Cellular processing determinants for the activation of damage signals in response to topoisomerase I-linked DNA breakage

Ting-Hsiang Huang, Hsiang-Chin Chen, Shang-Min Chou, Yu-Chen Yang, Jia-Rong Fan, Tsai-Kun Li

Research output: Contribution to journalArticle

8 Citations (Scopus)

Abstract

Recent studies have suggested an involvement of processing pathways for the initiation of cellular responses induced by topoisomerase-targeting drugs. Here, we showed that cellular exposure to camptothecin (CPT) induced formation of topoisomerase I cleavable complex (TOP1cc), degradation of TOP1 and activation of DNA damage responses (DDR). Transcription and proteasome-dependent proteolysis, but not replication, were involved in CPT-induced TOP1 degradation, while none of above three processing activities affected TOP1cc formation. Replication-and transcription-initiated processing (RIP and TIP) of TOP1cc were identified as two independent pathways, which contribute distinctly to various CPT-activated DDR. Specifically, in cycling cells, RIP-processed TOP1cc triggered the CPT-induced RPA phosphorylation. At higher CPT dosages, the TIP pathway is required for other DDR activation, including ATM, p53 and Chk1/2 phosphorylation. The TIP pathway was further demonstrated to be S-phase independent by using three nonreplicating cell models. Furthermore, the effect of proteasome inhibitors mimicked that of transcription inhibition on the CPT-induced activation of DDR, suggesting the involvement of proteasome in the TIP pathway. Interestingly, the TIP pathway was important for TOP1cc-activated, but not ionization radiation-activated ATM, p53 and Chk2 phosphorylation. We have also found that pharmacological interferences of TIP and RIP pathways distinctively modulated the CPT-induced cell killing with treatments at low and high dosages, respectively. Together, our results support that both RIP and TIP pathways of TOP1cc are required for the activation of CPT-induced DDR and cytotoxicity. © 2010 IBCB, SIBS, CAS. All rights reserved.
Original languageEnglish
Pages (from-to)1060-1075
Number of pages16
JournalCell Research
Volume20
Issue number9
DOIs
Publication statusPublished - 2010
Externally publishedYes

Fingerprint

Camptothecin
Type I DNA Topoisomerase
DNA Damage
Phosphorylation
Proteasome Endopeptidase Complex
Proteasome Inhibitors
Drug Delivery Systems
S Phase
Proteolysis
Pharmacology
Radiation

Keywords

  • cleavable complex
  • DNA damage responses
  • downregulation
  • processing
  • protein-linked DNA break
  • ATM protein
  • camptothecin
  • cell cycle protein
  • checkpoint kinase 2
  • DNA binding protein
  • DNA topoisomerase
  • protein p53
  • protein serine threonine kinase
  • replication factor A
  • RPA2 protein, human
  • tumor suppressor protein
  • article
  • DNA repair
  • DNA replication
  • DNA strand breakage
  • genetic transcription
  • genetics
  • human
  • metabolism
  • phosphorylation
  • signal transduction
  • tumor cell line
  • Camptothecin
  • Cell Cycle Proteins
  • Cell Line, Tumor
  • DNA Breaks
  • DNA Repair
  • DNA Replication
  • DNA Topoisomerases, Type I
  • DNA-Binding Proteins
  • Humans
  • Phosphorylation
  • Protein-Serine-Threonine Kinases
  • Replication Protein A
  • Signal Transduction
  • Transcription, Genetic
  • Tumor Suppressor Protein p53
  • Tumor Suppressor Proteins

Cite this

Cellular processing determinants for the activation of damage signals in response to topoisomerase I-linked DNA breakage. / Huang, Ting-Hsiang; Chen, Hsiang-Chin; Chou, Shang-Min; Yang, Yu-Chen; Fan, Jia-Rong; Li, Tsai-Kun.

In: Cell Research, Vol. 20, No. 9, 2010, p. 1060-1075.

