Control of topological states of DNA by transient breakage and subsequent rejoining of DNA strands. Topoisomerase II makes double-strand breaks. Indirectly involved in vitamin D-coupled transcription regulation via its association with the WINAC complex, a chromatin-remodeling complex recruited by vitamin D receptor (VDR), which is required for the ligand-bound VDR-mediated transrepression of the CYP27B1 gene.
We identified a human multiprotein complex (WINAC) that directly interacts with the vitamin D receptor (VDR) through the Williams syndrome transcription factor (WSTF). WINAC has ATP-dependent chromatin-remodeling activity and contains both SWI/SNF components and DNA replication-related factors. The latter might explain a WINAC requirement for normal S phase progression. WINAC mediates the recruitment of unliganded VDR to VDR target sites in promoters, while subsequent binding of coregulators requires ligand binding. This recruitment order exemplifies that an interaction of a sequence-specific regulator with a chromatin-remodeling complex can organize nucleosomal arrays at specific local sites in order to make promoters accessible for coregulators. Furthermore, overexpression of WSTF could restore the impaired recruitment of VDR to vitamin D regulated promoters in fibroblasts from Williams syndrome patients. This suggests that WINAC dysfunction contributes to Williams syndrome, which could therefore be considered, at least in part, a chromatin-remodeling factor disease.
A type II topoisomerase is essential for decatenating DNA replication products, and it accomplishes this task by passing one DNA duplex through a transient break in a second duplex. The B' domain of topoisomerase II contains three highly conserved motifs, EGDSA, PL(R/K)GK(I/L/M)LNVR, and IMTD(Q/A)DXD. We have investigated these motifs in topoisomerase II beta by mutagenesis, and report that they play a critical role in establishing the DNA cleavage-religation equilibrium. In addition, the mutations E477Q (EGDSA) and K505E (PLRGKILNVR) increase the optimal magnesium ion concentration for strand passage, without affecting the Mg(2+) dependence of ATP hydrolysis. It is likely that the binding affinity of the magnesium ion(s) specifically required for DNA cleavage has been reduced by these mutations. The crystal structure of yeast topo II indicates that residues E477 and K505 may help to position the three aspartate residues of the IMTD(Q/A)DXD motif for magnesium ion coordination, and we propose two possible locations for the magnesium ion binding site(s). These observations are consistent with a previous model in which the B' domain is positioned such that these acidic residues lie next to the active site tyrosine residue. A magnesium ion bound by these aspartate residues could therefore mediate the DNA cleavage-religation reaction.
Interacting selectively and non-covalently with chromatin, the network of fibers of DNA, protein, and sometimes RNA, that make up the chromosomes of the eukaryotic nucleus during interphase.
We visualized DNA topoisomerases in A431 cells and isolated chromosomes by isoenzyme-selective immunofluorescence microscopy. In interphase, topoisomerase I mainly had a homogeneous nuclear distribution. 10-15% of the cells exhibited granular patterns, 30% showed bright intranucleolar patches. Topoisomerase II isoenzymes showed spotted (alpha) or reticular (beta) nuclear patterns throughout interphase. In contrast to topoisomerase IIalpha, topoisomerase IIbeta was completely excluded from nucleoli. In mitosis, topoisomerase IIbeta diffused completely into the cytosol, whereas topoisomerases I and IIalpha remained chromosome bound. Chromosomal staining of topoisomerase I was homogeneous, whereas topoisomerase IIalpha accumulated in the long axes of the chromosome arms and in the centriols. Topoisomerase antigens were 2-3-fold higher in mitosis than in interphase, but specific activities of topoisomerase I and II were reduced 5- and 2.4-fold, respectively. These changes were associated with mitotic enzyme hyperphosphorylation. In interphase, topoisomerases could be completely linked to DNA by etoposide or camptothecin, whereas in mitosis, 50% of topoisomerase IIalpha escaped poisoning. Refractoriness to etoposide could be assigned to the salt-stable scaffold fraction of topoisomerase IIalpha, which increased from <2% in G1 phase to 48% in mitosis. Topoisomerases I and IIbeta remained completely extractable throughout the cell cycle. In summary, expression of topoisomerases increases towards mitosis, but specific activities decrease. Topoisomerase IIbeta is released from the heterochromatin, whereas topoisomerase I and IIalpha remain chromosome bound. Scaffold-associated topoisomerase IIalpha appears not to be involved in catalytic DNA turnover, though it may play a role in the replicational cycle of centriols, where it accumulates during M phase.
Catalysis of a DNA topological transformation by transiently cleaving a pair of complementary DNA strands to form a gate through which a second double-stranded DNA segment is passed, after which the severed strands in the first DNA segment are rejoined; product release is coupled to ATP binding and hydrolysis; changes the linking number in multiples of 2.
