Hartung Frank, Suer Stefanie, Knoll Alexander, Wurz-Wildersinn Rebecca, Puchta Holger
Botany II, University of Karlsruhe, Karlsruhe, Germany.
PLoS Genet. 2008 Dec;4(12):e1000285. doi: 10.1371/journal.pgen.1000285. Epub 2008 Dec 19.
Topoisomerases are enzymes with crucial functions in DNA metabolism. They are ubiquitously present in prokaryotes and eukaryotes and modify the steady-state level of DNA supercoiling. Biochemical analyses indicate that Topoisomerase 3alpha (TOP3alpha) functions together with a RecQ DNA helicase and a third partner, RMI1/BLAP75, in the resolution step of homologous recombination in a process called Holliday Junction dissolution in eukaryotes. Apart from that, little is known about the role of TOP3alpha in higher eukaryotes, as knockout mutants show early lethality or strong developmental defects. Using a hypomorphic insertion mutant of Arabidopsis thaliana (top3alpha-2), which is viable but completely sterile, we were able to define three different functions of the protein in mitosis and meiosis. The top3alpha-2 line exhibits fragmented chromosomes during mitosis and sensitivity to camptothecin, suggesting an important role in chromosome segregation partly overlapping with that of type IB topoisomerases. Furthermore, AtTOP3alpha, together with AtRECQ4A and AtRMI1, is involved in the suppression of crossover recombination in somatic cells as well as DNA repair in both mammals and A. thaliana. Surprisingly, AtTOP3alpha is also essential for meiosis. The phenotype of chromosome fragmentation, bridges, and telophase I arrest can be suppressed by AtSPO11 and AtRAD51 mutations, indicating that the protein is required for the resolution of recombination intermediates. As Atrmi1 mutants have a similar meiotic phenotype to Attop3alpha mutants, both proteins seem to be involved in a mechanism safeguarding the entangling of homologous chromosomes during meiosis. The requirement of AtTOP3alpha and AtRMI1 in a late step of meiotic recombination strongly hints at the possibility that the dissolution of double Holliday Junctions via a hemicatenane intermediate is indeed an indispensable step of meiotic recombination.
拓扑异构酶是在DNA代谢中具有关键功能的酶。它们普遍存在于原核生物和真核生物中，并调节DNA超螺旋的稳态水平。生化分析表明，拓扑异构酶3α（TOP3α）在真核生物中同源重组的解离步骤（称为霍利迪连接解离）中与RecQ DNA解旋酶及第三个伙伴RMI1/BLAP75共同发挥作用。除此之外，关于TOP3α在高等真核生物中的作用知之甚少，因为基因敲除突变体表现出早期致死性或严重的发育缺陷。利用拟南芥的一个亚效插入突变体（top3α-2），该突变体是可存活的但完全不育，我们能够确定该蛋白在有丝分裂和减数分裂中的三种不同功能。top3α-2株系在有丝分裂期间表现出染色体片段化以及对喜树碱敏感，这表明其在染色体分离中起重要作用，部分与IB型拓扑异构酶的作用重叠。此外，AtTOP3α与AtRECQ4A和AtRMI1一起参与体细胞中交叉重组的抑制以及哺乳动物和拟南芥中的DNA修复。令人惊讶的是，AtTOP3α对减数分裂也是必不可少的。AtSPO11和AtRAD51突变可抑制染色体片段化、桥接和减数第一次分裂末期停滞的表型，这表明该蛋白是重组中间体解离所必需的。由于Atrmi1突变体具有与Attop3α突变体相似的减数分裂表型，这两种蛋白似乎都参与了一种在减数分裂期间保护同源染色体缠结的机制。减数分裂重组后期对AtTOP3α和AtRMI1的需求强烈暗示，通过半连环中间体解离双霍利迪连接确实是减数分裂重组中不可或缺的一步。
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