Research output: Contribution to journalArticle

Huang, Ting-Hsiang ; Chen, Hsiang-Chin ; Chou, Shang-Min ; Yang, Yu-Chen ; Fan, Jia-Rong ; Li, Tsai-Kun. / Cellular processing determinants for the activation of damage signals in response to topoisomerase I-linked DNA breakage. In: Cell Research. 2010 ; Vol. 20, No. 9. pp. 1060-1075.
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abstract = "Recent studies have suggested an involvement of processing pathways for the initiation of cellular responses induced by topoisomerase-targeting drugs. Here, we showed that cellular exposure to camptothecin (CPT) induced formation of topoisomerase I cleavable complex (TOP1cc), degradation of TOP1 and activation of DNA damage responses (DDR). Transcription and proteasome-dependent proteolysis, but not replication, were involved in CPT-induced TOP1 degradation, while none of above three processing activities affected TOP1cc formation. Replication-and transcription-initiated processing (RIP and TIP) of TOP1cc were identified as two independent pathways, which contribute distinctly to various CPT-activated DDR. Specifically, in cycling cells, RIP-processed TOP1cc triggered the CPT-induced RPA phosphorylation. At higher CPT dosages, the TIP pathway is required for other DDR activation, including ATM, p53 and Chk1/2 phosphorylation. The TIP pathway was further demonstrated to be S-phase independent by using three nonreplicating cell models. Furthermore, the effect of proteasome inhibitors mimicked that of transcription inhibition on the CPT-induced activation of DDR, suggesting the involvement of proteasome in the TIP pathway. Interestingly, the TIP pathway was important for TOP1cc-activated, but not ionization radiation-activated ATM, p53 and Chk2 phosphorylation. We have also found that pharmacological interferences of TIP and RIP pathways distinctively modulated the CPT-induced cell killing with treatments at low and high dosages, respectively. Together, our results support that both RIP and TIP pathways of TOP1cc are required for the activation of CPT-induced DDR and cytotoxicity. {\circledC} 2010 IBCB, SIBS, CAS. All rights reserved.",
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author = "Ting-Hsiang Huang and Hsiang-Chin Chen and Shang-Min Chou and Yu-Chen Yang and Jia-Rong Fan and Tsai-Kun Li",
note = "被引用次數:8 Export Date: 7 April 2016 通訊地址: Li, T.-K.; Department and Graduate Institute of Microbiology, College of Medicine, National Taiwan University, Jen-Ai Road, Taipei 10018, Taiwan; 電子郵件: tsaikunli@ntu.edu.tw 化學物質/CAS: DNA topoisomerase, 80449-01-0; camptothecin, 7689-03-4; checkpoint kinase 2, 244634-79-5; Camptothecin, 7689-03-4; Cell Cycle Proteins; DNA Topoisomerases, Type I, 5.99.1.2; DNA-Binding Proteins; Protein-Serine-Threonine Kinases, 2.7.11.1; RPA2 protein, human, 2.7.7.7; Replication Protein A; Tumor Suppressor Protein p53; Tumor Suppressor Proteins; ataxia telangiectasia mutated protein, 2.7.1.37; checkpoint kinase 2, 2.7.1.37 參考文獻: Zhou, B.B., Bartek, J., Targeting the checkpoint kinases: Chemo-sensitization