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Updated: Oct 13, 2025
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Visualization of DNA Replication in the Vertebrate Model System DT40 using the DNA Fiber Technique
Published on: October 27, 2011
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Visualizing replication fork encounters with DNA interstrand crosslinks
Ryan C James1, Marina A Bellani2, Jing Zhang3
1Department of Molecular Biology and Genetics, Cornell University, Ithaca, NY, United States.
Methods in Enzymology
|November 15, 2021
Summary
Replication forks can restart DNA synthesis past interstrand crosslinks (ICLs). Researchers discovered two distinct replisome versions, one for euchromatin and one for heterochromatin, offering new insights into DNA replication challenges.
Area of Science:
- Molecular Biology
- Cell Biology
- Genetics
Background:
- Replication forks face challenges in euchromatin and heterochromatin during S phase.
- Impediments include DNA structures, transcription complexes, R-loops, DNA-protein complexes, and adducts.
- Replication stress and DNA Damage Response markers are commonly used to study these events.
Purpose of the Study:
- To directly examine the consequences for replisomes encountering DNA replication impediments.
- To visualize replication fork collisions with interstrand crosslinks (ICLs).
Main Methods:
- Developed a novel approach for imaging replication fork collisions with ICLs on DNA fibers.
- Visualized the encounter of replication tracts and an antigen-tagged ICL.
Main Results:
- Observed unexpected restart of DNA synthesis past an intact ICL.
- Identified two distinct replisome populations: one biased toward euchromatin, the other more prominent in heterochromatin.
Conclusions:
- DNA synthesis can restart past intact ICLs, challenging previous assumptions.
- Distinct replisome variants exist, tailored for euchromatin and heterochromatin environments.
- These findings provide direct insights into replisome behavior at replication impediments.
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