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Related Concept Videos

Restarting Stalled Replication Forks02:37

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,...
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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...
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In response to DNA damage, cells can pause the cell cycle to assess and repair the breaks. However, the cell must check the DNA at certain critical stages during the cell cycle. If the cell cycle pauses before DNA replication, the cells will contain twice the amount of DNA. On the other hand, if cells arrest after DNA replication but before mitosis, they will contain four times the normal amount of DNA. With a host of specialized proteins at their disposal,cells must use the right protein at...
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The double-stranded structure of DNA has two major advantages. First, it serves as a safe repository of genetic information where one strand serves as the back-up in case the other strand is damaged. Second, the double-helical structure can be wrapped around proteins called histones to form nucleosomes, which can then be tightly wound to form chromosomes. This way, DNA chains up to 2 inches long can be contained within microscopic structures in a cell. A double-stranded break not only damages...
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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...
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Since the discovery of the two BER pathways, there has been a debate about how a cell chooses one pathway over the other and the factors determining this selection. Numerous in vitro experiments have pointed out multiple determinants for the sub-pathway selection. These are:
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Related Experiment Video

Updated: Jul 13, 2025

Real-time Observation of the DNA Strand Exchange Reaction Mediated by Rad51
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RAD51 paralogs synergize with RAD51 to protect reversed forks from cellular nucleases.

Chia-Lun Guh1, Kai-Hang Lei1, Yi-An Chen2

  • 1Institute of Biochemical Sciences, National Taiwan University, Taipei, Taiwan.

Nucleic Acids Research
|October 16, 2023
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RAD51 paralogs, RAD51B-RAD51D-XRCC2 and RAD51C-XRCC3, do not reverse stalled replication forks. However, they synergize with RAD51 to protect these forks from degradation.

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Detection of Homologous Recombination Intermediates via Proximity Ligation and Quantitative PCR in Saccharomyces cerevisiae
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Area of Science:

  • DNA replication
  • DNA repair mechanisms
  • Molecular biology

Background:

  • Replication fork stability is crucial for genome integrity.
  • RAD51 paralogs (RAD51B, RAD51C, RAD51D, XRCC2, XRCC3) are implicated in DNA repair but their roles in fork reversal are unclear.
  • Stalled replication forks are prone to collapse, necessitating protective mechanisms.

Purpose of the Study:

  • To mechanistically dissect the functions of RAD51 paralog complexes BCDX2 (RAD51B-RAD51C-RAD51D-XRCC2) and CX3 (RAD51C-XRCC3) in forming and protecting reversed replication forks.
  • To elucidate the interplay between RAD51 paralogs and the recombinase RAD51 in maintaining fork stability.

Main Methods:

  • Purification of BCDX2 and CX3 complexes.
  • In vitro biochemical assays using reconstituted systems.
  • Analysis of fork reversal and protection activities in the presence of RAD51 paralog complexes and nucleases (MRE11, EXO1).

Main Results:

  • BCDX2 and CX3 complexes do not possess intrinsic fork reversal activity.
  • CX3 complex shows moderate ability to protect reversed forks.
  • BCDX2 complex significantly enhances the protective function of RAD51 against nucleolytic attack on DNA.
  • DNA protection by BCDX2 is dependent on the formation of a functional RAD51 nucleoprotein filament.

Conclusions:

  • RAD51 paralogs do not directly mediate replication fork reversal.
  • RAD51 paralog complexes, particularly BCDX2, play a critical role in protecting stressed replication forks by synergizing with RAD51.
  • This synergy involves the formation of RAD51 nucleoprotein filaments, highlighting a novel mechanism for preventing fork degradation and maintaining genome stability.