versus chemoprotection (2004) Nat Rev Cancer, 4, pp. 216-225; Falck, J., Coates, J., Jackson, S.P., Conserved modes of recruitment of ATM. ATR and DNA-PKcs to sites of DNA damage (2005) Nature, 434, pp. 605-611; Sancar, A., Lindsey-Boltz, L.A., Unsal-Kacmaz, K., Linn, S., Molecular mechanisms of mammalian DNA repair and the DNA damage checkpoints (2004) Annu Rev Biochem, 73, pp. 39-85; Li, T.K., Liu, L.F., Tumor cell death induced by topoisomerase-targeting drugs (2001) Annu Rev Pharmacol Toxicol, 41, pp. 53-77; Pommier, Y., Topoisomerase i inhibitors: Camptothecins and beyond (2006) Nat Rev Cancer, 6, pp. 789-802; Nitiss, J.L., Targeting DNA topoisomerase II in cancer chemotherapy (2009) Nat Rev Cancer, 9, pp. 338-350; Connelly, J.C., Leach, D.R., Repair of DNA covalently linked to protein (2004) Mol Cell, 13, pp. 307-316; Staker, B.L., Hjerrild, K., Feese, M.D., The mechanism of topoisomerase i poisoning by a camptothecin analog (2002) Proc Natl Acad Sci USA, 99, pp. 15387-15392; Stewart, L., Redinbo, M.R., Qiu, X., Hol, W.G., Champoux, J.J., A model for the mechanism of human topoisomerase i (1998) Science, 279, pp. 1534-1541; Redinbo, M.R., Stewart, L., Kuhn, P., Champoux, J.J., Hol, W.G., Crystal structures of human topoisomerase i in covalent and noncovalent complexes with DNA (1998) Science, 279, pp. 1504-1513; Sordet, O., Redon, C.E., Guirouilh-Barbat, J., Ataxia te-langiectasia mutated activation by transcription-and topoi-somerase I-induced DNA double-strand breaks (2009) EMBO Rep, 10, pp. 887-893; Lin, C.P., Ban, Y., Lyu, Y.L., Liu, L.F., Proteasome-dependent processing of topoisomerase I-DNA adducts into DNA double strand breaks at arrested replication forks (2009) J Biol Chem, 284, pp. 28084-28092; Lin, C.P., Ban, Y., Lyu, Y.L., Desai, S.D., Liu, L.F., A ubiquitin-proteasome pathway for the repair of topoisomerase I-DNA covalent complexes (2008) J Biol Chem, 283, pp. 21074-21083; Fan, J.R., Peng, A.L., Chen, H.C., Cellular processing pathways contribute to the activation of etoposide-induced DNA damage responses (2008) DNA Repair (Amst), 7, pp. 452-463; Zhang, A., Lyu, Y.L., Lin, C.P., A protease pathway for the repair of topoisomerase II-DNA covalent complexes (2006) J Biol Chem, 281, pp. 35997-36003; Takemura, H., Rao, V.A., Sordet, O., Defective Mre11-dependent activation of Chk2 by ataxia telangiectasia mutated in colorectal carcinoma cells in response to replication-dependent DNA double strand breaks (2006) J Biol Chem, 281, pp. 30814-30823; Wu, J.X., Liu, L.F., Processing of topoisomerase i cleavable complexes into DNA damage by transcription (1997) Nucleic Acids Res, 25, pp. 4181-4186; Tsao, Y.P., Russo, A., Nyamuswa, G., Silber, R., Liu, L.F., Interaction between replication forks and topoisomerase-I-DNA cleav-able complexes-studies in a cell-free Sv40 DNA-replication system (1993) Cancer Res, 53, pp. 5908-5914; Hsiang, Y.H., Lihou, M.G., Liu, L.F., Arrest of replication forks by drug-stabilized topoisomerase i-dna cleavable complexes as a mechanism of cell killing by camptothecin (1989) Cancer Res, 49, pp. 