This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Poon, R. Y. C. Articles by Hunter, T. Search for Related Content PubMed PubMed Citation Articles by Poon, R. Y. C. Articles by Hunter, T.

The Role of Cdc2 Feedback Loop Control in the DNA Damage Checkpoint in Mammalian Cells¹

Department of Biochemistry, The Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong [R. Y. C. P., M. S. C.]; Department of Microbiology, Faculty of Pharmaceutical Sciences, Kanazawa University, 13-1, Takara-machi, Kanazawa 920, Japan [K. Y.]; and The Salk Institute for Biological Studies, La Jolla, California 92037 [T. H.]

DNA damage inactivates cyclin-dependent kinases (CDKs) and arrests the cell cycle. Following DNA damage, the G₁-S CDKs are inhibited by a mechanism involving p53-dependent induction of p21^Cip1/Waf1; but how the Cdc2 is inhibited is less apparent. We found that the signal generated by the DNA damage checkpoint in G₂ was dominant over that from the spindle microtubule-assembly checkpoint, because the high Cdc2 activity present in nocodazole or Taxol-arrested cells was reduced by DNA damage. Phosphorylation of the inhibitory residues in Cdc2, Thr¹⁴, and Tyr¹⁵ coincided with the inactivation of Cdc2 after DNA damage. Interpretation of this result, however, was not straightforward due to the regulation of Thr¹⁴/Tyr¹⁵ phosphorylation by feedback loops; hence, their phosphorylation can in principle result merely from the inhibition of Cdc2 activity. Consistent with this, Thr¹⁴/Tyr¹⁵ phosphorylation was induced when Cdc2 kinase activity was inhibited with butyrolactone-I. Given these complications, we undertook a more critical analysis of the mechanisms that regulate Cdc2 after DNA damage. Caffeine reversed the DNA damage-induced inhibition of Cdc2 by causing dephosphorylation of Cdc2, and this dephosphorylation still occurred even when the Cdc2 feedback loops were blocked with butyrolactone-I. These data suggest that the DNA damage checkpoint in part acts through Thr¹⁴/Tyr¹⁵ phosphorylation by a mechanism independent of Cdc2 activity, and this phosphorylation can be accentuated by the Cdc2 feedback loops involving Thr¹⁴/Tyr¹⁵ protein kinases and phosphatases. The kinase activity of the Wee1Hu Tyr¹⁵ protein kinase was unaltered after DNA damage, but the phosphatase activity of Cdc25C was reduced. Thus, the decrease in Cdc25C activity may in part account for the DNA damage-induced increase in Thr¹⁴/Tyr¹⁵ phosphorylation.

¹ This work was supported in part by the USPHS Grants CA14195 and CA39780 (to T. H.), the Research Grants Council Grant DAG96/97-SC26 (to R. Y. C. P.), and the Dan Charitable Trust Fund International Fellowship (to R. Y. C. P. and K. Y.). T. H. is an American Cancer Society Research Professor.

² To whom requests for reprints should be addressed, at Department of Biochemistry, The Hong Kong University of Science and Technology, Clear Water Bay, Kowloon, Hong Kong. Phone: 852-2358-8703; Fax: 852-2358-1552; E-mail: bcrandy@usthk.ust.hk.

Received 6/12/97. Accepted 9/19/97.

This article has been cited by other articles:

				Z. Jin, E. Homola, S. Tiong, and S. D. Campbell Drosophila Myt1 Is the Major Cdk1 Inhibitory Kinase for Wing Imaginal Disc Development Genetics, December 1, 2008; 180(4): 2123 - 2133. [Abstract] [Full Text] [PDF]

				A. Blasina, J. Hallin, E. Chen, M. E. Arango, E. Kraynov, J. Register, S. Grant, S. Ninkovic, P. Chen, T. Nichols, et al. Breaching the DNA damage checkpoint via PF-00477736, a novel small-molecule inhibitor of checkpoint kinase 1 Mol. Cancer Ther., August 1, 2008; 7(8): 2394 - 2404. [Abstract] [Full Text] [PDF]

				H. Hochegger, D. Dejsuphong, E. Sonoda, A. Saberi, E. Rajendra, J. Kirk, T. Hunt, and S. Takeda An essential role for Cdk1 in S phase control is revealed via chemical genetics in vertebrate cells J. Cell Biol., July 10, 2007; 178(2): 257 - 268. [Abstract] [Full Text] [PDF]

				T. K. Fung, H. T. Ma, and R. Y.C. Poon Specialized Roles of the Two Mitotic Cyclins in Somatic Cells: Cyclin A as an Activator of M Phase-promoting Factor Mol. Biol. Cell, May 1, 2007; 18(5): 1861 - 1873. [Abstract] [Full Text] [PDF]

