Tipin-replication Protein A Interaction Mediates Chk1 Phosphorylation by ATR in Response to Genotoxic Stress

Overview

Journal J Biol Chem

Specialty Biochemistry

Date 2010 Mar 18

PMID 20233725

Citations 73

Authors

Michael G Kemp

Zafer Akan

Secil Yilmaz

Mary Grillo

Stephanie L Smith-Roe

Tae-Hong Kang

Marila Cordeiro-Stone

William K Kaufmann

Robert T Abraham

Aziz Sancar

Keziban Unsal-Kacmaz

Affiliations

Soon will be listed here.

Abstract

Mammalian Timeless is a multifunctional protein that performs essential roles in the circadian clock, chromosome cohesion, DNA replication fork protection, and DNA replication/DNA damage checkpoint pathways. The human Timeless exists in a tight complex with a smaller protein called Tipin (Timeless-interacting protein). Here we investigated the mechanism by which the Timeless-Tipin complex functions as a mediator in the ATR-Chk1 DNA damage checkpoint pathway. We find that the Timeless-Tipin complex specifically mediates Chk1 phosphorylation by ATR in response to DNA damage and replication stress through interaction of Tipin with the 34-kDa subunit of replication protein A (RPA). The Tipin-RPA interaction stabilizes Timeless-Tipin and Tipin-Claspin complexes on RPA-coated ssDNA and in doing so promotes Claspin-mediated phosphorylation of Chk1 by ATR. Our results therefore indicate that RPA-covered ssDNA not only supports recruitment and activation of ATR but also, through Tipin and Claspin, it plays an important role in the action of ATR on its critical downstream target Chk1.

Citing Articles

Metabolic Rewiring in the Face of Genomic Assault: Integrating DNA Damage Response and Cellular Metabolism.

Ma W, Zhou S Biomolecules. 2025; 15(2).

PMID: 40001471 PMC: 11852599. DOI: 10.3390/biom15020168.

Intrinsic PARG inhibitor sensitivity is mimicked by haploinsufficiency and rescued by nucleoside supplementation.

Coulson-Gilmer C, Littler S, Barnes B, Brady R, Anagho H, Pillay N NAR Cancer. 2024; 6(3):zcae030.

PMID: 39015544 PMC: 11249981. DOI: 10.1093/narcan/zcae030.

The TIMELESS and PARP1 interaction suppresses replication-associated DNA gap accumulation.

Saldanha J, Rageul J, Patel J, Phi A, Lo N, Park J Nucleic Acids Res. 2024; 52(11):6424-6440.

PMID: 38801073 PMC: 11194094. DOI: 10.1093/nar/gkae445.

The MCM2-7 Complex: Roles beyond DNA Unwinding.

Rankin B, Rankin S Biology (Basel). 2024; 13(4).

PMID: 38666870 PMC: 11048021. DOI: 10.3390/biology13040258.

Timeless-Tipin interactions with MCM and RPA mediate DNA replication stress response.

Prorok P, Wolf E, Cardoso M Front Cell Dev Biol. 2024; 12:1346534.

PMID: 38487270 PMC: 10939015. DOI: 10.3389/fcell.2024.1346534.

References

Lee J, Paull T . Activation and regulation of ATM kinase activity in response to DNA double-strand breaks. Oncogene. 2007; 26(56):7741-8. DOI: 10.1038/sj.onc.1210872. View

Stokes M, Rush J, MacNeill J, Min Ren J, Sprott K, Nardone J . Profiling of UV-induced ATM/ATR signaling pathways. Proc Natl Acad Sci U S A. 2007; 104(50):19855-60. PMC: 2148387. DOI: 10.1073/pnas.0707579104. View

Bucher N, Britten C . G2 checkpoint abrogation and checkpoint kinase-1 targeting in the treatment of cancer. Br J Cancer. 2008; 98(3):523-8. PMC: 2243162. DOI: 10.1038/sj.bjc.6604208. View

Cimprich K, Cortez D . ATR: an essential regulator of genome integrity. Nat Rev Mol Cell Biol. 2008; 9(8):616-27. PMC: 2663384. DOI: 10.1038/nrm2450. View

Liu Q, Guntuku S, Cui X, Matsuoka S, Cortez D, Tamai K . Chk1 is an essential kinase that is regulated by Atr and required for the G(2)/M DNA damage checkpoint. Genes Dev. 2000; 14(12):1448-59. PMC: 316686. View