5077-5082; Morris, E.J., Geller, H.M., Induction of neuronal apoptosis by camptothecin, an inhibitor of DNA topoisomerase-I: Evidence for cell cycle-independent toxicity (1996) J Cell Biol, 134, pp. 757-770; Shao, R.G., Cao, C.X., Zhang, H.L., Replication-mediated DNA damage by camptothecin induces phosphorylation of RPA by DNA-dependent protein kinase and dissociates RPA: DNA-PK complexes (1999) EMBO J, 18, pp. 1397-1406; Desai, S.D., Zhang, H., Rodriguez-Bauman, A., Transcription-dependent degradation of topoisomerase I-DNA covalent complexes (2003) Mol Cel Biol, 23, pp. 2341-2350; Desai, S.D., Li, T.K., Rodriguez-Bauman, A., Rubin, E.H., Liu, L.F., Ubiquitin/26S proteasome-mediated degradation of topoisomerase i as a resistance mechanism to camptothecin in tumor cells (2001) Cancer Res, 61, pp. 5926-5932; Daroui, P., Desai, S.D., Li, T.K., Liu, A.A., Liu, L.F., Hydrogen peroxide induces topoisomerase I-mediated DNA damage and cell death (2004) J Biol Chem, 279, pp. 14587-14594; Subramanian, D., Kraut, E., Staubus, A., Young, D.C., Muller, M.T., Analysis of topoisomerase I/DNA complexes in patients administered topotecan (1995) Cancer Res, 55, pp. 2097-2103; Afshari, C.A., Barrett, J.C., Disruption of G0-G1 arrest in quiescent and senescent cells treated with phosphatase inhibitors (1994) Cancer Res, 54, pp. 2317-2321; Li, T.K., Houghton, P.J., Desai, S.D., Characterization of ARC-111 as a novel topoisomerase I-targeting anticancer drug (2003) Cancer Res, 63, pp. 8400-8407; Siu, W.Y., Lau, A., Arooz, T., Topoisomerase poisons differentially activate DNA damage checkpoints through ataxia-telangiectasia mutated-dependent and-independent mechanisms (2004) Mol Cancer Ther, 3, pp. 621-632; Bakkenist, C.J., Kastan, M.B., DNA damage activates ATM through intermolecular autophosphorylation and dimer dissociation (2003) Nature, 421, pp. 499-506; D'Arpa, P., Beardmore, C., Liu, L.F., Involvement of nucleic acid synthesis in cell killing mechanisms of topoisomerase poisons (1990) Cancer Res, 50, pp. 6919-6924; Furuta, T., Takemura, H., Liao, Z.Y., Phosphorylation of histone H2AX and activation of Mre11, Rad50, and Nbs1 in response to replication-dependent DNA double-strand breaks induced by mammalian DNA topoisomerase i cleavage complexes (2003) J Biol Chem, 278, pp. 20303-20312; Liu, L.F., Desai, S.D., Li, T.K., Mao, Y., Sun, M., Sim, S.P., Mechanism of action of camptothecin (2000) Ann N y Acad Sci, 922, pp. 1-10; Adams, J., The development of proteasome inhibitors as anti-cancer drugs (2004) Cancer Cell, 5, pp. 417-421; Xu, Y., Villalona-Calero, M.A., Irinotecan: Mechanisms of tumor resistance and novel strategies for modulating its activity (2002) Ann Oncol, 13, pp. 1841-1851; Canman, C.E., Lim, D.S., Cimprich, K.A., Activation of the ATM kinase by ionizing radiation and phosphorylation of p53 (1998) Science, 281, pp. 1677-1679",
year = "2010",
doi = "10.1038/cr.2010.95",
language = "English",
volume = "20",
pages = "1060--1075",
journal = "Cell Research",
issn = "1001-0602",
publisher = "Nature Publishing Group",
number = "9",