A type II topoisomerase is essential for decatenating DNA replication products, and it accomplishes this task by passing one DNA duplex through a transient break in a second duplex. The B' domain of topoisomerase II contains three highly conserved motifs, EGDSA, PL(R/K)GK(I/L/M)LNVR, and IMTD(Q/A)DXD. We have investigated these motifs in topoisomerase II beta by mutagenesis, and report that they play a critical role in establishing the DNA cleavage-religation equilibrium. In addition, the mutations E477Q (EGDSA) and K505E (PLRGKILNVR) increase the optimal magnesium ion concentration for strand passage, without affecting the Mg(2+) dependence of ATP hydrolysis. It is likely that the binding affinity of the magnesium ion(s) specifically required for DNA cleavage has been reduced by these mutations. The crystal structure of yeast topo II indicates that residues E477 and K505 may help to position the three aspartate residues of the IMTD(Q/A)DXD motif for magnesium ion coordination, and we propose two possible locations for the magnesium ion binding site(s). These observations are consistent with a previous model in which the B' domain is positioned such that these acidic residues lie next to the active site tyrosine residue. A magnesium ion bound by these aspartate residues could therefore mediate the DNA cleavage-religation reaction.
DNA topoisomerase (topo) II modulates DNA topology and is essential for cell division. There are two isoforms of topo II (alpha and beta) that have limited functional redundancy, although their catalytic mechanisms appear the same. Using their COOH-terminal domains (CTDs) in yeast two-hybrid analysis, we have identified phospholipid scramblase 1 (PLSCR1) as a binding partner of both topo II alpha and beta. Although predominantly a plasma membrane protein involved in phosphatidylserine externalization, PLSCR1 can also be imported into the nucleus where it may have a tumour suppressor function. The interactions of PLSCR1 and topo II were confirmed by pull-down assays with topo II alpha and beta CTD fusion proteins and endogenous PLSCR1, and by co-immunoprecipitation of endogenous PLSCR1 and topo II alpha and beta from HeLa cell nuclear extracts. PLSCR1 also increased the decatenation activity of human topo IIalpha. A conserved basic sequence in the CTD of topo IIalpha was identified as being essential for binding to PLSCR1 and binding of the two proteins could be inhibited by a synthetic peptide corresponding to topo IIalpha amino acids 1430-1441. These studies reveal for the first time a physical and functional interaction between topo II and PLSCR1.
Histone deacetylases (HDACs) modify nucleosomal histones, have a key role in the regulation of gene transcription, and may be involved in cell-cycle regulation, differentiation and human cancer. Purified recombinant human HDAC1 protein was used to screen a cDNA expression library, and one of the clones identified encoded DNA topoisomerase II (Topo II), an enzyme known to have a role in transcriptional regulation and chromatin organization. Coimmunoprecipitation experiments indicate that HDAC1 and HDAC2 are associated with Topo II in vivo under normal physiological conditions. Complexes containing Topo II possess HDAC activities, and complexes containing HDAC1 or HDAC2 possess Topo II activities. HDAC and Topo II modify each other's activity in vitro and in vivo. Our results indicate the existence of a functionally coupled complex between these two enzymes and offer insights into the potential mechanisms of action of both enzymes.
Evidence
2:
Inferred from Physical InteractionUniProtKB
DNA topoisomerase II (topo II) is a ubiquitous nuclear enzyme that is involved in DNA replication, transcription, chromosome segregation, and apoptosis. Here we show by immunoprecipitation, pull down with glutathione S-transferase fusion proteins, and yeast two-hybrid analysis that both topo IIalpha and -beta physically interact with the histone deacetylase HDAC1. The in vitro DNA decatenation activity of recombinant topo IIalpha and -beta is inhibited by association with catalytically inactive, recombinant HDAC1. We provide evidence for the in vivo significance of the topo II-HDAC1 association, showing that inhibition of HDAC activity with trichostatin A suppresses apoptosis induced by the topo II poison etoposide, but not by the topoisomerase I inhibitor camptothecin. We suggest that chromatin remodeling by an HDAC-containing complex facilitates both topo II-catalyzed DNA rearrangement and etoposide-induced DNA damage in vivo.
Interacting selectively and non-covalently with a protein C-terminus, the end of any peptide chain at which the 1-carboxy function of a constituent amino acid is not attached in peptide linkage to another amino-acid residue.
Evidence
1:
Inferred from Physical InteractionUniProtKB
The p53 tumor suppressor protein is a critical regulator of cell cycle progression and apoptosis following exposure of cells to DNA damaging agents such as ionizing radiation or anticancer drugs. An important group of anticancer drugs, including compounds such as etoposide and doxorubicin (Adriamycin), interacts with DNA topoisomerase II (topo II), causing the accumulation of enzyme-DNA adducts that ultimately lead to double-strand breaks and cell death via apoptosis. Human topo IIbeta has previously been shown to interact with p53, and we have extended this analysis to show that both topo IIalpha and IIbeta interact with p53 in vivo and in vitro. Furthermore, we show that the regulatory C-terminal basic region of p53 (residues 364-393) is necessary and sufficient for interaction with DNA topo II.
