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Torsion-Driven Plectoneme Formation During Nanopore Translocation of DNA Polymers

Fei Zheng1,2,3, Antonio Suma4, Christopher Maffeo5, Kaikai Chen2, Mohammed Alawami1, Jingjie Sha3, Aleksei Aksimentiev5, Cristian Micheletti6,*, and Ulrich F. Keyser1,†

  • *Contact author: michelet@sissa.it
  • †Contact author: ufk20@cam.ac.uk
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Phys. Rev. X 15, 031041 – Published 12 August, 2025

DOI: https://doi.org/10.1103/spyg-kl86

Abstract

The transport of DNA polymers through nanoscale pores is central to many biological processes, from bacterial gene exchange to viral infection. In single-molecule nanopore sensing, the detection of nucleic acid and protein analytes relies on the passage of a long biopolymer through a nanoscale aperture. Understanding the dynamics of polymer translocation through nanopores, especially the relation between ionic current signal and polymer conformations, is thus essential for the successful identification of targets. Here, by analyzing ionic current traces of dsDNA translocation, we reveal that features up to now uniquely associated with knots are instead different structural motifs: plectonemes. By combining experiments and simulations, we demonstrate that such plectonemes form because of the solvent flow that induces rotation of the helical DNA fragment in the nanopore, causing torsion propagation outwards from the pore. Molecular dynamic simulations reveal that plectoneme nucleation is dominated by the applied torque, while the translocation time and size of the plectonemes depend on the coupling of torque and pulling force, a mechanism that might also be relevant for in vivo DNA organization. Experiments with nicked DNA constructs show that the number of plectonemes depends on the rotational constraints of the translocating molecules. Thus, our work introduces plectonemes as essential structural features that must be considered for accurate analysis of dsDNA polymers in the nanopore.

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Popular Summary

The motion of DNA through tiny channels called nanopores is a powerful tool used in genetic sensing and analysis, with broad applications in biology and medicine. Scientists have long believed that certain unusual signals in these experiments were caused by knots forming in the DNA. In our study, we challenge this assumption by showing that another type of DNA structure, called a plectoneme—where the DNA twists and coils around itself—also plays a major role.

To explore this, we combine detailed experiments with computer simulations. We observe that as DNA is pulled through a nanopore, the movement of fluid inside the pore causes the DNA to rotate. This rotation builds up twisting forces along the molecule, leading to the formation of plectonemes. We find that plectonemes are actually more common than knots and leave a distinct, longer-lasting signature in the nanopore signals. When we further alter the DNA to prevent twist propagation, plectoneme formation drops significantly.

This discovery reshapes how we interpret results from nanopore experiments and opens new opportunities for studying DNA structure and behavior. Being able to tell the difference between knots and plectonemes can improve our understanding of DNA organization inside living cells, how DNA entanglement changes during cell processes, and how to design more sensitive tools for biosensing and genetic analysis.

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References (59)