Mendez J, Stillman B . Chromatin association of human origin recognition complex, cdc6, and minichromosome maintenance proteins during the cell cycle: assembly of prereplication complexes in late mitosis. Mol Cell Biol. 2000; 20(22):8602-12. PMC: 102165. DOI: 10.1128/MCB.20.22.8602-8612.2000. View

MER G, Bochkarev A, Gupta R, Bochkareva E, Frappier L, Ingles C . Structural basis for the recognition of DNA repair proteins UNG2, XPA, and RAD52 by replication factor RPA. Cell. 2000; 103(3):449-56. DOI: 10.1016/s0092-8674(00)00136-7. View

Kumagai A, Dunphy W . Claspin, a novel protein required for the activation of Chk1 during a DNA replication checkpoint response in Xenopus egg extracts. Mol Cell. 2000; 6(4):839-49. DOI: 10.1016/s1097-2765(05)00092-4. View

Zhao H, Piwnica-Worms H . ATR-mediated checkpoint pathways regulate phosphorylation and activation of human Chk1. Mol Cell Biol. 2001; 21(13):4129-39. PMC: 87074. DOI: 10.1128/MCB.21.13.4129-4139.2001. View

10.

Mason A, Haring S, Pryor J, Staloch C, Gan T, Wold M . An alternative form of replication protein a prevents viral replication in vitro. J Biol Chem. 2009; 284(8):5324-31. PMC: 2643524. DOI: 10.1074/jbc.M808963200. View

11.

Choi J, Lindsey-Boltz L, Sancar A . Cooperative activation of the ATR checkpoint kinase by TopBP1 and damaged DNA. Nucleic Acids Res. 2009; 37(5):1501-9. PMC: 2655664. DOI: 10.1093/nar/gkn1075. View

12.

Kerzendorfer C, ODriscoll M . Human DNA damage response and repair deficiency syndromes: linking genomic instability and cell cycle checkpoint proficiency. DNA Repair (Amst). 2009; 8(9):1139-52. DOI: 10.1016/j.dnarep.2009.04.018. View

13.

Stracker T, Usui T, Petrini J . Taking the time to make important decisions: the checkpoint effector kinases Chk1 and Chk2 and the DNA damage response. DNA Repair (Amst). 2009; 8(9):1047-54. PMC: 2725228. DOI: 10.1016/j.dnarep.2009.04.012. View

14.

Tanaka H, Kubota Y, Tsujimura T, Kumano M, Masai H, Takisawa H . Replisome progression complex links DNA replication to sister chromatid cohesion in Xenopus egg extracts. Genes Cells. 2009; 14(8):949-63. DOI: 10.1111/j.1365-2443.2009.01322.x. View

15.

Zhang Y, Brognard J, Coughlin C, You Z, Dolled-Filhart M, Aslanian A . The F box protein Fbx6 regulates Chk1 stability and cellular sensitivity to replication stress. Mol Cell. 2009; 35(4):442-53. PMC: 2736145. DOI: 10.1016/j.molcel.2009.06.030. View

16.

Smith K, Fu M, Brown E . Tim-Tipin dysfunction creates an indispensible reliance on the ATR-Chk1 pathway for continued DNA synthesis. J Cell Biol. 2009; 187(1):15-23. PMC: 2762102. DOI: 10.1083/jcb.200905006. View

17.

Vassin V, Anantha R, Sokolova E, Kanner S, Borowiec J . Human RPA phosphorylation by ATR stimulates DNA synthesis and prevents ssDNA accumulation during DNA-replication stress. J Cell Sci. 2009; 122(Pt 22):4070-80. PMC: 2776501. DOI: 10.1242/jcs.053702. View

18.

Lindsey-Boltz L, Sercin O, Choi J, Sancar A . Reconstitution of human claspin-mediated phosphorylation of Chk1 by the ATR (ataxia telangiectasia-mutated and rad3-related) checkpoint kinase. J Biol Chem. 2009; 284(48):33107-14. PMC: 2785152. DOI: 10.1074/jbc.M109.064485. View

19.

Kemp M, Mason A, Carreira A, Reardon J, Haring S, Borgstahl G . An alternative form of replication protein a expressed in normal human tissues supports DNA repair. J Biol Chem. 2009; 285(7):4788-97. PMC: 2836084. DOI: 10.1074/jbc.M109.079418. View

20.

Haring S, Humphreys T, Wold M . A naturally occurring human RPA subunit homolog does not support DNA replication or cell-cycle progression. Nucleic Acids Res. 2009; 38(3):846-58. PMC: 2817474. DOI: 10.1093/nar/gkp1062. View