}

TY - JOUR

T1 - Cellular processing determinants for the activation of damage signals in response to topoisomerase I-linked DNA breakage

AU - Huang, Ting-Hsiang

AU - Chen, Hsiang-Chin

AU - Chou, Shang-Min

AU - Yang, Yu-Chen

AU - Fan, Jia-Rong

AU - Li, Tsai-Kun

N1 - 被引用次數:8 Export Date: 7 April 2016 通訊地址: Li, T.-K.; Department and Graduate Institute of Microbiology, College of Medicine, National Taiwan University, Jen-Ai Road, Taipei 10018, Taiwan; 電子郵件: tsaikunli@ntu.edu.tw 化學物質/CAS: DNA topoisomerase, 80449-01-0; camptothecin, 7689-03-4; checkpoint kinase 2, 244634-79-5; Camptothecin, 7689-03-4; Cell Cycle Proteins; DNA Topoisomerases, Type I, 5.99.1.2; DNA-Binding Proteins; Protein-Serine-Threonine Kinases, 2.7.11.1; RPA2 protein, human, 2.7.7.7; Replication Protein A; Tumor Suppressor Protein p53; Tumor Suppressor Proteins; ataxia telangiectasia mutated protein, 2.7.1.37; checkpoint kinase 2, 2.7.1.37 參考文獻: Zhou, B.B., Bartek, J., Targeting the checkpoint kinases: Chemo-sensitization versus chemoprotection (2004) Nat Rev Cancer, 4, pp. 216-225; Falck, J., Coates, J., Jackson, S.P., Conserved modes of recruitment of ATM. ATR and DNA-PKcs to sites of DNA damage (2005) Nature, 434, pp. 605-611; Sancar, A., Lindsey-Boltz, L.A., Unsal-Kacmaz, K., Linn, S., Molecular mechanisms of mammalian DNA repair and the DNA damage checkpoints (2004) Annu Rev Biochem, 73, pp. 39-85; Li, T.K., Liu, L.F., Tumor cell death induced by topoisomerase-targeting drugs (2001) Annu Rev Pharmacol Toxicol, 41, pp. 53-77; Pommier, Y., Topoisomerase i inhibitors: Camptothecins and beyond (2006) Nat Rev Cancer, 6, pp. 789-802; Nitiss, J.L., Targeting DNA topoisomerase II in cancer chemotherapy (2009) Nat Rev Cancer, 9, pp. 338-350; Connelly, J.C., Leach, D.R., Repair of DNA covalently linked to protein (2004) Mol Cell, 13, pp. 307-316; Staker, B.L., Hjerrild, K., Feese, M.D., The mechanism of topoisomerase i poisoning by a camptothecin analog (2002) Proc Natl Acad Sci USA, 99, pp. 15387-15392; Stewart, L., Redinbo, M.R., Qiu, X., Hol, W.G., Champoux, J.J., A model for the mechanism of human topoisomerase i (1998) Science, 279, pp. 1534-1541; Redinbo, M.R., Stewart, L., Kuhn, P., Champoux, J.J., Hol, W.G., Crystal structures of human topoisomerase i in covalent and noncovalent complexes with DNA (1998) Science, 279, pp. 1504-1513; Sordet, O., Redon, C.E., Guirouilh-Barbat, J., Ataxia te-langiectasia mutated activation by transcription-and topoi-somerase I-induced DNA double-strand breaks (2009) EMBO Rep, 10, pp. 887-893; Lin, C.P., Ban, Y., Lyu, Y.L., Liu, L.F., Proteasome-dependent processing of topoisomerase I-DNA adducts into DNA double strand breaks at arrested replication forks (2009) J Biol Chem, 284, pp. 28084-28092; Lin, C.P., Ban, Y., Lyu, Y.L., Desai, S.D., Liu, L.F., A ubiquitin-proteasome pathway for the repair of topoisomerase I-DNA covalent complexes (2008) J Biol Chem, 283, pp. 21074-21083; Fan, J.R., Peng, A.L., Chen, H.C., Cellular processing pathways contribute to the activation of etoposide-induced DNA damage responses (2008) DNA Repair (Amst), 7, pp. 452-463; Zhang, A., Lyu, Y.L., Lin, C.P., A protease pathway for the repair of topoisomerase II-DNA covalent complexes (2006) J Biol Chem, 281, pp. 35997-36003; Takemura, H., Rao, V.A., Sordet, O., Defective Mre11-dependent activation of Chk2 by ataxia telangiectasia mutated in colorectal carcinoma cells in response to replication-dependent DNA double strand breaks (2006) J Biol