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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...
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...
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...
S-Cdk Initiates DNA Replication02:38

S-Cdk Initiates DNA Replication

The cell cycle is a series of events leading to DNA duplication followed by the division of cell content to form two daughter cells. The cell cycle progresses in four stages—the cell increases in size (gap 1 or G1-phase), duplicates its DNA (synthesis or S-phase), prepares to divide (gap 2 or G2-phase), and divides (mitosis or M-phase).
Two states at the origin of replication
In eukaryotes, the initiation of replication occurs at many sites on the chromosomes, called the origins of replication.
M-Cdk Drives Transition Into Mitosis02:15

M-Cdk Drives Transition Into Mitosis

Checkpoints throughout the cell cycle serve as safeguards and gatekeepers, allowing the cell cycle to progress in favorable conditions and slow or halt it in problematic ones. This regulation is known as the cell cycle control system.
Cyclin-dependent kinases, or Cdks, work in concert with cyclins to control cell cycle transitions. M-Cdk, a complex of Cdk1 bound to M cyclin, is a well-known example of this coordinated control that drives the transition from the G2 to the M phase.
M cyclin...

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Hybrid Ensemble and Single-molecule Assay to Image the Motion of Fully Reconstituted CMG
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Mcm2 phosphorylation and the response to replicative stress.

Brent E Stead1, Christopher J Brandl, Matthew K Sandre

  • 1Department of Biochemistry, Schulich School of Medicine & Dentistry, University of Western Ontario, London, ON, N6A 5C1, Canada.

BMC Genetics
|May 9, 2012
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Summary

Dbf4-dependent kinase Cdc7 (DDK) phosphorylation of Mcm2 is crucial for the DNA damage response. Loss of these sites impairs cell proliferation and increases mutation rates, highlighting DDK

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Area of Science:

  • Cell cycle regulation
  • DNA replication
  • DNA repair

Background:

  • The minichromosome maintenance (Mcm)2-7 complex functions as the replicative helicase in eukaryotes.
  • Dbf4-dependent kinase Cdc7 (DDK) phosphorylates Mcm2, a key regulator of cell proliferation and response to replicative stress.
  • Previous studies indicated Mcm2 phosphorylation sites are essential for responding to caffeine and methyl methanesulfonate (MMS).

Purpose of the Study:

  • To investigate the role of DDK-mediated phosphorylation of Mcm2 in response to various genotoxic agents and DNA damage.
  • To identify genes that interact synthetically lethally or suppress the phenotypes associated with loss of Mcm2 phosphorylation sites.

Main Methods:

  • Created a Saccharomyces cerevisiae strain with alanine mutations at DDK phosphorylation sites (S164 and S170) in Mcm2 (mcm2AA).
  • Assessed sensitivity of mcm2AA strains to hydroxyurea (HU), 5-fluorouracil (5-FU), and phleomycin.
  • Performed genome-wide screens to identify synthetic lethal interactions and suppressors of mcm2AA phenotypes.
  • Measured spontaneous mutation rates using CAN1 forward mutation assay.

Main Results:

  • The mcm2AA strain exhibited sensitivity to HU and 5-FU, but not phleomycin.
  • Synthetic lethal screens identified genes involved in genome integrity and oxidative stress.
  • The mcm2AA strain showed an increased spontaneous mutation rate, while a phosphomimetic mcm2EE strain showed a decreased rate.
  • Suppressor screens identified genes that decrease DNA damage, enhance homologous recombination, or slow replication forks.

Conclusions:

  • DDK-mediated phosphorylation of Mcm2 is essential for the cellular response to replicative stress and certain types of DNA damage.
  • Phosphorylation of Mcm2 likely modulates Mcm2-7 helicase activity, stabilizing replication forks under stress conditions.
  • This phosphorylation event plays a critical role in maintaining genome stability.