				S.-M. Jangi, J. L. Diaz-Perez, B. Ochoa-Lizarralde, I. Martin-Ruiz, A. Asumendi, G. Perez-Yarza, J. Gardeazabal, J. L. Diaz-Ramon, and M. D. Boyano H1 histamine receptor antagonists induce genotoxic and caspase-2-dependent apoptosis in human melanoma cells Carcinogenesis, September 1, 2006; 27(9): 1787 - 1796. [Abstract] [Full Text] [PDF]

				P. Bozko, M. Sabisz, A. K. Larsen, and A. Skladanowski Cross-talk between DNA damage and cell survival checkpoints during G2 and mitosis: pharmacologic implications Mol. Cancer Ther., December 1, 2005; 4(12): 2016 - 2025. [Abstract] [Full Text] [PDF]

				Y. Hirose, M. Katayama, O. K. Mirzoeva, M. S. Berger, and R. O. Pieper Akt Activation Suppresses Chk2-Mediated, Methylating Agent-Induced G2 Arrest and Protects from Temozolomide-Induced Mitotic Catastrophe and Cellular Senescence Cancer Res., June 1, 2005; 65(11): 4861 - 4869. [Abstract] [Full Text] [PDF]

				E. Bayart, O. Grigorieva, S. Leibovitch, R. Onclercq-Delic, and M. Amor-Gueret A Major Role for Mitotic cdc2 Kinase Inactivation in the Establishment of the Mitotic DNA Damage Checkpoint Cancer Res., December 15, 2004; 64(24): 8954 - 8959. [Abstract] [Full Text] [PDF]

				Y.-P. Lu, Y.-R. Lou, Q.-Y. Peng, J.-G. Xie, and A. H. Conney Stimulatory Effect of Topical Application of Caffeine on UVB-Induced Apoptosis in the Epidermis of p53 and Bax Knockout Mice Cancer Res., July 15, 2004; 64(14): 5020 - 5027. [Abstract] [Full Text] [PDF]

				W. Y. Siu, A. Lau, T. Arooz, J. P.H. Chow, H. T.B. Ho, and R. Y.C. Poon Topoisomerase poisons differentially activate DNA damage checkpoints through ataxia-telangiectasia mutated-dependent and -independent mechanisms Mol. Cancer Ther., May 1, 2004; 3(5): 621 - 632. [Abstract] [Full Text] [PDF]

				J. P. H. Chow, W. Y. Siu, H. T. B. Ho, K. H. T. Ma, C. C. Ho, and R. Y. C. Poon Differential Contribution of Inhibitory Phosphorylation of CDC2 and CDK2 for Unperturbed Cell Cycle Control and DNA Integrity Checkpoints J. Biol. Chem., October 17, 2003; 278(42): 40815 - 40828. [Abstract] [Full Text] [PDF]

				S. M. Post, A. E. Tomkinson, and E. Y.-H. P. Lee The human checkpoint Rad protein Rad17 is chromatin-associated throughout the cell cycle, localizes to DNA replication sites, and interacts with DNA polymerase {epsilon} Nucleic Acids Res., October 1, 2003; 31(19): 5568 - 5575. [Abstract] [Full Text] [PDF]

				J. P.H. Chow, W. Y. Siu, T. K. Fung, W. M. Chan, A. Lau, T. Arooz, C.-P. Ng, K. Yamashita, and R. Y.C. Poon DNA Damage during the Spindle-Assembly Checkpoint Degrades CDC25A, Inhibits Cyclin-CDC2 Complexes, and Reverses Cells to Interphase Mol. Biol. Cell, October 1, 2003; 14(10): 3989 - 4002. [Abstract] [Full Text] [PDF]

				D. Cortez Caffeine Inhibits Checkpoint Responses without Inhibiting the Ataxia-Telangiectasia-mutated (ATM) and ATM- and Rad3-related (ATR) Protein Kinases J. Biol. Chem., September 26, 2003; 278(39): 37139 - 37145. [Abstract] [Full Text] [PDF]

				M. Motwani, C. Rizzo, F. Sirotnak, Y. She, and G. K. Schwartz Flavopiridol Enhances the Effect of Docetaxel in Vitro and in Vivo in Human Gastric Cancer Cells Mol. Cancer Ther., June 1, 2003; 2(6): 549 - 555. [Abstract] [Full Text] [PDF]

