TY - JOUR
T1 - Crosstalk between repair pathways elicits double-strand breaks in alkylated dna and implications for the action of temozolomide
AU - Fuchs, Robert P.
AU - Isogawa, Asako
AU - Paulo, Joao A.
AU - Onizuka, Kazumitsu
AU - Takahashi, Tatsuro
AU - Amunugama, Ravindra
AU - Duxin, Julien P.
AU - Fujii, Shingo
N1 - Funding Information:
We thank Johannes Walter (Harvard Medical School) for providing space, advice, and materials, and Paul Modrich (Duke Univ) and Bernd Kaina (Medical Univ, Mainz) for insightful reading and suggestions. This work was partially supported by NIH/NIGMS grant R01 GM132129 (to JAP) and MEXT/JSPS KAKENHI JP20H03186 and JP20H05392 (to TT).
Publisher Copyright:
© Fuchs et al.
PY - 2021/7
Y1 - 2021/7
N2 - Temozolomide (TMZ), a DNA methylating agent, is the primary chemotherapeutic drug used in glioblastoma treatment. TMZ induces mostly N-alkylation adducts (N7-methylguanine and N3-methyladenine) and some O6-methylguanine (O6 mG) adducts. Current models propose that during DNA replication, thymine is incorporated across from O6 mG, promoting a futile cycle of mismatch repair (MMR) that leads to DNA double-strand breaks (DSBs). To revisit the mechanism of O6 mG processing, we reacted plasmid DNA with N-methyl-N-nitrosourea (MNU), a temozolomide mimic, and incubated it in Xenopus egg-derived extracts. We have shown that in this system, MMR proteins are enriched on MNU-treated DNA and we observed robust, MMR-dependent, repair synthesis. Our evidence also suggests that MMR, initiated at O6 mG:C sites, is strongly stimulated in cis by repair processing of other lesions, such as N-alkylation adducts. Importantly, MNU-treated plasmids display DSBs in extracts, the frequency of which increases linearly with the square of alkylation dose. We suggest that DSBs result from two independent repair processes, one involving MMR at O6 mG:C sites and the other involving base excision repair acting at a nearby N-alkylation adduct. We propose a new, replication-independent mechanism of action of TMZ, which operates in addition to the well-studied cell cycle-dependent mode of action.
AB - Temozolomide (TMZ), a DNA methylating agent, is the primary chemotherapeutic drug used in glioblastoma treatment. TMZ induces mostly N-alkylation adducts (N7-methylguanine and N3-methyladenine) and some O6-methylguanine (O6 mG) adducts. Current models propose that during DNA replication, thymine is incorporated across from O6 mG, promoting a futile cycle of mismatch repair (MMR) that leads to DNA double-strand breaks (DSBs). To revisit the mechanism of O6 mG processing, we reacted plasmid DNA with N-methyl-N-nitrosourea (MNU), a temozolomide mimic, and incubated it in Xenopus egg-derived extracts. We have shown that in this system, MMR proteins are enriched on MNU-treated DNA and we observed robust, MMR-dependent, repair synthesis. Our evidence also suggests that MMR, initiated at O6 mG:C sites, is strongly stimulated in cis by repair processing of other lesions, such as N-alkylation adducts. Importantly, MNU-treated plasmids display DSBs in extracts, the frequency of which increases linearly with the square of alkylation dose. We suggest that DSBs result from two independent repair processes, one involving MMR at O6 mG:C sites and the other involving base excision repair acting at a nearby N-alkylation adduct. We propose a new, replication-independent mechanism of action of TMZ, which operates in addition to the well-studied cell cycle-dependent mode of action.
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U2 - 10.7554/eLife.69544
DO - 10.7554/eLife.69544
M3 - Article
C2 - 34236314
AN - SCOPUS:85111555353
VL - 10
JO - eLife
JF - eLife
SN - 2050-084X
M1 - e69544
ER -