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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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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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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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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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In eukaryotic cells, DNA replication is highly conserved and tightly regulated. Multiple linear chromosomes must be duplicated with high fidelity before cell division, so there are many proteins that fulfill specialized roles in the replication process. Replication occurs in three phases: initiation, elongation, and termination, and ends with two complete sets of chromosomes in the nucleus.
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For successful DNA replication, the unwinding of double-stranded DNA must be accompanied by stabilization and protection of the separated single strands of the DNA. This crucial task is performed by single-strand DNA-binding (SSB) proteins. They bind to the DNA in a sequence-independent manner, which means that the nitrogenous bases of the DNA need not be present in a specific order for binding of SSB proteins to it. The binding of SSB proteins straightens single-stranded DNA (ssDNA) and makes...
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Sequence and chromatin features guide DNA double-strand break resection initiation.

Robert Gnügge1, Giordano Reginato2, Petr Cejka2

  • 1Department of Microbiology & Immunology, Columbia University Irving Medical Center, New York, NY 10032, USA.

Molecular Cell
|March 14, 2023
PubMed
Summary
This summary is machine-generated.

The Mre11-Rad50-Xrs2 (MRX) complex initiates DNA repair by nicking DNA double-strand breaks (DSBs). Ku70-Ku80 directs these nicks, influenced by DNA sequence and chromatin structure.

Keywords:
CtIPDNA double-strand breakDNA repairMre11Sae2chromatinhomologous recombinationresection

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Area of Science:

  • Molecular Biology
  • Genetics
  • DNA Repair Mechanisms

Background:

  • DNA double-strand breaks (DSBs) are critical DNA lesions that threaten genome integrity.
  • Accurate and efficient repair of DSBs is essential for cell survival.
  • The Mre11-Rad50-Xrs2 (MRX) complex plays a key role in initiating DSB repair via homologous recombination by processing DNA ends.

Purpose of the Study:

  • To elucidate the mechanisms of MRX complex-mediated 5' strand-specific nicking at DSB ends.
  • To investigate the influence of DNA sequence and chromatin context on MRX nicking activity.
  • To understand how MRX-DNA interactions facilitate the initiation of homologous recombination repair.

Main Methods:

  • Deep sequencing-based assay to map MRX nicks at single-nucleotide resolution.
  • Analysis of MRX nicking patterns in the yeast genome near multiple DSBs.
  • Identification of sequence motifs and DNA properties influencing MRX cleavage.

Main Results:

  • The Ku70-Ku80 complex directs MRX nicks to DSB-proximal sites.
  • Repetitive MRX cleavage extends DNA resection tracts, facilitating repair.
  • A specific DNA sequence motif and meltability profile are preferentially nicked by MRX.
  • Nucleosomes and active transcription impede MRX incision activity.

Conclusions:

  • MRX complex activity is precisely regulated by DNA end-binding proteins like Ku70-Ku80.
  • Local DNA sequence features and chromatin accessibility significantly shape MRX function in DSB repair.
  • These findings provide critical insights into the regulation of DNA double-strand break repair initiation.