Chem, 281, pp. 30814-30823; Wu, J.X., Liu, L.F., Processing of topoisomerase i cleavable complexes into DNA damage by transcription (1997) Nucleic Acids Res, 25, pp. 4181-4186; Tsao, Y.P., Russo, A., Nyamuswa, G., Silber, R., Liu, L.F., Interaction between replication forks and topoisomerase-I-DNA cleav-able complexes-studies in a cell-free Sv40 DNA-replication system (1993) Cancer Res, 53, pp. 5908-5914; Hsiang, Y.H., Lihou, M.G., Liu, L.F., Arrest of replication forks by drug-stabilized topoisomerase i-dna cleavable complexes as a mechanism of cell killing by camptothecin (1989) Cancer Res, 49, pp. 5077-5082; Morris, E.J., Geller, H.M., Induction of neuronal apoptosis by camptothecin, an inhibitor of DNA topoisomerase-I: Evidence for cell cycle-independent toxicity (1996) J Cell Biol, 134, pp. 757-770; Shao, R.G., Cao, C.X., Zhang, H.L., Replication-mediated DNA damage by camptothecin induces phosphorylation of RPA by DNA-dependent protein kinase and dissociates RPA: DNA-PK complexes (1999) EMBO J, 18, pp. 1397-1406; Desai, S.D., Zhang, H., Rodriguez-Bauman, A., Transcription-dependent degradation of topoisomerase I-DNA covalent complexes (2003) Mol Cel Biol, 23, pp. 2341-2350; Desai, S.D., Li, T.K., Rodriguez-Bauman, A., Rubin, E.H., Liu, L.F., Ubiquitin/26S proteasome-mediated degradation of topoisomerase i as a resistance mechanism to camptothecin in tumor cells (2001) Cancer Res, 61, pp. 5926-5932; Daroui, P., Desai, S.D., Li, T.K., Liu, A.A., Liu, L.F., Hydrogen peroxide induces topoisomerase I-mediated DNA damage and cell death (2004) J Biol Chem, 279, pp. 14587-14594; Subramanian, D., Kraut, E., Staubus, A., Young, D.C., Muller, M.T., Analysis of topoisomerase I/DNA complexes in patients administered topotecan (1995) Cancer Res, 55, pp. 2097-2103; Afshari, C.A., Barrett, J.C., Disruption of G0-G1 arrest in quiescent and senescent cells treated with phosphatase inhibitors (1994) Cancer Res, 54, pp. 2317-2321; Li, T.K., Houghton, P.J., Desai, S.D., Characterization of ARC-111 as a novel topoisomerase I-targeting anticancer drug (2003) Cancer Res, 63, pp. 8400-8407; Siu, W.Y., Lau, A., Arooz, T., Topoisomerase poisons differentially activate DNA damage checkpoints through ataxia-telangiectasia mutated-dependent and-independent mechanisms (2004) Mol Cancer Ther, 3, pp. 621-632; Bakkenist, C.J., Kastan, M.B., DNA damage activates ATM through intermolecular autophosphorylation and dimer dissociation (2003) Nature, 421, pp. 499-506; D'Arpa, P., Beardmore, C., Liu, L.F., Involvement of nucleic acid synthesis in cell killing mechanisms of topoisomerase poisons (1990) Cancer Res, 50, pp. 6919-6924; Furuta, T., Takemura, H., Liao, Z.Y., Phosphorylation of histone H2AX and activation of Mre11, Rad50, and Nbs1 in response to replication-dependent DNA double-strand breaks induced by mammalian DNA topoisomerase i cleavage complexes (2003) J Biol Chem, 278, pp. 20303-20312; Liu, L.F., Desai, S.D., Li, T.K., Mao, Y., Sun, M., Sim, S.P., Mechanism of action of camptothecin (2000) Ann N y Acad Sci, 922, pp. 1-10; Adams, J., The development of proteasome inhibitors as anti-cancer drugs (2004) Cancer Cell, 5, pp. 417-421; Xu, Y., Villalona-Calero, M.A., Irinotecan: Mechanisms of tumor resistance and novel strategies for modulating its activity (2002) Ann Oncol, 13, pp. 1841-1851; Canman, C.E., Lim, D.S., Cimprich, K.A., Activation of the ATM kinase by ionizing radiation and phosphorylation of p53 (1998) Science, 281, pp. 1677-1679