J. Biol. Chem. 274, 26529-26536 (1999)[PubMed:10473615]
Eukaryotic topoisomerase II is a nuclear enzyme essential for DNA metabolism and chromosome dynamics. The enzyme has a dimeric structure, and subunit dimerization is vital to the cellular functions and activities of the enzyme. Two biochemical approaches based on metal ion affinity chromatography and immunoprecipitation have been carried out to map the dimerization region(s) in human topoisomerase IIalpha. The results demonstrate that two regions spanning amino acids 1053-1069 and 1124-1143 are both essential for dimerization. The regions correspond to the interaction domains revealed in yeast topoisomerase II after crystallization of a central fragment of this enzyme, indicating that the overall C-terminal dimerization structure of eukaryotic topoisomerase II is conserved from yeast to human. Furthermore, linker insertion analysis has demonstrated that the two dimerization regions are located in a highly flexible part of the enzyme. Topoisomerase IIalpha mutant enzymes unable to dimerize via the C-terminal primary dimerization regions due to lack of one of the defined dimerization regions can still be forced to dimerize if DNA and an ATP analog are added to the reaction mixture. The result indicates that secondary interactions occur by ATP analog-mediated clamp closing when the subunits are brought together on DNA.
DNA topoisomerase II is an essential nuclear enzyme that modulates DNA processes by altering the topological state of double-stranded DNA. This enzyme is required for chromosome condensation and segregation; however, the regulatory mechanism of its activation is largely unknown. Here we demonstrate that topoisomerase IIalpha is activated in response to genotoxic stress. Concomitant with the activation, the expression of topoisomerase IIalpha is increased following DNA damage. The results also demonstrate that the proapoptotic kinase protein kinase C delta (PKCdelta) interacts with topoisomerase IIalpha. This association is in an S-phase-specific manner and is required for stabilization and catalytic activation of topoisomerase IIalpha in response to DNA damage. Conversely, inhibition of PKCdelta activity attenuates DNA damage-induced activation of topoisomerase IIalpha. Finally, aberrant activation of topoisomerase IIalpha by PKCdelta is associated with induction of apoptosis upon exposure to genotoxic agents. These findings indicate that PKCdelta regulates topoisomerase IIalpha and thereby cell fate in the genotoxic stress response.
The process in which genetic material, in the form of chromosomes, is organized into specific structures and then physically separated and apportioned to two or more sets. In eukaryotes, chromosome segregation begins with the alignment of chromosomes at the metaphase plate, includes chromosome separation, and ends when chromosomes have completed movement to the spindle poles.
A type II topoisomerase is essential for decatenating DNA replication products, and it accomplishes this task by passing one DNA duplex through a transient break in a second duplex. The B' domain of topoisomerase II contains three highly conserved motifs, EGDSA, PL(R/K)GK(I/L/M)LNVR, and IMTD(Q/A)DXD. We have investigated these motifs in topoisomerase II beta by mutagenesis, and report that they play a critical role in establishing the DNA cleavage-religation equilibrium. In addition, the mutations E477Q (EGDSA) and K505E (PLRGKILNVR) increase the optimal magnesium ion concentration for strand passage, without affecting the Mg(2+) dependence of ATP hydrolysis. It is likely that the binding affinity of the magnesium ion(s) specifically required for DNA cleavage has been reduced by these mutations. The crystal structure of yeast topo II indicates that residues E477 and K505 may help to position the three aspartate residues of the IMTD(Q/A)DXD motif for magnesium ion coordination, and we propose two possible locations for the magnesium ion binding site(s). These observations are consistent with a previous model in which the B' domain is positioned such that these acidic residues lie next to the active site tyrosine residue. A magnesium ion bound by these aspartate residues could therefore mediate the DNA cleavage-religation reaction.
The process whose specific outcome is the progression of the forebrain over time, from its formation to the mature structure. The forebrain is the anterior of the three primary divisions of the developing chordate brain or the corresponding part of the adult brain (in vertebrates, includes especially the cerebral hemispheres, the thalamus, and the hypothalamus and especially in higher vertebrates is the main control center for sensory and associative information processing, visceral functions, and voluntary motor functions).
A mitotic cell cycle checkpoint that blocks cell cycle progression from G2 to M phase until sister chromatids are sufficiently separated, with concatenated molecules topologically resolved.
The exchange, reciprocal or nonreciprocal, of genetic material between one DNA molecule and a homologous region of DNA that occurs during mitotic cell cycles.
