DNA Damage Checkpoint Signaling Pathways in Human Cancer

ABSTRACT In Xenopus laevis embryos, the midblastula transition (MBT) at the 12th cell division marks initiation of critical developmental events, including zygotic transcription and the abrupt inclusion of gap phases into the cell cycle. Interestingly, although an ionizing radiation-induced checkpoint response is absent in pre-MBT embryos, introduction of a threshold amount of undamaged plasmid or sperm DNA allows a DNA damage checkpoint response to be activated. We show here that undamaged threshold DNA directly participates in checkpoint signaling, as judged by several dynamic changes, including H2AX phosphorylation, ATM phosphorylation and loading onto chromatin, and Chk1/Chk2 phosphorylation and release from nuclear DNA. These responses on physically separate threshold DNA require γ-H2AX and are triggered by an ATM-dependent soluble signal initiated by damaged DNA. The signal persists in egg extracts even after damaged DNA is removed from the system, indicating that the absence of damaged DNA is not sufficient to end the checkpoint response. The results identify a novel mechanism by which undamaged DNA enhances checkpoint signaling and provide an example of how the transition to cell cycle checkpoint activation during development is accomplished by maternally programmed increases in the DNA-to-cytoplasm ratio.

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The Fission Yeast Rad32 (Mre11)-Rad50-Nbs1 Complex Is Required for the S-Phase DNA Damage Checkpoint

Molecular and Cellular Biology ◽

10.1128/mcb.23.18.6564-6573.2003 ◽

2003 ◽

Vol 23 (18) ◽

pp. 6564-6573 ◽

Cited By ~ 53

Author(s):

Charly Chahwan ◽

Toru M. Nakamura ◽

Sasirekha Sivakumar ◽

Paul Russell ◽

Nicholas Rhind

Keyword(s):

Dna Damage ◽

Fission Yeast ◽

Chromosome Instability ◽

S Phase ◽

Comparative Genomic ◽

Dna Damage Checkpoint ◽

Checkpoint Signaling ◽

Damage Repair ◽

Genomic Approach ◽

Telomere Regulation

ABSTRACT Mre11, Rad50, and Nbs1 form a conserved heterotrimeric complex that is involved in recombination and DNA damage checkpoints. Mutations in this complex disrupt the S-phase DNA damage checkpoint, the checkpoint which slows replication in response to DNA damage, and cause chromosome instability and cancer in humans. However, how these proteins function and specifically where they act in the checkpoint signaling pathway remain crucial questions. We identified fission yeast Nbs1 by using a comparative genomic approach and showed that the genes for human Nbs1 and fission yeast Nbs1 and that for their budding yeast counterpart, Xrs2, are members of an evolutionarily related but rapidly diverging gene family. Fission yeast Nbs1, Rad32 (the homolog of Mre11), and Rad50 are involved in DNA damage repair, telomere regulation, and the S-phase DNA damage checkpoint. However, they are not required for G2 DNA damage checkpoint. Our results suggest that a complex of Rad32, Rad50, and Nbs1 acts specifically in the S-phase branch of the DNA damage checkpoint and is not involved in general DNA damage recognition or signaling.

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The inhibition of checkpoint activation by telomeres does not involve exclusion of dimethylation of histone H4 lysine 20 (H4K20me2)

F1000Research ◽

10.12688/f1000research.15166.1 ◽

2018 ◽

Vol 7 ◽

pp. 1027

Author(s):

Julien Audry ◽

Jinyu Wang ◽

Jessica R. Eisenstatt ◽

Kathleen L. Berkner ◽

Kurt W. Runge

Keyword(s):

Dna Damage ◽

Protein Complexes ◽

Dna Damage Checkpoint ◽

Short Report ◽

Dna Repeats ◽

Histone H4 ◽

Wild Type ◽

Telomeric Dna ◽

Checkpoint Signaling ◽

Eukaryotic Chromosome

DNA double-strand (DSBs) breaks activate the DNA damage checkpoint machinery to pause or halt the cell cycle. Telomeres, the specific DNA-protein complexes at linear eukaryotic chromosome ends, are capped DSBs that do not activate DNA damage checkpoints. This “checkpoint privileged” status of telomeres was previously investigated in the yeast Schizosaccharomyces pombe lacking the major double-stranded telomere DNA binding protein Taz1. Telomeric DNA repeats in cells lacking Taz1 are 10 times longer than normal and contain single-stranded DNA regions. DNA damage checkpoint proteins associate with these damaged telomeres, but the DNA damage checkpoint is not activated. This severing of the DNA damage checkpoint signaling pathway was reported to stem from exclusion of histone H4 lysine 20 dimethylation (H4K20me2) from telomeric nucleosomes in both wild type cells and cells lacking Taz1. However, experiments to identify the mechanism of this exclusion failed, prompting our re-evaluation of H4K20me2 levels at telomeric chromatin. In this short report, we used an extensive series of controls to identify an antibody specific for the H4K20me2 modification and show that the level of this modification is the same at telomeres and internal loci in both wild type cells and those lacking Taz1. Consequently, telomeres must block activation of the DNA Damage Response by another mechanism that remains to be determined.

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