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. 2023 Apr 19;616(7958):822–827. doi: 10.1038/s41586-023-05961-5

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© The Author(s) 2023

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 2 — a,b, Examples of loop extrusion blocked by (a) or passing (b) CTCF (cyan) labelled with Janelia Fluor 646 (JF646). DNA loops (red) were visualized by Sytox Orange and perpendicular buffer flow. Scale bar, 2 µm. c, Cohesin-mediated DNA loop extrusion encountering N-terminally oriented JF646-labelled CTCF (cyan). Growth of DNA loop stops after encountering CTCF at around 30 s and around 50 s. Scale bar, 2 µm. d, The same as c, but for a passing event. CTCF passes into the loop at 70 s and translocates with it. Scale bar, 2 µm. e, The fraction of loop-extrusion events blocked after encountering N- or C-terminally oriented CTCF or dCas9. Data are mean ± 95% binomial confidence interval. n = 119, 115 and 19 from 13, 3 and 3 independent experiments for N-terminal, C-terminal and dCas9 encounters, respectively. The force range between 0.04 and 0.08 pN was best covered and was therefore chosen to compare the overall blocking efficiency (Extended Data Fig. 5c,d). f, The DNA tension at the moment of the encounter was calculated by the amount of DNA outside the loop and the DNA end-to-end length (Supplementary Note). g, The loop-extrusion blocking probability of N- or C-terminally oriented CTCF depends on DNA tension. Data are mean ± 95% binomial confidence interval. The solid lines are fits of the form 1 − exp(−F/F₀), which were used to compute the force at which 100% blocking is achieved (N-terminal encounters: P_block(F) = 147(1 − e⁻^F/0.125 pN); C-terminal encounters: P_block(F) = 115(1 − e^{−F/0.357 pN}). n per bin for N-terminal (N) and C-terminal (C) encounters: 0–0.015 pN: 17 (N) and 12 (C); 0.015–0.026 pN: 75 (N) and 77 (C); 0.026–0.05 pN: 72 (N) and 53 (C); 0.05–0.072 pN: 89 (N) and 34 (C); 0.096–0.119 pN: 40 (N) and 6 (C); and 0.119–0.142 pN: 3 (N) and 0 (C). The bin for C-terminal encounters at the highest DNA tension regime is not shown owing to insufficient observations (n < 3). Sample sizes refer to biological replicates from 13 independent experiments for N-terminal encounters and 3 independent experiments for C-terminal encounters.

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