence, and ChIP-seq approaches. We focused on key histone modifications and employed HeLa-S3 cells as a model system. Generally, in spite of the general hypoacetylation observed during mitosis, we observed a global concordance between the genomic organization of histone modifications in interphase and mitosis, suggesting that the epigenomic landscape may serve as a component of the mitotic bookmarking process. Next, we investigated the nucleosome that enters nucleosome depleted regions (NDRs) during mitosis. We observed that in ∼60% of the NDRs, the entering nucleosome is distinct from the surrounding highly acetylated nucleosomes and appears to have either low levels of acetylation or high levels of phosphorylation in adjacent residues (since adjacent phosphorylation may interfere with the ability to detect acetylation). Inhibition of histone deacetylases (HDACs) by the small molecule TSA reverts this pattern, suggesting that these nucleosomes are specifically deacetylated during mitosis. Altogether, by merging multiple approaches, our study provides evidence to support a model where histone modifications may play a role in mitotic bookmarking and uncovers new insights into the deposition of nucleosomes during mitosis.

CTCF sites display cell cycle dependent dynamics in factor binding and nucleosome positioning

“…Here we find that despite remaining highly accessible, nucleosome-sized ATAC-seq fragments are detected at TSSs in mitosis indicating that nucleosomes are able to occupy these sites, and that the spacing between flanking nucleosomes becomes larger and more variable compared to genome-wide average, similar to what we observed at and around CTCF sites. These observations are consistent with a recent study that found that a large fraction of NDRs at TSSs become filled in by a nucleosome that is marked with H3K4 methylation (Javasky et al 2017). The fact that TSSs remain hyperaccessible suggests either that TSSs become occupied by nucleosomes in only a subset of cells in the population, or that nucleosomes rearrange in all cells, creating linkers between adjacent nucleosomes at TSS that are relatively large and thus more accessible than the genome-wide average.…”

“…In line with previous studies and our CUT&RUN data at and around CTCF sites, we found that H2A.Z and H3K4me3 levels are maintained at TSSs in prometaphase (Supplemental Fig. S13; (Lin et al 2016;Varier et al 2010;Wang and Higgins 2013;Nekrasov et al 2012;Kelly et al 2010;Javasky et al 2017)). Interestingly, but not surprisingly, we again observe loss of depletion of histone signal at TSSs in prometaphase, similar to CTCF sites.…”

“…Many histone marks and variants have been suggested to serve as bookmarks for factor binding in mitosis (reviewed in (Wang and Higgins 2013)). Both H2A.Z and H3K4 methylation states have been observed throughout the cell cycle (Nekrasov et al 2012;Kelly et al 2010;Liu et al 2017;Javasky et al 2017). Using CUT&RUN we observed that H2A.Z binding and H3K4 methylation states are maintained at CTCF sites during mitosis, even though CTCF binding is temporarily lost (Fig.…”

Regulation of transcription reactivation dynamics exiting mitosis

“…As a consequence, most TFs and the RNAPII are evicted from mitotic chromosomes 9 and RNA synthesis is drastically reduced [1]. 10 In spite of this global decrease of gene expression during mitosis, proliferating cells are able to maintain 11 their cell identity and propagate regulatory transcriptional programs from mother to daughter cells [2]. 12 Mitotic bookmarking has been proposed as a potential mechanism that could be involved in the transmission 13 of regulatory information during the cell-cycle [3].…”

“…However, it has been shown for a handful of MFs, known 18 as bookmarking factors (BFs) [6][7][8][9], their ability to interact specifically with at least a fraction of their 19 interphase target sites during mitosis, indicating that chromosomes are not as compacted as previously 20 thought [9]. In fact, chromatin accessibility and nucleosomes landscape during mitosis remain unchanged 21 on bookmarked regions bound by known BFs [11,12]. This ability of BFs to maintain chromatin structure 22 locally could promote a quick transcription reactivation exiting mitosis.…”