DNA Polymerase Delta Governs Parental Histone Transfer to DNA Replication Lagging Strand
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA1057206 | ENA |
REPOSITORIES: ENA
Dataset's files
| Action | DRS | |||
|---|---|---|---|---|
| SRR27352000_1.fastq.gz | Fastqsanger.gz | |||
| SRR27352000_2.fastq.gz | Fastqsanger.gz | |||
| SRR27352001_1.fastq.gz | Fastqsanger.gz | |||
| SRR27352001_2.fastq.gz | Fastqsanger.gz | |||
| SRR27352002_1.fastq.gz | Fastqsanger.gz |
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Project description:Chromatin replication is intricately intertwined with DNA replication, and the recycling of parental histones is essential for epigenetic inheritance. The transfer of parental histones to both the DNA replication leading and lagging strands involves two distinct pathways: the leading strand utilizes DNA polymerase ε subunits Dpb3/Dpb4, while the lagging strand is facilitated by the MCM helicase subunit Mcm2. However, the process by which Mcm2, moving along the leading strand, facilitates the transfer of parental histones to the lagging strand remains unclear. Our study reveals that the deletion of Pol32, a non-essential subunit of major lagging-strand DNA polymerase δ, results in a predominant transfer of parental histone H3-H4 to the replication leading strand. Biochemical analyses further demonstrate that Pol32 can bind histone H3-H4 both in vivo and in vitro. The interaction of Pol32 with parental histone H3-H4 is disrupted by the mutation of Mcm2's histone H3-H4 binding domain. In conclusion, our findings identify the DNA polymerase delta subunit Pol32 as a novel key histone chaperone downstream of Mcm2, mediating the transfer of parental histones to the lagging strand during DNA replication.
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Project description:How parental histone H3-H4 tetramers, the primary carrier of epigenetic modifications, are transferred to leading and lagging strands of DNA replication forks following DNA replication is an important question that remains not well understood. Here we show that DNA polymerase clamp PCNA and its partner involved in lagging strand DNA synthesis, Pol d, regulate parental histone transfer to lagging strands. Mutations at PCNA as well as at subunits of Pol d that impair the PCNA-Pol d interaction affect parental histone transfer to lagging strands, and this defect unlikely arises from their impacts on DNA synthesis. Moreover, Pol d interacts with H3-H4 in vitro. We suggest that the PCNA-Pol d complex, best known for its role in lagging strand DNA synthesis and DNA repair, couples lagging strand DNA synthesis to the transfer of the parental histones H3-H4 for the inheritance of chromatin structures following DNA replication and possibly DNA repair.
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Project description:How parental histones, the carriers of epigenetic modifications, are deposited onto replicating DNA remains poorly understood. Here, we develop the eSPAN method (enrichment and sequencing of protein associated nascent DNA) in mouse embryonic stem (ES) cells, which detects the distribution of histones with distinct modifications on leading and lagging strands in a relatively small number of cells. We show that DNA polymerase α (Pol α), which synthesizes primers used for synthesis of both leading and lagging strands, binds histone H3-H4 preferentially. The Pol α mutant defective in histone binding in vitro impairs the transfer of parental H3-H4 to lagging strand in both yeast and mouse ES cells. Finally, dysregulation of both coding genes and non-coding endogenous retroviruses (ERVs) is detected in mutant ES cells defective in parental histone transfer. Together, we report an efficient eSPAN method for analysis of DNA replication coupled processes in mouse ES cells and reveal the mechanism of Pol α in parental histone transfer.
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Project description:Chromatin replication is intricately intertwined with the recycling of parental histones to the newly duplicated DNA strands for faithful genetic and epigenetic inheritance. The transfer of parental histones occurs through two distinct pathways: leading strand deposition, mediated by the DNA polymerase ε subunits Dpb3/Dpb4, and lagging strand deposition, facilitated by the MCM helicase subunit Mcm2. However, the mechanism of the facilitation of Mcm2 transferring parental histones to the lagging strand while moving along the leading strand remains unclear. Here, we show that the deletion of Pol32, a nonessential subunit of major lagging-strand DNA polymerase δ, results in a predominant transfer of parental histone H3-H4 to the leading strand during replication. Biochemical analyses further demonstrate that Pol32 can bind histone H3-H4 both in vivo and in vitro. The interaction of Pol32 with parental histone H3-H4 is disrupted through the mutation of the histone H3-H4 binding domain within Mcm2. Our findings identify the DNA polymerase δ subunit Pol32 as a critical histone chaperone downstream of Mcm2, mediating the transfer of parental histones to the lagging strand during DNA replication.
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Project description:Although essential for epigenetic inheritance, the transfer of parental histone (H3-H4)2 tetramers that contain epigenetic modifications to replicating DNA strands is poorly understood. Here, we show that the Mcm2-Ctf4-Pol axis facilitates the transfer of parental (H3-H4)2 tetramers to lagging strands of DNA replication forks. Mutating the H3-H4 binding domain of Mcm2, a subunit of the CMG (Cdc45-MCM-GINS) replicative helicase that translocates along leading strands, impairs the transfer of parental (H3-H4)2 to lagging strands. Similar effects are observed in Ctf4 and Pol-primase mutants that disrupt the connection of the CMG helicase via Ctf4-Pol to lagging strands. Our results support a model whereby parental (H3-H4)2 displaced from nucleosomes through leading-strand DNA replication are transferred to lagging strands for nucleosome assembly via the CMG-Ctf4-Pol complex.
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