Yeast Gene for a Tyr-DNA Phosphodiesterase that Repairs Topoisomerase I Complexes

Pouliot, Jeffrey J.; Yao, Kevin C.; Robertson, Carol A.; Nash, Howard A.

doi:10.1126/science.286.5439.552

Cited by 358 publications

(342 citation statements)

References 17 publications

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“…11), the fact that ED expression does not enhance sensitivity to camptothecin exposure suggests that APE1 is not a major player in the removal of this form of DNA damage. Indeed, evidence indicates that tyrosyl DNA phosphodiesterase serves as the primary repair enzyme for such 3 ¶-blocking lesions in mammalian cells (49,50). The observation that ED induction does not alter cellular sensitivity to etoposide, mitomycin C, and cisplatin is consistent with APE1 not playing a known role in the repair of 5 ¶-trapped topoisomerase-DNA intermediates or interstrand/intrastrand DNA cross-links.…”

Section: Discussionsupporting

confidence: 56%

A Dominant-Negative Form of the Major Human Abasic Endonuclease Enhances Cellular Sensitivity to Laboratory and Clinical DNA-Damaging Agents

McNeill

Wilson

2007

Molecular Cancer Research

109

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Apurinic/apyrimidinic (AP) endonuclease 1 (APE1) is the primary enzyme in mammals for the repair of abasic sites in DNA, as well as a variety of 3 ¶ damages that arise upon oxidation or as products of enzymatic processing. If left unrepaired, APE1 substrates can promote mutagenic and cytotoxic outcomes. We describe herein a dominant-negative form of APE1 that lacks detectable nuclease activity and binds substrate DNA with a 13-fold higher affinity than the wild-type protein. This mutant form of APE1, termed ED, possesses two amino acid substitutions at active site residues Glu 96 (changed to Gln) and Asp 210 (changed to Asn). In vitro biochemical assays reveal that ED impedes wild-type APE1 AP site incision function, presumably by binding AP-DNA and blocking normal lesion processing. Moreover, tetracycline-regulated (tet-on) expression of ED in Chinese hamster ovary cells enhances the cytotoxic effects of the laboratory DNA-damaging agents, methyl methanesulfonate (MMS; 5.4-fold) and hydrogen peroxide (1.5-fold). This MMS-induced, ED-dependent cell killing coincides with a hyperaccumulation of AP sites, implying that excessive DNA damage is the cause of cell death. Because an objective of the study was to identify a protein reagent that could be used in targeted gene therapy protocols, the effects of ED on cellular sensitivity to a number of chemotherapeutic compounds was tested. We show herein that ED expression sensitizes Chinese hamster ovary cells to the killing effects of the alkylating agent 1,3-bis(2-chloroethyl)-1-nitrosourea (also known as carmustine) and the chain terminating nucleoside analogue dideoxycytidine (also known as zalcitabine), but not to the radiomimetic bleomycin, the nucleoside analogue B-D-arabinofuranosylcytosine (also known as cytarabine), the topoisomerase inhibitors camptothecin and etoposide, or the cross-linking agents mitomycin C and cisplatin. Transient expression of ED in the human cancer cell line NCI-H1299 enhanced cellular sensitivity to MMS, 1,3-bis(2-chloroethyl)-1-nitrosourea, and dideoxycytidine, demonstrating the potential usefulness of this strategy in the treatment of human tumors.

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Section: Discussionsupporting

confidence: 56%

A Dominant-Negative Form of the Major Human Abasic Endonuclease Enhances Cellular Sensitivity to Laboratory and Clinical DNA-Damaging Agents

McNeill

Wilson

2007

Molecular Cancer Research

109

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“…Because Top1 is the only known enzyme to generate 3′-phosphotyrosine linkages in vivo, the observed enzymatic activity was proposed to be associated with the repair of DNA lesions, which develop from irreversible Top1-DNA cleavage complexes [19,20]. The yeast gene encoding this activity was subsequently cloned and appropriately named tyrosyl-DNA phosphodiesterase 1 (Tdp1).…”

