DNA-PKcs is required to maintain stability of Chk1 and Claspin for optimal replication stress response.
The ataxia telangiectasia mutated and Rad3-related (ATR)-checkpoint kinase 1 (Chk1) axis is the major signaling pathway activated in response to replication stress and is essential for the intra-S checkpoint. ATR phosphorylates and activates a number of molecules to coordinate cell cycle progression. Chk1 is the major effector downstream from ATR and plays a critical role in intra-S checkpoint on replication stress. Activation of Chk1 kinase also requires its association with Claspin, an adaptor protein essential for Chk1 protein stability, recruitment and ATR-dependent Chk1 phosphorylation. We have previously reported that, on replication stress, the catalytic subunit of DNA-dependent protein kinase (DNA-PKcs) is rapidly phosphorylated by ATR at the stalled replication forks and is required for cellular resistance to replication stresses although the impact of DNA-PKcs onto the ATR signaling pathway remains elusive. Here we report that ATR-dependent Chk1 phosphorylation and Chk1 signaling are compromised in the absence of DNA-PKcs. Our investigation reveals that DNA-PKcs is required to maintain Chk1-Claspin complex stability and transcriptional regulation of Claspin expression. The impaired Chk1 activity results in a defective intra-S checkpoint response in DNA-PKcs-deficient cells. Taken together, these results suggest that DNA-PKcs, in addition to its direct role in DNA damage repair, facilitates ATR-Chk1 signaling pathway in response to replication stress.
Division of Molecular Radiation Biology, Department of Radiation Oncology, University of Texas, Southwestern Medical Center at Dallas, Dallas, TX 75390, USA and Division of Molecular Pharmacology, Department of Pathophysiological and Therapeutic Science, Tottori University, Japan.
Yu-Fen
YF
Lin
Hung-Ying
HY
Shih
Zengfu
Z
Shang
Shinji
S
Matsunaga
Benjamin Pc
BP
Chen
D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism*; D064007:Ataxia Telangiectasia Mutated Proteins / Q000378:metabolism; D045744:Cell Line, Tumor; D000071877:Checkpoint Kinase 1; D002843:Chromatin / Q000378:metabolism; D004261:DNA Replication; D051747:DNA-Activated Protein Kinase / Q000235:genetics / Q000502:physiology*; D006801:Humans; D009154:Mutation; D009687:Nuclear Proteins / Q000235:genetics / Q000502:physiology*; D011494:Protein Kinases / Q000378:metabolism*; D055550:Protein Stability; D059807:S Phase Cell Cycle Checkpoints; D013312:Stress, Physiological / Q000235:genetics
D048868:Adaptor Proteins, Signal Transducing; C488158:CLSPN protein, human; D002843:Chromatin; D009687:Nuclear Proteins; D011494:Protein Kinases; C576597:ATR protein, human; D064007:Ataxia Telangiectasia Mutated Proteins; C000605867:CHEK1 protein, human; D000071877:Checkpoint Kinase 1; D051747:DNA-Activated Protein Kinase; C497263:PRKDC protein, human