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The Replisome

The Replisome

DNA replication is carried out by a large complex of proteins that act in a coordinated matter to achieve high-fidelity DNA replication. Together this complex is known as the DNA replication machinery or the replisome.
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Restarting Stalled Replication Forks

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DNA replication is initiated at sites containing predefined DNA sequences known as origins of replication. DNA is unwound at these sites by the minichromosome maintenance (MCM) helicase and other factors such as Cdc45 and the associated GINS complex.The unwound single strands are protected by replication protein A (RPA) until DNA polymerase starts synthesizing DNA at the 5’ end of the strand in the same direction as the replication fork. To prevent the replication fork from falling apart,...

Homologous Recombination

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The basic reaction of homologous recombination (HR) involves two chromatids that contain DNA sequences sharing a significant stretch of identity. One of these sequences uses a strand from another as a template to synthesize DNA in an enzyme-catalyzed reaction. The final product is a novel amalgamation of the two substrates. To ensure an accurate recombination of sequences, HR is restricted to the S and G2 phases of the cell cycle. At these stages, the DNA has been replicated already and the...

Replication in Eukaryotes

Replication in Eukaryotes

Nucleosome Remodeling

Nucleosome Remodeling

Nucleosomes are the basic units of chromatin compaction. Each nucleosome consists of the DNA bound tightly around a histone core, which makes the DNA inaccessible to DNA binding proteins such as DNA polymerase and RNA polymerase. Hence, the fundamental problem is to ensure access to DNA when appropriate, despite the compact and protective chromatin structure.
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Related Experiment Video

Updated: Oct 15, 2025

Author Spotlight: Investigating the Motion Dynamics of the Eukaryotic Replisome Components at the Single-Molecule Level

Author Spotlight: Investigating the Motion Dynamics of the Eukaryotic Replisome Components at the Single-Molecule Level

Published on: July 26, 2024

A conserved mechanism for regulating replisome disassembly in eukaryotes.

Michael Jenkyn-Bedford¹, Morgan L Jones¹, Yasemin Baris¹

¹MRC Laboratory of Molecular Biology, Cambridge, UK.

|October 26, 2021

Summary

Replisome disassembly, the final step in DNA replication, is regulated by ubiquitylation of the CMG helicase. DNA at replication forks prevents premature CMG ubiquitylation, ensuring genome stability.

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Related Experiment Videos

Last Updated: Oct 15, 2025

Author Spotlight: Investigating the Motion Dynamics of the Eukaryotic Replisome Components at the Single-Molecule Level

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Published on: July 26, 2024

Inducing a Site Specific Replication Blockage in E. coli Using a Fluorescent Repressor Operator System

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Published on: August 21, 2016

Strand-Specific Analysis of Proteins at Replicating DNA Strands by Enrichment and Sequencing of Protein-Associated Nascent DNA Method

Strand-Specific Analysis of Proteins at Replicating DNA Strands by Enrichment and Sequencing of Protein-Associated Nascent DNA Method

Published on: May 2, 2025

Area of Science:

Molecular Biology
Genetics
Biochemistry

Background:

Replisome disassembly concludes eukaryotic DNA replication, initiated by CMG helicase ubiquitylation.
Ubiquitylation is suppressed before replication termination to prevent fork collapse, mediated by replication fork DNA.
The mechanism by which E3 ligases discriminate between elongating and terminated replisomes remains unclear.

Purpose of the Study:

To elucidate the structural basis for the selective ubiquitylation of CMG by SCF^Dia2 and CUL2^LRR1 ligases.
To understand how replication fork DNA suppresses CMG ubiquitylation during elongation.

Main Methods:

High-resolution cryo-electron microscopy (cryo-EM) of budding yeast and human replisome-E3 ligase complexes.
Structural analysis of Dia2 and LRR1 interactions with the CMG helicase.

Main Results:

Structures reveal that LRR domains of Dia2 and LRR1 bind a conserved site on CMG, including MCM3/MCM5.
This LRR-MCM interaction is critical for replisome disassembly.
The excluded DNA strand at replication forks sterically hinders this interaction, preventing CMG ubiquitylation during elongation.

Conclusions:

A conserved mechanism regulates replisome disassembly, involving DNA-mediated suppression of CMG ubiquitylation before termination.
DNA plays a crucial role in maintaining replisome integrity by preventing premature disassembly.