				Z.-y. Huang, Y. Wu, N. Hedrick, and D. H. Gutmann T-Cadherin-Mediated Cell Growth Regulation Involves G2 Phase Arrest and Requires p21CIP1/WAF1 Expression Mol. Cell. Biol., January 15, 2003; 23(2): 566 - 578. [Abstract] [Full Text] [PDF]

				T. K. Fung, W. Y. Siu, C. H. Yam, A. Lau, and R. Y. C. Poon Cyclin F Is Degraded during G2-M by Mechanisms Fundamentally Different from Other Cyclins J. Biol. Chem., September 13, 2002; 277(38): 35140 - 35149. [Abstract] [Full Text] [PDF]

				S. Dutertre, R. Sekhri, L. A. Tintignac, R. Onclercq-Delic, B. Chatton, C. Jaulin, and M. Amor-Gueret Dephosphorylation and Subcellular Compartment Change of the Mitotic Bloom's Syndrome DNA Helicase in Response to Ionizing Radiation J. Biol. Chem., February 15, 2002; 277(8): 6280 - 6286. [Abstract] [Full Text] [PDF]

				U. Aytac, F.-X. Claret, L. Ho, K. Sato, K. Ohnuma, G. B. Mills, F. Cabanillas, C. Morimoto, and N. H. Dang Expression of CD26 and Its Associated Dipeptidyl Peptidase IV Enzyme Activity Enhances Sensitivity to Doxorubicin-induced Cell Cycle Arrest at the G2/M Checkpoint Cancer Res., October 1, 2001; 61(19): 7204 - 7210. [Abstract] [Full Text] [PDF]

				B. Hu, J. Mitra, S. van den Heuvel, and G. H. Enders S and G2 Phase Roles for Cdk2 Revealed by Inducible Expression of a Dominant-Negative Mutant in Human Cells Mol. Cell. Biol., April 15, 2001; 21(8): 2755 - 2766. [Abstract] [Full Text]

				V. Gottifredi, O. Karni-Schmidt, S.-Y. Shieh, and C. Prives p53 Down-Regulates CHK1 through p21 and the Retinoblastoma Protein Mol. Cell. Biol., February 15, 2001; 21(4): 1066 - 1076. [Abstract] [Full Text]

				K. Kikuchi, K. Naito, J. Noguchi, A. Shimada, H. Kaneko, M. Yamashita, F. Aoki, H. Tojo, and Y. Toyoda Maturation/M-Phase Promoting Factor: A Regulator of Aging in Porcine Oocytes Biol Reprod, September 1, 2000; 63(3): 715 - 722. [Abstract] [Full Text]

				X. Wang, C. H. McGowan, M. Zhao, L. He, J. S. Downey, C. Fearns, Y. Wang, S. Huang, and J. Han Involvement of the MKK6-p38gamma Cascade in gamma -Radiation-Induced Cell Cycle Arrest Mol. Cell. Biol., July 1, 2000; 20(13): 4543 - 4552. [Abstract] [Full Text]

				N. Mailand, J. Falck, C. Lukas, R. G. Syljuåsen, M. Welcker, J. Bartek, and J. Lukas Rapid Destruction of Human Cdc25A in Response to DNA Damage Science, May 26, 2000; 288(5470): 1425 - 1429. [Abstract] [Full Text]

				J.-M. Darbon, M. Penary, N. Escalas, F. Casagrande, F. Goubin-Gramatica, C. Baudouin, and B. Ducommun Distinct Chk2 Activation Pathways Are Triggered by Genistein and DNA-damaging Agents in Human Melanoma Cells J. Biol. Chem., May 12, 2000; 275(20): 15363 - 15369. [Abstract] [Full Text] [PDF]

				Y. Wang, C. Jacobs, K. E. Hook, H. Duan, R. N. Booher, and Y. Sun Binding of 14-3-3{beta} to the Carboxyl Terminus of Wee1 Increases Wee1 Stability, Kinase Activity, and G2-M Cell Population Cell Growth Differ., April 1, 2000; 11(4): 211 - 219. [Abstract] [Full Text]

				J. A. Koutcher, M. Motwani, K. L. Zakian, X.-K. Li, C. Matei, J. P. Dyke, D. Ballon, H. H. Yoo, and G. K. Schwartz The in Vivo Effect of Bryostatin-1 on Paclitaxel-induced Tumor Growth, Mitotic Entry, and Blood Flow Clin. Cancer Res., April 1, 2000; 6(4): 1498 - 1507. [Abstract] [Full Text]

				V. W. HU and D. S. HEIKKA Radiolabeling revisited: metabolic labeling with 35S-methionine inhibits cell cycle progression, proliferation, and survival FASEB J, March 1, 2000; 14(3): 448 - 454. [Abstract] [Full Text]

