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Related Concept Videos

DNA Damage can Stall the Cell Cycle02:36

DNA Damage can Stall the Cell Cycle

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...
DNA Damage Can Stall the Cell Cycle02:36

DNA Damage Can Stall the Cell Cycle

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...
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...
Homologous Recombination02:31

Homologous Recombination

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...
The Replisome03:01

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.
The synthesis of the leading and lagging strands is a highly coordinated process. To explain this, the “Trombone model” was proposed by Bruce Alberts in 1980. The DNA loop formation starts when a primer is synthesized on the parent lagging strand. The loop grows with the...
The Replisome03:01

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.
The synthesis of the leading and lagging strands is a highly coordinated process. To explain this, the “Trombone model” was proposed by Bruce Alberts in 1980. The DNA loop formation starts when a primer is synthesized on the parent lagging strand. The loop grows with the...

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Related Experiment Video

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Quantification of γH2AX Foci in Response to Ionising Radiation
06:53

Quantification of γH2AX Foci in Response to Ionising Radiation

Published on: April 6, 2010

A PP4-phosphatase complex dephosphorylates gamma-H2AX generated during DNA replication.

Dipanjan Chowdhury1, Xingzhi Xu, Xueyan Zhong

  • 1Immune Disease Institute, Harvard Medical School, Boston, MA 02115, USA. dipanjan_chowdhury@dfci.harvard.edu

Molecular Cell
|July 11, 2008
PubMed
Summary

A novel protein phosphatase, PP4, dephosphorylates gamma-H2AX, a marker of DNA replication breaks. This dephosphorylation is crucial for efficient DNA repair and cell survival when DNA replication is inhibited.

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Last Updated: Jul 3, 2026

Quantification of γH2AX Foci in Response to Ionising Radiation
06:53

Quantification of γH2AX Foci in Response to Ionising Radiation

Published on: April 6, 2010

Evaluation of the Spatial Distribution of γH2AX following Ionizing Radiation
09:28

Evaluation of the Spatial Distribution of γH2AX following Ionizing Radiation

Published on: August 7, 2010

Quantitation of γH2AX Foci in Tissue Samples
08:48

Quantitation of γH2AX Foci in Tissue Samples

Published on: June 28, 2010

Area of Science:

  • Molecular Biology
  • Cell Biology
  • Biochemistry

Background:

  • Histone H2A variant H2AX is phosphorylated to gamma-H2AX at DNA double-strand breaks, recruiting DNA repair factors.
  • Protein phosphatase 2A (PP2A) is known to resolve gamma-H2AX foci after exogenous DNA damage.
  • The specific phosphatases involved in resolving gamma-H2AX during DNA replication remain largely unknown.

Purpose of the Study:

  • To identify and characterize phosphatases involved in dephosphorylating gamma-H2AX during DNA replication.
  • To investigate the role of the identified phosphatase complex in DNA repair and cellular response to replication stress.

Main Methods:

  • Identification of a three-protein PP4 phosphatase complex (PP4C, PP4R2, PP4R3beta) in mammalian cells.
  • In vitro dephosphorylation assays using gamma-H2AX within mononucleosomes.
  • Silencing of the PP4 complex using genetic methods.
  • Assessment of DNA repair efficiency and cell sensitivity to DNA replication inhibitors and radiomimetic drugs.

Main Results:

  • The PP4 complex specifically dephosphorylates ATR-mediated gamma-H2AX generated during DNA replication.
  • PP4 efficiently dephosphorylates gamma-H2AX in vitro and acts directly on the substrate in cells.
  • Silencing PP4 leads to inefficient repair of replication-mediated DNA breaks and hypersensitivity to replication inhibitors.
  • PP4 activity is distinct from its effect on ATR or checkpoint kinase activity.

Conclusions:

  • The PP4 phosphatase complex plays a critical role in eliminating gamma-H2AX foci arising from DNA replication stress.
  • Efficient resolution of gamma-H2AX by PP4 is essential for repairing replication-mediated DNA breaks and maintaining genomic stability.
  • Distinct phosphatases, potentially with overlapping functions, are involved in resolving gamma-H2AX at different types of DNA damage.