Mechanisms to Repair Stalled Topoisomerase II-DNA Covalent Complexes

Abstract

DNA topoisomerases regulate the topological state of DNA, relaxing DNA supercoils and resolving catenanes and knots that result from biological processes such as transcription and replication. DNA topoisomerase II (TOP2) enzymes achieve this by binding DNA and introducing an enzyme-bridged DNA double-strand break (DSB) where each protomer of the dimeric enzyme is covalently attached to the 5' end of the cleaved DNA via an active site tyrosine phosphodiester linkage. The enzyme then passes a second DNA duplex though the DNA break, before religation and release of the enzyme. However, this activity is potentially hazardous to the cell, as failure to complete religation leads to persistent TOP2 protein-DNA covalent complexes which are cytotoxic. Indeed, this property of topoisomerase has been exploited in cancer therapy in the form of topoisomerase poisons which block the religation stage of the reaction cycle, leading to an accumulation of topoisomerase-DNA adducts. A number of parallel cellular processes have been identified that lead to removal of these covalent TOP2-DNA complexes facilitating repair of the resulting protein-free DSB by standard DNA repair pathways. These pathways presumably arose to repair spontaneous stalled or poisoned TOP2-DNA complexes, but understanding their mechanisms also has implications for cancer therapy, particularly resistance to anti-cancer TOP2 poisons and the genotoxic side effects of these drugs. Here we review recent progress in the understanding of the processing to TOP2 DNA covalent complexes., The basic components and mechanisms plus the additional layer of complexity posed by the post-translational modifications that modulate these pathways. SIGNIFICANCE STATEMENTMultiple pathways have been reported for removal and repair of TOP2-DNA covalent complexes to ensure the timely and efficient repair of TOP2-DNA covalent adducts to protect the genome. Post-translational modifications such as ubiquitination and SUMOylation are involved in the regulation of TOP2-DNA complex repair. Small molecule inhibitors of these post translational modifications may help to improve outcomes of TOP2 poison chemotherapy, for example by increasing TOP2 poison cytotoxicity and reducing genotoxicity, but this remains to be determined.

To Break or Not to Break: The Role of TOP2B in Transcription

“…Canonically, the transient enzyme-linked strand breaks formed during the reaction cycle remain buried within the enzyme as TOP2-covalent DNA complexes (TOP2-CCs) and are rapidly resolved after strand-passage (Figure 1A,B). However, if religation is impeded, TOP2-CCs can be processed to protein free DSBs (pf-DSBs) [5] which can initiate a DNA damage response including phosphorylation of histone H2AX (Figure 1C). This leads to the question of whether these promoter-associated, signalinduced, TOP2B-dependent DSBs are the incidental consequence of occasional failure of TOP2 to quickly complete its reaction cycle or are part of a mechanism that specifically requires the formation of distinct pf-DNA DSBs for efficient transcriptional induction (see Section 6).…”

“…In this article, we will elaborate on the specific roles of TOP2B in transcription focusing on the nature of the DSBs observed at activated promoters and on the reported protein interactions of TOP2B with established protein complexes and individual proteins associated with transcription-related processes. Figure 1C) [5]. Topoisomerases are involved in many types of DNA transaction, from replication and recombination to DNA repair and gene expression [6].…”

Deep learning-assisted high-content screening identifies isoliquiritigenin as an inhibitor of DNA double-strand breaks for preventing doxorubicin-induced cardiotoxicity

“…Anthracyclines bind to DNA and topoisomerase II (topo II), which hinders the re-ligation of the DNA double strands, and persistent DNA double-strand break (DSB) was generated consequently. DSB is lethal to cells and brings about p53 activation and cell death [ 13 ]. Zhang et al [ 14 ] found that the deletion of topo IIβ from cardiomyocytes prevented mice from developing AIC, suggesting the critical role of topoisomerase-mediated DSB in the pathomechanism of AIC.…”

Requirements for MRN endonuclease processing of topoisomerase II-mediated DNA damage in mammalian cells

“…Therefore, cells have a diverse repertoire of DNA repair and damage tolerance mechanisms to prevent topoisomerase-induced genome instability (Weickert and Stingele, 2022). These various pathways include DNA damage signaling, removal of proteins bound to DNA by proteolysis (Li and Liu, 2001), and a variety of nucleolytic enzymes that can excise phosphotyrosyl-linked peptides (Yang et al, 1996;Cortes Ledesma et al, 2009) or carry out endonucleolytic cleavage to remove DNA/protein crosslinks (Swan et al, 2022) MRE11 is a double strand break (DSB) repair enzyme with endonuclease and 3′ to 5′ exonuclease activities, acting in a functional protein complex with RAD50 and NBS1. The MRE11-RAD50-NBS1 (MRN) complex plays a central role in sensing and initial processing of DSB termini to promote HR or NHEJmediated repair (Paull, 2018).…”