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The DNA Replication Fork01:02

The DNA Replication Fork

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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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Restarting Stalled Replication Forks02:37

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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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Replication in Eukaryotes02:31

Replication in Eukaryotes

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Overview
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DNA Damage can Stall the Cell Cycle02:37

DNA Damage can Stall the Cell Cycle

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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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DNA Replication02:40

DNA Replication

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DNA replication involves the separation of the two strands of the double helix, with each strand serving as a template from which the new complementary strand is copied.  After replication, each double-stranded DNA includes one parental or “old” strand and one “new” strand. This is known as semiconservative replication. The resulting DNA molecules have the same sequence and are divided equally into the two daughter cells.
Replication in Prokaryotes
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Homologous Recombination02:31

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...
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Related Experiment Video

Updated: Aug 7, 2025

Quantifying Replication Stress in Ovarian Cancer Cells Using Single-Stranded DNA Immunofluorescence
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Quantifying Replication Stress in Ovarian Cancer Cells Using Single-Stranded DNA Immunofluorescence

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The TIMELESS effort for timely DNA replication and protection.

Jinal A Patel1, Hyungjin Kim2,3

  • 1Department of Pharmacological Sciences, State University of New York at Stony Brook, Basic Sciences Tower 8-125, 101 Nicolls Rd, Stony Brook, NY, 11794, USA.

Cellular and Molecular Life Sciences : CMLS
|March 9, 2023
PubMed
Summary

The TIMELESS (TIM) protein is crucial for DNA replication fork stability and preventing genome instability. Its upregulation in cancer suggests TIM as a potential therapeutic target for cancer treatment.

Keywords:
ATR-CHK1 checkpointDNA replicationFork protection complexGenome stabilityReplisome

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Area of Science:

  • Molecular Biology
  • Genetics
  • Cancer Biology

Background:

  • Accurate genome replication is vital for cell survival and tumor prevention.
  • DNA replication forks are susceptible to damage, leading to stalling, collapse, and genome instability.
  • The fork protection complex (FPC), with TIMELESS (TIM) as a key scaffold, maintains replication fork integrity.

Purpose of the Study:

  • To elucidate the multifaceted roles of TIMELESS (TIM) in DNA replication and stalled fork protection.
  • To explore the collaborative functions of TIM with other genome surveillance factors.
  • To investigate the potential of targeting TIM as a cancer therapy.

Main Methods:

  • Review of recent advances in understanding TIMELESS (TIM) function.
  • Analysis of TIM's role in coupling CMG helicase and replicative polymerase activities.
  • Examination of TIM's interaction with other replication machinery proteins.

Main Results:

  • Loss of TIM or FPC impairs fork progression, increases fork stalling and breakage, and defects replication checkpoint activation.
  • TIMELESS (TIM) is essential for protecting both active and stalled replication forks.
  • TIMELESS (TIM) is upregulated in various cancers, indicating a potential therapeutic vulnerability.

Conclusions:

  • TIMELESS (TIM) plays a pivotal role in maintaining DNA replication fork integrity and genome stability.
  • Dysregulation of TIM contributes to tumorigenesis by promoting genome instability.
  • Targeting the upregulated TIMELESS (TIM) in cancer cells offers a promising therapeutic strategy.