PY - 2010

Y1 - 2010

N2 - Recent studies have suggested an involvement of processing pathways for the initiation of cellular responses induced by topoisomerase-targeting drugs. Here, we showed that cellular exposure to camptothecin (CPT) induced formation of topoisomerase I cleavable complex (TOP1cc), degradation of TOP1 and activation of DNA damage responses (DDR). Transcription and proteasome-dependent proteolysis, but not replication, were involved in CPT-induced TOP1 degradation, while none of above three processing activities affected TOP1cc formation. Replication-and transcription-initiated processing (RIP and TIP) of TOP1cc were identified as two independent pathways, which contribute distinctly to various CPT-activated DDR. Specifically, in cycling cells, RIP-processed TOP1cc triggered the CPT-induced RPA phosphorylation. At higher CPT dosages, the TIP pathway is required for other DDR activation, including ATM, p53 and Chk1/2 phosphorylation. The TIP pathway was further demonstrated to be S-phase independent by using three nonreplicating cell models. Furthermore, the effect of proteasome inhibitors mimicked that of transcription inhibition on the CPT-induced activation of DDR, suggesting the involvement of proteasome in the TIP pathway. Interestingly, the TIP pathway was important for TOP1cc-activated, but not ionization radiation-activated ATM, p53 and Chk2 phosphorylation. We have also found that pharmacological interferences of TIP and RIP pathways distinctively modulated the CPT-induced cell killing with treatments at low and high dosages, respectively. Together, our results support that both RIP and TIP pathways of TOP1cc are required for the activation of CPT-induced DDR and cytotoxicity. © 2010 IBCB, SIBS, CAS. All rights reserved.

AB - Recent studies have suggested an involvement of processing pathways for the initiation of cellular responses induced by topoisomerase-targeting drugs. Here, we showed that cellular exposure to camptothecin (CPT) induced formation of topoisomerase I cleavable complex (TOP1cc), degradation of TOP1 and activation of DNA damage responses (DDR). Transcription and proteasome-dependent proteolysis, but not replication, were involved in CPT-induced TOP1 degradation, while none of above three processing activities affected TOP1cc formation. Replication-and transcription-initiated processing (RIP and TIP) of TOP1cc were identified as two independent pathways, which contribute distinctly to various CPT-activated DDR. Specifically, in cycling cells, RIP-processed TOP1cc triggered the CPT-induced RPA phosphorylation. At higher CPT dosages, the TIP pathway is required for other DDR activation, including ATM, p53 and Chk1/2 phosphorylation. The TIP pathway was further demonstrated to be S-phase independent by using three nonreplicating cell models. Furthermore, the effect of proteasome inhibitors mimicked that of transcription inhibition on the CPT-induced activation of DDR, suggesting the involvement of proteasome in the TIP pathway. Interestingly, the TIP pathway was important for TOP1cc-activated, but not ionization radiation-activated ATM, p53 and Chk2 phosphorylation. We have also found that pharmacological interferences of TIP and RIP pathways distinctively modulated the CPT-induced cell killing with treatments at low and high dosages, respectively. Together, our results support that both RIP and TIP pathways of TOP1cc are required for the activation of CPT-induced DDR and cytotoxicity. © 2010 IBCB, SIBS, CAS. All rights reserved.

KW - cleavable complex

KW - DNA damage responses

KW - downregulation

KW - processing

KW - protein-linked DNA break

KW - ATM protein

KW - camptothecin

KW - cell cycle protein

KW - checkpoint kinase 2

KW - DNA binding protein

KW - DNA topoisomerase

KW - protein p53

KW - protein serine threonine kinase

KW - replication factor A

KW - RPA2 protein, human

KW - tumor suppressor protein

KW - article

KW - DNA repair

KW - DNA replication

KW - DNA strand breakage

KW - genetic transcription

KW - genetics

KW - human

KW - metabolism

KW - phosphorylation

KW - signal transduction

KW - tumor cell line

KW - Camptothecin

KW - Cell Cycle Proteins

KW - Cell Line, Tumor

KW - DNA Breaks

KW - DNA Repair

KW - DNA Replication

KW - DNA Topoisomerases, Type I

KW - DNA-Binding Proteins

KW - Humans

KW - Phosphorylation

KW - Protein-Serine-Threonine Kinases

KW - Replication Protein A

KW - Signal Transduction

KW - Transcription, Genetic

KW - Tumor Suppressor Protein p53

KW - Tumor Suppressor Proteins

U2 - 10.1038/cr.2010.95

DO - 10.1038/cr.2010.95

M3 - Article

VL - 20

SP - 1060

EP - 1075

JO - Cell Research

JF - Cell Research

SN - 1001-0602

IS - 9

ER -