Genotoxin-induced Rad9-Hus1-Rad1 (9-1-1) Chromatin Association Is an Early Checkpoint Signaling Event

Rad17, Rad1, Hus1, and Rad9 are key participants in checkpoint signaling pathways that block cell cycle progression in response to genotoxins. Biochemical and molecular modeling data predict that Rad9, Hus1, and Rad1 form a heterotrimeric complex, dubbed 9-1-1, which is loaded onto chromatin by a co...

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Bibliographic Details
Published in:The Journal of biological chemistry 2002-11, Vol.277 (46), p.43809-43812
Main Authors: Roos-Mattjus, Pia, Vroman, Benjamin T, Burtelow, Matthew A, Rauen, Matthew, Eapen, Alex K, Karnitz, Larry M
Format: Article
Language:English
Subjects:
Cell Cycle Proteins - chemistry
Cell Cycle Proteins - metabolism
Chromatin - metabolism
DNA - metabolism
DNA Damage
DNA, Complementary - metabolism
DNA-Binding Proteins
Endonucleases - chemistry
Endonucleases - metabolism
G1 Phase
Humans
Immunoblotting
K562 Cells
Mutation
Phosphorylation
Protein Binding
Schizosaccharomyces pombe Proteins
Signal Transduction
Transfection
Tumor Cells, Cultured
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Summary:Rad17, Rad1, Hus1, and Rad9 are key participants in checkpoint signaling pathways that block cell cycle progression in response to genotoxins. Biochemical and molecular modeling data predict that Rad9, Hus1, and Rad1 form a heterotrimeric complex, dubbed 9-1-1, which is loaded onto chromatin by a complex of Rad17 and the four small replication factor C (RFC) subunits (Rad17-RFC) in response to DNA damage. It is unclear what checkpoint proteins or checkpoint signaling events regulate the association of the 9-1-1 complex with DNA. Here we show that genotoxin-induced chromatin binding of 9-1-1 does not require the Rad9-inducible phosphorylation site (Ser-272). Although we found that Rad9 undergoes an additional phosphatidylinositol 3-kinase-related kinase (PIKK)-dependent posttranslational modification, we also show that genotoxin-triggered 9-1-1 chromatin binding does not depend on the catalytic activity of the PIKKs ataxia telangiectasia-mutated (ATM), ataxia telangiectasia and Rad3-related (ATR), or DNA-PK. Additionally, 9-1-1 chromatin binding does not require DNA replication, suggesting that the complex can be loaded onto DNA in response to DNA structures other than stalled DNA replication forks. Collectively, these studies demonstrate that 9-1-1 chromatin binding is a proximal event in the checkpoint signaling cascade.
ISSN:0021-9258
1083-351X
DOI:10.1074/jbc.M207272200