Mokhtari Parisa D, Seyedjavadi Tannos, Liyanaarachchi Thulni A, Courcelle Charmain T, Courcelle Justin
Dept of Biology, Portland State University, Portland, Oregon, United States of America.
PLoS Genet. 2025 Sep 8;21(9):e1011857. doi: 10.1371/journal.pgen.1011857. eCollection 2025 Sep.
The ability to complete DNA replication as replisomes converge has recently been shown to be a highly regulated, multi-enzymatic process. Converging forks also are likely to generate unique supercoiled, tangled, or knotted substrates. These structures are typically resolved by one of the four topoisomerases encoded by Escherichia coli. However, identifying the cellular substrates and specific function for these essential enzymes which contain overlapping biochemical activities has remained challenging. Here, we show that Topo I and Topo IV are required to allow converging forks to complete chromosome replication. Impaired Topo I function leads to amplifications where forks converge, whereas inactivation of Topo IV prevents forks from converging and produces a dramatic loss of this chromosome region. The results are consistent with previous studies suggesting Topo I suppresses illegitimate initiations in the terminus region by disrupting R- and D-loops and demonstrate a specific requirement for Topo IV acting before replication completes to allow convergent forks to reach their doubling point. We propose that the positive supercoils arising between convergent forks are converted to precatenanes and resolved by Topo IV, when diminishing space may preclude gyrase from binding and functioning.
最近研究表明，随着复制体的汇聚完成DNA复制的能力是一个受到高度调控的多酶过程。汇聚的复制叉也可能产生独特的超螺旋、缠结或打结的底物。这些结构通常由大肠杆菌编码的四种拓扑异构酶之一来解决。然而，确定这些具有重叠生化活性的必需酶的细胞底物和特定功能仍然具有挑战性。在这里，我们表明拓扑异构酶I（Topo I）和拓扑异构酶IV（Topo IV）是使汇聚的复制叉完成染色体复制所必需的。拓扑异构酶I功能受损会导致复制叉汇聚处发生扩增，而拓扑异构酶IV失活会阻止复制叉汇聚，并导致该染色体区域的显著丢失。这些结果与之前的研究一致，即拓扑异构酶I通过破坏R环和D环来抑制末端区域的非法起始，并证明了在复制完成前拓扑异构酶IV发挥作用以使汇聚的复制叉到达其加倍点的特定需求。我们提出，当空间减小可能会阻止回旋酶结合和发挥功能时，汇聚的复制叉之间产生的正超螺旋会转化为前连环体并由拓扑异构酶IV解决。
