PubMed Health⌕ Search
Home/Archive/PubMed

PubMed · 42711322

In vitro reconstitution of chromatin replication recapitulates symmetric histone recycling.

Fritz Nagae·Shizuko Endo·Yasuto Murayama·Tsuyoshi Terakawa
View original source ↗Export metadata ↓

Abstract

Symmetric histone recycling is vital for maintaining epigenetic inheritance upon eukaryotic DNA replication. Recent genome-wide studies have uncovered key determinants of this process, but how these factors collectively support parental histone transfer remains incompletely understood. Here, we successfully reconstitute histone recycling with 24 purified proteins and analyze the products digested by Micrococcal nuclease with Repli-pore-seq, the newly developed pipeline combining nanopore sequencing and deep-learning-based classification. As a result, we identify histones symmetrically recycled as tetrasomes or hexasomes on nucleosome-favorable sequences. We also observe the discordance of the recycled position between lagging and leading strands on the GC-rich DNA sequences. Moreover, removal of Pol δ, Pol32, Dpb3/4, Ctf4, Csm3/Tof1, or Mrc1 disrupts the balance of histone recycling between the two daughter strands, whereas removal of Ctf4, Csm3/Tof1, or Mrc1 additionally alters the positions at which histones were recycled. Furthermore, addition of the lagging-strand maturation factors Fen1 and Cdc9 enhances histone recycling to the lagging strand. These findings provide critical insights into the molecular players and mechanisms underlying symmetric histone recycling.

Explore related subjects

HistonesDNA ReplicationSaccharomyces cerevisiae ProteinsChromatinSaccharomyces cerevisiaeNucleosomesJournal Article

Keep this discovery

Explore connections, maps & timelines

BibTeXRIS

Fritz Nagae, Shizuko Endo, Yasuto Murayama, Tsuyoshi Terakawa. 2026-08-11. In vitro reconstitution of chromatin replication recapitulates symmetric histone recycling.. https://doi.org/10.1038/s41467-026-76520-5

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related citations

PubMed2026

ATX1-COMPASS-like complex participates in the bud dormancy release of tree peony by regulating H3K4me3 modification.

Bud dormancy release in woody plants is crucial for survival, regrowth, flowering, and fruiting. Tree peony (Paeonia suffruticosa), an important ornamental and economic plant, undergoes bud endodormancy in winter, and sufficient chilling duration and exogenous gibberellins (GAs) can effectively break the dormancy. However, the epigenetic regulation mechanism remains poorly understood. Here, immunoblotting revealed that H3K4me3, but not H3K4me1 or H3K4me2, was associated with chilling- and GA3-induced dormancy release. Chromatin immunoprecipitation sequencing (ChIP-seq) combined with RNA-seq results revealed that H3K4me3 enriched near transcription start sites (TSS). H3K4me3 enrichment genes (HEGs) and differentially expressed genes (DEGs) were commonly enriched in KEGG pathways, such as plant hormone signal transduction and MAPK signaling. The expression patterns of these marker genes, such as EARLY BUD-BREAK 3 (PsEBB3), CYCLIND3.1 (PsCYCD3.1), CYCLIND3.3 (PsCYCD3.3), and β-1,3-glucanase 6 (PsBG6), were correlated with their H3K4me3 enrichment and were validated by chromatin immunoprecipitation-quantitative PCR (ChIP-qPCR). Four COMPASS-like component homologs and one histone methyltransferase were screened; among them, PsWDR5a, PsRBL, PsASH2R, and PsATX1 were upregulated by prolonged chilling and GA3 treatments. Yeast two-hybrid (Y2H), yeast three-hybrid (Y3H), luciferase complementation (LCA), and co-immunoprecipitation (Co-IP) analyses revealed that PsRBL interacts with PsWDR5a and PsASH2R as a bridge. PsATX1 was confirmed as an H3K4me3 methyltransferase and interacted with PsWDR5a and PsRBL to form the PsATX1-COMPASS-like complex using Y2H, LCA, and Co-IP assays. Functional analyses showed that PsWDR5a, PsRBL, PsASH2R, and PsATX1 significantly promoted budburst by elevating genomic H3K4me3 levels. Our findings provide insights into the epigenetic regulation of dormancy transitions in woody perennials.

Histones↗
PubMed2026

MacroH2A2-Enriched Domains Are Largely Stable Across the Cell Cycle but Focally Displaced at Mitotic Regulatory Elements.

The macroH2A variants mH2A1 and mH2A2 are structurally similar but not identical. Our previous study demonstrated that mH2A1 is reloaded during cell-cycle progression, but whether mH2A2 follows similar dynamics has remained unclear. Here, we used native ChIP-seq in synchronized Huh-7 cells to profile both variants at G1/S and G2/M. Although mH2A1- and mH2A2-enriched domains overlapped extensively, mH2A2 domains were largely stable across the cell cycle, in sharp contrast to the dynamic reloading of mH2A1. Only a small subset of mH2A2 domains showed phase-specific deposition or displacement. Among these, G1/S-unique mH2A2 domains were preferentially located in the active A compartment and coincided with reduced chromatin accessibility at binding sites for cell-cycle regulators. These G1/S-unique domains co-localize with genes involved in mitotic progression within the same A compartment, suggesting potential regulatory roles in both chromatin organization and transcriptional regulation. These findings refine the classical view of macroH2A variants as static repressive marks: mH2A2 is not entirely static, but its cell-cycle dynamics are far more restricted than those of its paralog mH2A1, occurring only at a small subset of genomic loci, with a regulatory logic distinct from that of mH2A1.

Histones↗
PubMed2026

Improved spike-in normalization clarifies the relationship between active histone modifications and transcription.

Spike-in normalization enables quantitative analysis of chromatin immunoprecipitation sequencing (ChIP-seq) signal. Here we introduce a robust dual spike-in normalization approach for ChIP-seq (ChIP-wrangler), optimize parameters and verify its accuracy in quantifying changes in ChIP-seq signal and detecting technical artifacts. We use ChIP-wrangler to revisit recent claims that active histone marks depend on transcription. We show that acute depletion of RNA polymerase II (RNAPII) has a modest impact on H3K27ac levels, with only 6% of peaks significantly changing after RNAPII depletion, indicating that histone acetylation maintenance is not entirely dependent on ongoing transcription. Promoters and enhancers are differentially affected, with 82% of decreasing acetylation peaks located at promoter-distal elements with enhancer-related motifs. ChIP-wrangler provides increased rigor and 'guardrails' for successful spike-in normalization and, as applied here, refines the understanding of crosstalk between RNAPII activity and transcription-associated histone marks.

Histones↗