The cleavage and rejoining of intermediates, such as Holliday junctions, formed during meiotic recombination to produce two intact molecules in which genetic material has been exchanged.
A type II topoisomerase is essential for decatenating DNA replication products, and it accomplishes this task by passing one DNA duplex through a transient break in a second duplex. The B' domain of topoisomerase II contains three highly conserved motifs, EGDSA, PL(R/K)GK(I/L/M)LNVR, and IMTD(Q/A)DXD. We have investigated these motifs in topoisomerase II beta by mutagenesis, and report that they play a critical role in establishing the DNA cleavage-religation equilibrium. In addition, the mutations E477Q (EGDSA) and K505E (PLRGKILNVR) increase the optimal magnesium ion concentration for strand passage, without affecting the Mg(2+) dependence of ATP hydrolysis. It is likely that the binding affinity of the magnesium ion(s) specifically required for DNA cleavage has been reduced by these mutations. The crystal structure of yeast topo II indicates that residues E477 and K505 may help to position the three aspartate residues of the IMTD(Q/A)DXD motif for magnesium ion coordination, and we propose two possible locations for the magnesium ion binding site(s). These observations are consistent with a previous model in which the B' domain is positioned such that these acidic residues lie next to the active site tyrosine residue. A magnesium ion bound by these aspartate residues could therefore mediate the DNA cleavage-religation reaction.
Magnesium. Binds two Mg(2+) per subunit. The magnesium ions form salt bridges with both the protein and the DNA. Can also accept other divalent metal cations, such an Mn(2+) and Ca(2+).
A type II topoisomerase is essential for decatenating DNA replication products, and it accomplishes this task by passing one DNA duplex through a transient break in a second duplex. The B' domain of topoisomerase II contains three highly conserved motifs, EGDSA, PL(R/K)GK(I/L/M)LNVR, and IMTD(Q/A)DXD. We have investigated these motifs in topoisomerase II beta by mutagenesis, and report that they play a critical role in establishing the DNA cleavage-religation equilibrium. In addition, the mutations E477Q (EGDSA) and K505E (PLRGKILNVR) increase the optimal magnesium ion concentration for strand passage, without affecting the Mg(2+) dependence of ATP hydrolysis. It is likely that the binding affinity of the magnesium ion(s) specifically required for DNA cleavage has been reduced by these mutations. The crystal structure of yeast topo II indicates that residues E477 and K505 may help to position the three aspartate residues of the IMTD(Q/A)DXD motif for magnesium ion coordination, and we propose two possible locations for the magnesium ion binding site(s). These observations are consistent with a previous model in which the B' domain is positioned such that these acidic residues lie next to the active site tyrosine residue. A magnesium ion bound by these aspartate residues could therefore mediate the DNA cleavage-religation reaction.
Type II topoisomerases (TOP2s) resolve the topological problems of DNA by transiently cleaving both strands of a DNA duplex to form a cleavage complex through which another DNA segment can be transported. Several widely prescribed anticancer drugs increase the population of TOP2 cleavage complex, which leads to TOP2-mediated chromosome DNA breakage and death of cancer cells. We present the crystal structure of a large fragment of human TOP2β complexed to DNA and to the anticancer drug etoposide to reveal structural details of drug-induced stabilization of a cleavage complex. The interplay between the protein, the DNA, and the drug explains the structure-activity relations of etoposide derivatives and the molecular basis of drug-resistant mutations. The analysis of protein-drug interactions provides information applicable for developing an isoform-specific TOP2-targeting strategy.
Enzyme that catalyzes the 1,1-, 1,2- or 1,3-hydrogen shift. The 1,1- hydrogen shift is an inversion at an asymmetric carbon center (racemases, epimerases). The 1,2-hydrogen shift involved a hydrogen transfer between two adjacent carbon atoms, one undergoing oxidation, the other reduction (aldose-ketose isomerases). The 1,3-hydrogen shifts are allylic or azaallylic (when nitrogen is one of the three atoms) isomerizations.
Enzymes capable of altering the degree of supercoiling of double- stranded DNA molecules. Various topoisomerases can increase or relax supercoiling, convert single-stranded rings to intertwined double- stranded rings, tie and untie knots in single stranded and duplex rings or catenate and decatenate duplex rings. Any enzyme that cleaves only one strand of a DNA duplex and then reseals it is classified as a type I topoisomerase (Topo I). Type II topoisomerases (Topo II) change DNA topology by breaking and rejoining double-stranded DNA.
A reference proteome is a set of protein sequences derived from a complete proteome which constitutes a defined standard for a particular user community. Reference proteomes are manually defined according to a number of criteria. They cover the proteomes of well- studied model organisms and other proteomes of interest for biomedical and biotechnological research. Reference proteomes have been selected to provide broad coverage of the tree of life, and constitute a representative cross-section of the taxonomic diversity to be found within UniProtKB.
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