Section: Biological Functions and Catalytic Mechanismmentioning

confidence: 99%

Tyrosyl-DNA Phosphodiesterase as a Target for Anticancer Therapy

Dexheimer¹,

Antony²,

Marchand³

et al. 2008

ACAMC

145

203

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Tyrosyl-DNA phosphodiesterase 1 (Tdp1) is a recently discovered enzyme that catalyzes the hydrolysis of 3′-phosphotyrosyl bonds. Such linkages form in vivo following the DNA processing activity of topoisomerase I (Top1). For this reason, Tdp1 has been implicated in the repair of irreversible Top1-DNA covalent complexes, which can be generated by either exogenous or endogenous factors. Tdp1 has been regarded as a potential therapeutic co-target of Top1 in that it seemingly counteracts the effects of Top1 inhibitors, such as camptothecin and its clinically used derivatives. Thus, by reducing the repair of Top1-DNA lesions, Tdp1 inhibitors have the potential to augment the anticancer activity of Top1 inhibitors provided there is a presence of genetic abnormalities related to DNA checkpoint and repair pathways. Human Tdp1 can also hydrolyze other 3′-end DNA alterations including 3′-phosphoglycolates and 3′-abasic sites indicating it may function as a general 3′-DNA phosphodiesterase and repair enzyme. The importance of Tdp1 in humans is highlighted by the observation that a recessive mutation in the human TDP1 gene is responsible for the inherited disorder, spinocerebellar ataxia with axonal neuropathy (SCAN1). This review provides a summary of the biochemical and cellular processes performed by Tdp1 as well as the rationale behind the development of Tdp1 inhibitors for anticancer therapy.

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“…TOP1 can become covalently attached to the DNA, forming TOP1 cleavage complexes (TOP1ccs) [36], and many types of endogenous DNA modifications can cause TOP1ccs [37]. The observation that spinocerebellar ataxia with axonal neuropathy (SCAN1) results from mutations in the TDP1 gene [22], which encodes a protein involved in the repair of TOP1ccs [38,39] indicates the importance of repair of such complexes in preventing ataxia. In view of the high levels of TOP1 in Purkinje neurons, it follows that these cells would be at elevated risk for TOP1ccs, and thus the loss of ability to repair such complexes would be particularly severe in these cells.…”

Section: The Mrn-complex and Topoisomerase I Are Also Concentrated Inmentioning

confidence: 99%

ATM, the Mre11/Rad50/Nbs1 complex, and topoisomerase I are concentrated in the nucleus of Purkinje neurons in the juvenile human brain

et al. 2007

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The genetic disease ataxia telangiectasia (AT) results from mutations in the ataxia telangiectasia mutated (ATM) gene. AT patients develop a progressive degeneration of cerebellar Purkinje neurons. Surprisingly, while ATM plays a criticial role in the cellular reponse to DNA damage, previous studies have localized ATM to the cytoplasm of rodent and human Purkinje neurons. Here we show that ATM is primarily localized to the nucleus in cerebellar Purkinje neurons in postmortem human brain tissue samples, although some light cytoplasmic ATM staining was also observed. No ATM staining was observed in brain tissue samples from AT patients, verifying the specificity of the antibody. We also found that antibodies against components of the Mre11/Rad50/Nbs1 (MRN) complex showed strong staining in Purkinje cell nuclei. However, while ATM is present in both the nucleoplasm and nucleolus, MRN proteins are excluded from the nucleolus. We also observed very high levels of topoisomerase 1 (TOP1) in the nucleus, and specifically the nucleolus, of human Purkinje neurons. Our results have direct implications for understanding the mechanisms of neurodegeneration in AT and AT-like disorder.

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Yeast Gene for a Tyr-DNA Phosphodiesterase that Repairs Topoisomerase I Complexes

Cited by 358 publications

References 17 publications

A Dominant-Negative Form of the Major Human Abasic Endonuclease Enhances Cellular Sensitivity to Laboratory and Clinical DNA-Damaging Agents

A Dominant-Negative Form of the Major Human Abasic Endonuclease Enhances Cellular Sensitivity to Laboratory and Clinical DNA-Damaging Agents

Tyrosyl-DNA Phosphodiesterase as a Target for Anticancer Therapy

ATM, the Mre11/Rad50/Nbs1 complex, and topoisomerase I are concentrated in the nucleus of Purkinje neurons in the juvenile human brain

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