Related Experiment Video

Updated: May 24, 2026

Direct Restart of a Replication Fork Stalled by a Head-On RNA Polymerase
07:27

Direct Restart of a Replication Fork Stalled by a Head-On RNA Polymerase

Published on: April 29, 2010

Topoisomerase I poisoning results in PARP-mediated replication fork reversal

Arnab Ray Chaudhuri1, Yoshitami Hashimoto, Raquel Herrador

  • 1Institute of Molecular Cancer Research, University of Zurich, Zurich, Switzerland.

Nature Structural & Molecular Biology
|March 6, 2012
PubMed

Insights

Topoisomerase I (Top1) poisons stall DNA replication forks, promoting reversal to prevent double-strand breaks (DSBs). This fork reversal, dependent on Poly(ADP-ribose) polymerase, limits DNA damage from replication stress.

Area of Science:

  • Molecular Biology
  • Genetics
  • Biochemistry

Background:

  • Topoisomerase I (Top1) relieves DNA torsional stress during replication and transcription.
  • Top1 inhibitors, like camptothecin, are used in cancer chemotherapy.
  • Inhibitor cytotoxicity is often associated with double-strand break (DSB) formation.

Purpose of the Study:

  • To investigate the role of Top1 poisons in DNA replication fork dynamics.
  • To determine the relationship between Top1 inhibition, replication fork reversal, and DSB formation.
  • To identify cellular mechanisms that prevent chromosome breakage under replication stress.

Main Methods:

  • Utilized yeast, mammalian cell lines, and Xenopus laevis egg extracts.
  • Administered Top1 poisons to induce replication stress.
  • Monitored replication fork progression and reversal.
  • Assessed Poly(ADP-ribose) polymerase activity and single-stranded break repair.

Main Results:

  • Top1 poisons rapidly induce replication-fork slowing and reversal.
  • Replication fork reversal can be uncoupled from DSB formation at sublethal doses.
  • Poly(ADP-ribose) polymerase activity is essential for effective fork reversal.
  • Fork reversal limits the formation of DSBs.

Conclusions:

  • Replication fork reversal is a protective mechanism against chromosome breakage during replication stress.
  • Poly(ADP-ribose) polymerase plays a critical role in fork reversal.
  • Proteins involved in fork reversal and restart modulate the cytotoxicity of Top1-targeting chemotherapeutics.

More Related Videos

Detection of DNA Breaks in Dividing Human Cells by Neutral Comet Assay05:55

Detection of DNA Breaks in Dividing Human Cells by Neutral Comet Assay

Published on: August 23, 2024

Detection of Post-Replicative Gaps Accumulation and Repair in Human Cells Using the DNA Fiber Assay10:32

Detection of Post-Replicative Gaps Accumulation and Repair in Human Cells Using the DNA Fiber Assay

Published on: February 3, 2022

Related Concept Videos

Restarting Stalled Replication Forks02:37

Restarting Stalled Replication Forks

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, a...
Restarting Stalled Replication Forks02:37

Restarting Stalled Replication Forks

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, a...
DNA Topoisomerases02:02

DNA Topoisomerases

Topoisomerases are enzymes that relax overwound DNA molecules during various cell processes, including DNA replication and transcription. These enzymes regulate positive and negative DNA supercoiling without changing the nucleotide sequence. DNA overwinding in a clockwise direction results in positively supercoiled DNA, whereas underwinding in a counterclockwise direction produces negatively supercoiled DNA.
Types and Mechanism of action
Topoisomerases are divided into two main types.  Type I...
The DNA Replication Fork01:02

The DNA Replication Fork

An organism’s genome needs to be duplicated in an efficient and error-free manner for its growth and survival. The replication fork is a Y-shaped active region where two strands of DNA are separated and replicated continuously. The coupling of DNA unzipping and complementary strand synthesis is a characteristic feature of a replication fork.   Organisms with small circular DNA, such as E. coli, often have a single origin of replication; therefore, they have only two replication forks, one in...
The DNA Replication Fork01:02

The DNA Replication Fork

An organism’s genome needs to be duplicated in an efficient and error-free manner for its growth and survival. The replication fork is a Y-shaped active region where two strands of DNA are separated and replicated continuously. The coupling of DNA unzipping and complementary strand synthesis is a characteristic feature of a replication fork.   Organisms with small circular DNA, such as E. coli, often have a single origin of replication; therefore, they have only two replication forks, one in...
Translesion DNA Polymerases02:10

Translesion DNA Polymerases

Translesion (TLS) polymerases rescue stalled DNA polymerases at sites of damaged bases by replacing the replicative polymerase and installing a nucleotide across the damaged site. Doing so, TLS allows additional time for the cell to repair the damage before resuming regular DNA replication.
TLS polymerases are found in all three domains of life - archaea, bacteria, and eukaryotes. Of the different classes of TLS polymerases, members of the Y family are fitted with specialized structures that...