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Cdc45 is limiting for replication initiation in humans.

Carsten Köhler1, Dennis Koalick1, Anja Fabricius1

  • 1a Research group Biochemistry, Leibniz Institute for Age Research - Fritz Lipmann Institute , Jena , Germany.

Cell Cycle (Georgetown, Tex.)
|February 27, 2016
PubMed
Summary
This summary is machine-generated.

Overexpressing Cdc45 increases DNA replication origin firing but slows elongation, causing replication stress and apoptosis. This occurs due to RPA exhaustion and subsequent DNA breaks, activating ATM/Chk2 signaling.

Keywords:
ApoptosisCMG helicaseRPAgenome instabilityorigin firingreplication catastrophesingle-strand DNA

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Area of Science:

  • Molecular Biology
  • Cell Biology
  • Genetics

Background:

  • Cdc45 is a crucial component of the CMG helicase, essential for eukaryotic DNA replication.
  • Cdc45's role in regulating replication origin firing is critical for genome stability.
  • Dysregulation of DNA replication can lead to cell cycle arrest and apoptosis.

Purpose of the Study:

  • To investigate the consequences of Cdc45 overexpression on DNA replication.
  • To elucidate the signaling pathways involved in replication stress induced by altered Cdc45 levels.
  • To understand the link between origin firing, fork elongation, and cell fate.

Main Methods:

  • Cell culture and manipulation of Cdc45 expression levels.
  • Analysis of DNA replication dynamics, including origin firing and fork elongation rates.
  • Assessment of DNA damage markers (H2AX phosphorylation) and cell death pathways (apoptosis).
  • Investigation of signaling cascades, including ATR/Chk1 and ATM/Chk2 pathways.

Main Results:

  • Cdc45 overexpression resulted in increased origin firing but significantly reduced fork elongation rates.
  • Cells exhibited replication fork asymmetry, S phase arrest, and apoptosis.
  • Accumulation of single-stranded DNA and a diminished ATR/Chk1 response were observed.
  • A mild ATM/Chk2 activation correlated with H2AX phosphorylation and apoptosis.

Conclusions:

  • Elevated Cdc45 levels lead to replication catastrophe by exhausting RPA and impairing the ATR/Chk1 pathway.
  • Subsequent DNA breaks trigger ATM/Chk2-mediated H2AX phosphorylation, culminating in apoptosis.
  • Cdc45 levels must be tightly regulated to maintain replication fidelity and prevent genomic instability.