Chk1 is required to maintain Claspin stability

Chini, Claudia Christiano Silva; Wood, J.D.; Chen, Junjie

doi:10.1038/sj.onc.1209447

Cited by 55 publications

(66 citation statements)

References 23 publications

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“…These proteins may regulate each other's expression, and indeed, Chk1 has been shown to regulate the stability of Claspin. 32 However, the abundance of these proteins is higher during S phase in normal human fibroblasts and other cell lines. 4,11,12,33,34 Equivalent amounts of protein were loaded per well for SDS-PAGE, but the S-phase fraction in NHF1-hTERT cultures depleted of Chk1, Timeless, Tipin or Claspin was reduced (data not shown), which could account for the lower levels of these proteins when any one of them was targeted by siRNA.…”

Section: Resultsmentioning

confidence: 99%

Timeless functions independently of the Tim-Tipin complex to promote sister chromatid cohesion in normal human fibroblasts

Smith‐Roe

Patel²,

Simpson³

et al. 2011

Cell Cycle

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Section: Resultsmentioning

confidence: 99%

Timeless functions independently of the Tim-Tipin complex to promote sister chromatid cohesion in normal human fibroblasts

Smith‐Roe

Patel²,

Simpson³

et al. 2011

Cell Cycle

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“…And secondly, both the full-length protein and caspase cleavage fragments of Claspin are stabilised in cells exposed to different proteasome inhibitors (Figure 4a and b). During the preparation of this manuscript, a report was published in which the proteasomal degradation of Claspin in normal cells was described; 25 however, the degradation of Claspin by the proteasome during apoptosis is a novel observation. The proteasome plays a complex role in cell death, having both negative and positive effects on apoptosis.…”

Section: Discussionmentioning

confidence: 99%

Cleavage and degradation of Claspin during apoptosis by caspases and the proteasome

et al. 2007

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Apoptosis plays a crucial role in development and tissue homeostasis. Some key survival pathways, such as DNA damage checkpoints and DNA repair, have been described to be inactivated during apoptosis. Here, we describe the processing of the human checkpoint protein Claspin during apoptosis. We observed cleavage of Claspin into multiple fragments in vivo. In vitro cleavage with caspases 3 and 7 of various fragments of the protein, revealed cut sites near the N-and C-termini of the protein.Using mass spectrometry, we identified a novel caspase cleavage site in Claspin at Asp25. Importantly, in addition to cleavage by caspases, we observed a proteasome-dependent degradation of Claspin under apoptotic conditions, resulting in a reduction of the levels of both full-length Claspin and its cleavage products. This degradation was not dependent upon the DSGxxS phosphodegron motif required for SCF b-TrCP -mediated ubiquitination of Claspin. Finally, downregulation of Claspin protein levels by short interfering RNA resulted in an increase in apoptotic induction both in the presence and absence of DNA damage. We conclude that Claspin has antiapoptotic activity and is degraded by two different pathways during apoptosis. The resulting disappearance of Claspin from the cells further promotes apoptosis.

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“…The antibody was affinity-purified using SulfoLink Plus Immobilization kit (Pierce). BACH1, Claspin, Chk2, 53BP1, MDC1, and H2AX antibodies have been previously described (8,12). Polyclonal anti-rabbit MCPH1 was purchased from Abcam (ab2612) for Western blotting.…”

Section: Methodsmentioning

confidence: 99%

MCPH1 Functions in an H2AX-dependent but MDC1-independent Pathway in Response to DNA Damage

Wood

Singh

Mer

et al. 2007

Journal of Biological Chemistry

107

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Microcephalin (MCPH1) is one of the causative genes for the autosomal recessive disorder, primary microcephaly, characterized by dramatic reduction in brain size and mental retardation. MCPH1 also functions in the DNA damage response, participating in cell cycle checkpoint control. However, how MCPH1 is regulated in the DNA damage response still remains unknown. Here we report that the ability of MCPH1 to localize to the sites of DNA double-strand breaks depends on its C-terminal tandem BRCT domains. Although MCPH1 foci formation depends on H2AX phosphorylation after DNA damage, it can occur independently of MDC1. We also show that MCPH1 binds to a phospho-H2AX peptide in vitro with an affinity similar to that of MDC1, and overexpression of wild type, but not C-BRCT mutants of MCPH1, can interfere with the foci formation of MDC1 and 53BP1. Collectively, our data suggest MCPH1 is recruited to double-strand breaks via its interaction with ␥H2AX, which is mediated by MCPH1 C-terminal BRCT domains. These observations support that MCPH1 acts early in DNA damage responsive pathways.Cell cycle checkpoints are critical for detecting damaged DNA and delaying cell cycle progression to allow time for DNA repair. If the damage is too great, they can also trigger cell death. Increasing evidence has shown that defects within these DNA damage checkpoints can lead to genetic instability, a hallmark of cancer cells (1). The DNA damage response is a complex network of proteins that can sense DNA lesions and relay and amplify the signal to downstream effectors to promote accurate and safe transmission of genetic material to the next generation. This pathway is usually classified as having groups of proteins involved in different steps; that is, sensors, mediators, and effectors. Canonically, the DNA damage response is a signal transduction cascade triggered by a series of phosphorylation-dependent events that activate proteins involved in transducing the DNA damage signal (ionizing radiation (IR) 3 or stalled replication) to different effector proteins that evoke specific effects, namely halting cell cycle progression, activating DNA repair mechanisms and transcription, and triggering apoptosis. The phosphatidylinositol 3-kinase-like kinase ataxiatelangiectasia-mutated (ATM) becomes active upon IR and relocalizes to the sites of double-strand breaks and phosphorylates H2AX, a histone variant that marks sites of damaged DNA. This then allows for the recruitment of other mediator proteins like MDC1, BRCA1, and 53BP1. These proteins help promote the damage signal to Chk1 and Chk2, which can phosphorylate various substrates to modulate the cell cycle, DNA repair, and apoptosis. However, increasing evidence has shown that the DNA damage response is not a linear pathway; DNA damage proteins can also act both upstream and downstream of their respective functions and maintain cross-talk with other proteins. Instead of a linear pathway as previously thought, there is an intricate network of proteins that function independently and dependen...

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Chk1 is required to maintain Claspin stability

Cited by 55 publications

References 23 publications

Timeless functions independently of the Tim-Tipin complex to promote sister chromatid cohesion in normal human fibroblasts

Timeless functions independently of the Tim-Tipin complex to promote sister chromatid cohesion in normal human fibroblasts

Cleavage and degradation of Claspin during apoptosis by caspases and the proteasome

MCPH1 Functions in an H2AX-dependent but MDC1-independent Pathway in Response to DNA Damage

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