A protease pathway for the repair of topoisomerase II-DNA covalent complexes.
Despite rapid advances in the field of DNA repair, little is known about the repair of protein-DNA adducts. Previous studies have demonstrated that topoisomerase II (TopII)-DNA adducts (TopII-DNA covalent complexes) are rapidly degraded by the proteasome. It has been hypothesized that proteasomal degradation of TopII-DNA covalent adducts exposes TopII-concealed DNA double-strand breaks (DSBs) for repair. To test this hypothesis, the anticancer drug, VP-16 (etoposide), was employed to induce TopII-DNA covalent complexes in mammalian cells, and the involvement of proteasome in processing TopII-DNA covalent complexes into DSBs was investigated. Consistent with the hypothesis, VP-16-induced DSBs as monitored by neutral comet assay, as well as DNA damage signals (e.g. gamma-H2AX) were significantly reduced in the presence of the proteasome inhibitor, MG132. Using both top2beta knock-out mouse embryonic fibroblasts and Top2beta small interfering RNA knockdown PC12 cells, as well as postmitotic neurons in which TopIIalpha was absent, we showed that VP-16-induced DNA damage signals were attenuated upon proteasome inhibition, suggesting the involvement of proteasome in the repair/processing of both TopIIalpha-DNA and TopIIbeta-DNA adducts. By contrast, hydrogen peroxide-induced gamma-H2AX was unaffected upon proteasome inhibition, suggesting a specific requirement of the proteasome pathway in the processing of TopII-DNA covalent complexes into DNA damage.
Zhang
Ailing
A
Department of Pharmacology, University of Medicine and Dentistry of New Jersey-Robert Wood Johnson Medical School, Piscataway, New Jersey 08854, USA.
Lyu
Yi Lisa
YL
Lin
Chao-Po
CP
Zhou
Nai
N
Azarova
Anna M
AM
Wood
Laurence M
LM
Liu
Leroy F
LF
D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology; D020552:Comet Assay; D004247:DNA / Q000737:chemistry*; D004250:DNA Topoisomerases, Type II / Q000737:chemistry*; D005347:Fibroblasts / Q000378:metabolism; D006657:Histones / Q000378:metabolism; D051379:Mice; D009474:Neurons / Q000378:metabolism; D016716:PC12 Cells; D010447:Peptide Hydrolases / Q000737:chemistry*; D011485:Protein Binding; D034741:RNA, Small Interfering / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley
D000970:Antineoplastic Agents; D006657:Histones; D034741:RNA, Small Interfering; D004247:DNA; D010447:Peptide Hydrolases; D004250:DNA Topoisomerases, Type II