DNA Topology Lab, Molecular Biology Institute of Barcelona (IBMB), CSIC, Barcelona, Spain.
DNA Motors Group, MRC London Institute of Medical Sciences (LMS), London, UK.
EMBO J. 2023 Feb 1;42(3):e111913. doi: 10.15252/embj.2022111913. Epub 2022 Dec 19.
Condensin, an SMC (structural maintenance of chromosomes) protein complex, extrudes DNA loops using an ATP-dependent mechanism that remains to be elucidated. Here, we show how condensin activity alters the topology of the interacting DNA. High condensin concentrations restrain positive DNA supercoils. However, in experimental conditions of DNA loop extrusion, condensin restrains negative supercoils. Namely, following ATP-mediated loading onto DNA, each condensin complex constrains a DNA linking number difference (∆Lk) of -0.4. This ∆Lk increases to -0.8 during ATP binding and resets to -0.4 upon ATP hydrolysis. These changes in DNA topology do not involve DNA unwinding, do not spread outside the condensin-DNA complex and can occur in the absence of the condensin subunit Ycg1. These findings indicate that during ATP binding, a short DNA domain delimited by condensin is pinched into a negatively supercoiled loop. We propose that this loop is the feeding segment of DNA that is subsequently merged to enlarge an extruding loop. Such a "pinch and merge" mechanism implies that two DNA-binding sites produce the feeding loop, while a third site, plausibly involving Ycg1, might anchor the extruding loop.
凝聚素是一种 SMC（染色体结构维持）蛋白复合物，它利用依赖于 ATP 的机制来挤出 DNA 环，但其具体机制仍有待阐明。在这里，我们展示了凝聚素活性如何改变相互作用 DNA 的拓扑结构。高浓度的凝聚素可抑制正 DNA 超螺旋。然而，在 DNA 环挤出的实验条件下，凝聚素会抑制负超螺旋。也就是说，在 ATP 介导加载到 DNA 上后，每个凝聚素复合物都会限制 DNA 连接数差异（∆Lk）为-0.4。当 ATP 结合时，∆Lk 增加到-0.8，并在 ATP 水解后重置为-0.4。这些 DNA 拓扑结构的变化不涉及 DNA 解旋，不会在凝聚素-DNA 复合物之外扩散，并且可以在没有凝聚素亚基 Ycg1 的情况下发生。这些发现表明，在 ATP 结合期间，由凝聚素限定的短 DNA 域被夹成一个负超螺旋环。我们提出，这个环是随后被合并以扩大挤出环的 DNA 进料段。这种“夹合和合并”机制意味着两个 DNA 结合位点产生进料环，而第三个位点（可能涉及 Ycg1）可能会固定挤出环。
