Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:5.99.1.2 (topoisomerase)
9,166 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Topoisomerase I-mediated DNA damage induced by camptothecin has been shown to induce rapid small ubiquitin-related modifier (SUMO)-1 conjugation to topoisomerase I. In the current study, we show that topoisomerase II-mediated DNA damage induced by teniposide (VM-26) results in the formation of high molecular weight conjugates of both topoisomerase IIalpha and IIbeta isozymes in HeLa cells. Immunological characterization of these conjugates suggests that both topoisomerase IIalpha and IIbeta isozymes are conjugated to SUMO-1. The involvement of SUMO-1/UBC9 in the modification of topoisomerase II isozymes is also supported by the demonstration of physical interaction between topoisomerase II and SUMO-1/UBC9. Surprisingly, ICRF-193, which does not induce topoisomerase II-mediated DNA damage but traps topoisomerase II into a circular clamp conformation, is also shown to induce similar SUMO-1 conjugation to topoisomerase II isozymes. In addition, we show that both oxidative and heat shock stresses, which can cause protein damage, rapidly increase nuclear SUMO-1 conjugates. These studies raise the question on whether SUMO-1 conjugation to topoisomerases is an indirect result of a DNA damage response or a direct result because of protein conformational changes.
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PMID:SUMO-1 conjugation to human DNA topoisomerase II isozymes. 1086 13

DNA topoisomerase I and II have been shown to be modified with a ubiquitin-like protein SUMO in response to their specific inhibitors called 'poisons'. These drugs also damage DNA by stabilizing the enzyme-DNA cleavable complex and induce a degradation of the enzymes through the 26S proteasome system. A plausible link between sumoylation and degradation has not yet been elucidated. We demonstrate here that topoisomerase IIbeta, but not its isoform IIalpha, is selectively degraded through proteasome by exposure to the catalytic inhibitor ICRF-193 which does not damage DNA. The beta isoform immunoprecipitated from ICRF-treated cells was modified by multiple modifiers, SUMO-2/3, SUMO-1, and polyubiquitin. When the SUMO conjugating enzyme Ubc9 was conditionally knocked out, the ICRF-induced degradation of topoisomerase IIbeta did not occur, suggesting that the SUMO modification pathway is essential for the degradation.
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PMID:The SUMO pathway is required for selective degradation of DNA topoisomerase IIbeta induced by a catalytic inhibitor ICRF-193(1). 1283 72

We have analyzed the abundance of SUMO-conjugated species during the cell cycle in Xenopus egg extracts. The predominant SUMO conjugation products associated with mitotic chromosomes arose from SUMO conjugation of topoisomerase II. Topoisomerase II was modified exclusively by SUMO-2/3 during mitosis under normal circumstances, although we observed conjugation of topoisomerase II to SUMO-1 in extracts with exogenous SUMO-1 protein. Inhibition of SUMO modification by a dominant-negative mutant of the SUMO-conjugating enzyme Ubc9 (dnUbc9) did not detectably alter topoisomerase II activity, but it did increase the amount of unmodified topoisomerase II retained on mitotic chromosomes after high salt washing. dnUbc9 did not disrupt the assembly of condensed mitotic chromosomes or block progression of extracts through mitosis, but it did block the dissociation of sister chromatids at the metaphase-anaphase transition. Together, our results suggest that SUMO conjugation is important for chromosome segregation in metazoan systems, and that mobilization of topoisomerase II from mitotic chromatin may be a key target of this modification.
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PMID:SUMO-2/3 regulates topoisomerase II in mitosis. 1459 74

The dynamic, posttranslational modification of proteins with a small ubiquitin-like modifier (SUMO) tag has been recognized as an important cellular regulatory mechanism relevant to a number of cancers as well as normal embryonic development. As part of a program aimed toward the identification of inhibitors of SUMO-conjugating enzymes, we developed a microfluidic electrophoretic mobility shift assay to monitor sumoylation events in real time. We disclose herein the use of this assay to identify a cell-permeable compound capable of blocking the transfer of SUMO-1 from the E2 enzyme Ubc9 to the substrate. We screened a small collection of compounds and identified an oxygenated flavonoid derivative that inhibits sumoylation in vitro. Next, we carried out an in-depth mechanistic analysis that ruled out many common false-positive mechanisms such as aggregation or alkylation. Furthermore, we report that this flavonoid inhibits a single step in the sumoylation cascade: the transfer of SUMO from the E2 enzyme (Ubc9) thioester conjugate to the substrate. In addition to having a unique mechanism of action, this inhibitor has a discrete structure-activity relationship uncharacteristic of a promiscuous inhibitor. Cell-based studies showed that the flavonoid inhibits the sumoylation of topoisomerase-I in response to camptothecin treatment in two different breast cancer cell lines, while isomeric analogs are inactive. Importantly, this compound blocks sumoylation while not affecting ubiquitylation in cells. This work identifies a point of entry for pharmacologic inhibition of the sumoylation cascade and may serve as the basis for continued study of additional pharmacophores that modulate SUMO-conjugating enzymes such as Ubc9.
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PMID:An electrophoretic mobility shift assay identifies a mechanistically unique inhibitor of protein sumoylation. 2360 49

Topoisomerases form transient covalent DNA cleavage complexes to perform their reactions. Topoisomerase I cleavage complexes (TOP1ccs) are trapped by camptothecin and TOP2ccs by etoposide. Proteolysis of the trapped topoisomerase DNA-protein cross-links (TOP-DPCs) is a key step for some pathways to repair these lesions. We describe a pathway that features a prominent role of the small ubiquitin-like modifier (SUMO) modification for both TOP1- and TOP2-DPC repair. Both undergo rapid and sequential SUMO-2/3 and SUMO-1 modifications in human cells. The SUMO ligase PIAS4 is required for these modifications. RNF4, a SUMO-targeted ubiquitin ligase (STUbL), then ubiquitylates the TOP-DPCs for their subsequent degradation by the proteasome. This pathway is conserved in yeast with Siz1 and Slx5-Slx8, the orthologs of human PIAS4 and RNF4.
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PMID:A conserved SUMO pathway repairs topoisomerase DNA-protein cross-links by engaging ubiquitin-mediated proteasomal degradation. 3318 14

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