				C. H. Yam, W. Y. Siu, A. Lau, and R. Y. C. Poon Degradation of Cyclin A Does Not Require Its Phosphorylation by CDC2 and Cyclin-dependent Kinase 2 J. Biol. Chem., February 4, 2000; 275(5): 3158 - 3167. [Abstract] [Full Text] [PDF]

				R. JORQUERA and R. M. TANGUAY Cyclin B-dependent kinase and caspase-1 activation precedes mitochondrial dysfunction in fumarylacetoacetate-induced apoptosis FASEB J, December 1, 1999; 13(15): 2284 - 2298. [Abstract] [Full Text]

				J.-S. Park, S. Carter, D. B. Reardon, R. Schmidt-Ullrich, P. Dent, and P. B. Fisher Roles for Basal and Stimulated p21Cip-1/WAF1/MDA6 Expression and Mitogen-activated Protein Kinase Signaling in Radiation-induced Cell Cycle Checkpoint Control in Carcinoma Cells Mol. Biol. Cell, December 1, 1999; 10(12): 4231 - 4246. [Abstract] [Full Text]

				W. R. Taylor, S. E. DePrimo, A. Agarwal, M. L. Agarwal, A. H. Schönthal, K. S. Katula, and G. R. Stark Mechanisms of G2 Arrest in Response to Overexpression of p53 Mol. Biol. Cell, November 1, 1999; 10(11): 3607 - 3622. [Abstract] [Full Text]

				B. D. Aguda A quantitative analysis of the kinetics of the G2 DNA damage checkpoint system PNAS, September 28, 1999; 96(20): 11352 - 11357. [Abstract] [Full Text] [PDF]

				M. Motwani, T. M. Delohery, and G. K. Schwartz Sequential Dependent Enhancement of Caspase Activation and Apoptosis by Flavopiridol on Paclitaxel-treated Human Gastric and Breast Cancer Cells Clin. Cancer Res., July 1, 1999; 5(7): 1876 - 1883. [Abstract] [Full Text] [PDF]

				N. Wells, N Watanabe, T Tokusumi, W Jiang, M. Verdecia, and T Hunter The C-terminal domain of the Cdc2 inhibitory kinase Myt1 interacts with Cdc2 complexes and is required for inhibition of G(2)/M progression J. Cell Sci., January 10, 1999; 112(19): 3361 - 3371. [Abstract] [PDF]

				S. Wang, C Norbury, A. Harris, and T Toda Caffeine can override the S-M checkpoint in fission yeast J. Cell Sci., January 3, 1999; 112(6): 927 - 937. [Abstract] [PDF]

				C. H. Yam, R. W. M. Ng, W. Yi Siu, A. W. S. Lau, and R. Y. C. Poon Regulation of Cyclin A-Cdk2 by SCF Component Skp1 and F-Box Protein Skp2 Mol. Cell. Biol., January 1, 1999; 19(1): 635 - 645. [Abstract] [Full Text] [PDF]

				Z. Weng, A.-C. Fluckiger, S. Nisitani, M. I. Wahl, L. Q. Le, C. A. Hunter, A. A. Fernal, M. M. Le Beau, and O. N. Witte A DNA damage and stress inducible G protein-coupled receptor blocks cells in G2/M PNAS, October 13, 1998; 95(21): 12334 - 12339. [Abstract] [Full Text] [PDF]

				A. Kumagai, Z. Guo, K. H. Emami, S. X. Wang, and W. G. Dunphy The Xenopus Chk1 Protein Kinase Mediates a Caffeine-sensitive Pathway of Checkpoint Control in Cell-free Extracts J. Cell Biol., September 21, 1998; 142(6): 1559 - 1569. [Abstract] [Full Text] [PDF]

				P. Gowdy, H. Anderson, and M Roberge Entry into mitosis without Cdc2 kinase activation J. Cell Sci., January 11, 1998; 111(22): 3401 - 3410. [Abstract] [PDF]

				G. Damia, Y. Sanchez, E. Erba, and M. Broggini DNA Damage Induces p53-dependent Down-regulation of hCHK1 J. Biol. Chem., March 30, 2001; 276(14): 10641 - 10645. [Abstract] [Full Text] [PDF]

				X. Cortes-Bratti, C. Karlsson, T. Lagergard, M. Thelestam, and T. Frisan The Haemophilus ducreyi Cytolethal Distending Toxin Induces Cell Cycle Arrest and Apoptosis via the DNA Damage Checkpoint Pathways J. Biol. Chem., February 9, 2001; 276(7): 5296 - 5302. [Abstract] [Full Text] [PDF]