  1. Y.-L. Ying, Z.-L. Hu, S. Zhang, Y. Qing, A. Fragasso, G. Maglia, A. Meller, H. Bayley, C. Dekker, and Y.-T. Long, Nanopore-based technologies beyond DNA sequencing, Nat. Nanotechnol. 17, 1136 (2022).
  2. L. Xue, H. Yamazaki, R. Ren, M. Wanunu, A. P. Ivanov, and J. B. Edel, Solid-state nanopore sensors, Nat. Rev. Mater. 5, 931 (2020).
  3. M. Vayssières, N. Marechal, L. Yun, B. L. Duran, N. K. Murugasamy, J. M. Fogg, L. Zechiedrich, M. Nadal, and V. Lamour, Structural basis of DNA crossover capture by Escherichia Coli DNA gyrase, Science 384, 227 (2024).
  4. K. Coshic, C. Maffeo, D. Winogradoff, and A. Aksimentiev, The structure and physical properties of a packaged bacteriophage particle, Nature (London) 627, 905 (2024).
  5. M. Zhang, R. Nixon, F. Schaufelberger et al., Mechanical scission of a knotted polymer, Nat. Chem. 16, 1366 (2024).
  6. E. Kim, A. M. Gonzalez, B. Pradhan, J. van der Torre, and C. Dekker, Condensin-driven loop extrusion on supercoiled DNA, Nat. Struct. Mol. Biol. 29, 719 (2022).
  7. J. Lee, M. Wu, J. T. Inman et al., Chromatinization modulates topoisomerase II processivity, Nat. Commun. 14, 6844 (2023).
  8. A. L. Pyne, A. Noy, K. H. Main et al., Base-pair resolution analysis of the effect of supercoiling on DNA flexibility and major groove recognition by triplex-forming oligonucleotides, Nat. Commun. 12, 1053 (2021).
  9. A. D. Bates and A. Maxwell, DNA Topology (Oxford University Press, USA, 2005).
  10. A. Dorey and S. Howorka, Nanopore DNA sequencing technologies and their applications towards single-molecule proteomics, Nat. Chem. 16, 314 (2024).
  11. E. A. Manrao, I. M. Derrington, A. H. Laszlo, K. W. Langford, M. K. Hopper, N. Gillgren, M. Pavlenok, M. Niederweis, and J. H. Gundlach, Reading DNA at single-nucleotide resolution with a mutant MspA nanopore and phi29 DNA polymerase, Nat. Biotechnol. 30, 349 (2012).
  12. G. M. Cherf, K. R. Lieberman, H. Rashid, C. E. Lam, K. Karplus, and M. Akeson, Automated forward and reverse ratcheting of DNA in a nanopore at 5-Å precision, Nat. Biotechnol. 30, 344 (2012).
  13. I. M. Derrington, J. M. Craig, E. Stava et al., Subangstrom single-molecule measurements of motor proteins using a nanopore, Nat. Biotechnol. 33, 1073 (2015).
  14. K. Chen, I. Jou, N. Ermann, M. Muthukumar, U. F. Keyser, and N. A. W. Bell, Dynamics of driven polymer transport through a nanopore, Nat. Phys. 17, 1043 (2021).
  15. N. A. Bell, K. Chen, S. Ghosal, M. Ricci, and U. F. Keyser, Asymmetric dynamics of DNA entering and exiting a strongly confining nanopore, Nat. Commun. 8, 380 (2017).
  16. C.-Y. Lin, R. Fotis, Z. Xia, K. Kavetsky, Y.-C. Chou, D. J. Niedzwiecki, M. Biondi, F. Thei, and M. Drndić, Ultrafast polymer dynamics through a nanopore, Nano Lett. 22, 8719 (2022).
  17. J. Li, M. Gershow, D. Stein, E. Brandin, and J. A. Golovchenko, DNA molecules and configurations in a solid-state nanopore microscope, Nat. Mater. 2, 611 (2003).
  18. W. Reisner, K. J. Morton, R. Riehn, Y. M. Wang, Z. Yu, M. Rosen, J. C. Sturm, S. Y. Chou, E. Frey, and R. H. Austin, Statics and dynamics of single DNA molecules confined in nanochannels, Phys. Rev. Lett. 94, 196101 (2005).
  19. Y. Qi, L. Zeng, A. Khorshid, R. J. Hill, and W. W. Reisner, Compression of nanoslit confined polymer solutions, Macromolecules 51, 617 (2018).
  20. A. Khorshid, S. Amin, Z. Zhang, T. Sakaue, and W. W. Reisner, Nonequilibrium dynamics of nanochannel confined DNA, Macromolecules 49, 1933 (2016).
  21. N. Ermann, N. Hanikel, V. Wang, K. Chen, N. E. Weckman, and U. F. Keyser, Promoting single-file DNA translocations through nanopores using electro-osmotic flow, J. Chem. Phys. 149, 163311 (2018).
  22. R. Kumar Sharma, I. Agrawal, L. Dai, P. S. Doyle, and S. Garaj, Complex DNA knots detected with a nanopore sensor, Nat. Commun. 10, 4473 (2019).
  23. K. Briggs, G. Madejski, M. Magill, K. Kastritis, H. W. de Haan, J. L. McGrath, and V. Tabard-Cossa, DNA translocations through nanopores under nanoscale preconfinement, Nano Lett. 18, 660 (2018).
  24. B. N. Anderson, M. Muthukumar, and A. Meller, pH tuning of DNA translocation time through organically functionalized nanopores, ACS Nano 7, 1408 (2013).
  25. X. Shi, A.-K. Pumm, C. Maffeo et al., A DNA turbine powered by a transmembrane potential across a nanopore, Nat. Nanotechnol. 19, 338 (2024).
  26. C. Maffeo, L. Quednau, J. Wilson, and A. Aksimentiev, DNA double helix, a tiny electromotor, Nat. Nanotechnol. 18, 238 (2023).
  27. X. Shi, A.-K. Pumm, J. Isensee, W. Zhao, D. Verschueren, A. Martin-Gonzalez, R. Golestanian, H. Dietz, and C. Dekker, Sustained unidirectional rotation of a self-organized DNA rotor on a nanopore, Nat. Phys. 18, 1105 (2022).
  28. F. Zheng, M. Alawami, J. Zhu, C. M. Platnich, J. Sha, U. F. Keyser, and K. Chen, DNA carrier-assisted molecular ping-pong in an asymmetric nanopore, Nano Lett. 23, 11145 (2023).
  29. C. Plesa, L. Cornelissen, M. W. Tuijtel, and C. Dekker, Non-equilibrium folding of individual DNA molecules recaptured up to 1000 times in a solid state nanopore, Nanotechnology 24, 475101 (2013).
  30. M. Mihovilovic, N. Hagerty, and D. Stein, Statistics of DNA capture by a solid-state nanopore, Phys. Rev. Lett. 110, 028102 (2013).
  31. J. Sarabadani, T. Ikonen, and T. Ala-Nissila, Iso-flux tension propagation theory of driven polymer translocation: The role of initial configurations, J. Chem. Phys. 141, 214907 (2014).
  32. V. V. Palyulin, T. Ala-Nissila, and R. Metzler, Polymer translocation: The first two decades and the recent diversification, Soft Matter 10, 9016 (2014).
  33. J. Sarabadani, T. Ikonen, and T. Ala-Nissila, Theory of polymer translocation through a flickering nanopore under an alternating driving force, J. Chem. Phys. 143, 074905 (2015).
  34. J. Sarabadani and T. Ala-Nissila, Theory of pore-driven and end-pulled polymer translocation dynamics through a nanopore: An overview, J. Phys. Condens. Matter 30, 274002 (2018).
  35. T. Ikonen, A. Bhattacharya, T. Ala-Nissila, and W. Sung, Influence of non-universal effects on dynamical scaling in driven polymer translocation, J. Chem. Phys. 137, 085101 (2012).
  36. C. Plesa, D. Verschueren, S. Pud, J. van der Torre, J. W. Ruitenberg, M. J. Witteveen, M. P. Jonsson, A. Y. Grosberg, Y. Rabin, and C. Dekker, Direct observation of DNA knots using a solid-state nanopore, Nat. Nanotechnol. 11, 1093 (2016).
  37. L. Olavarrieta, P. Hernandez, D. B. Krimer, and J. B. Schvartzman, DNA knotting caused by head-on collision of transcription and replication, J. Mol. Biol. 322, 1 (2002).
  38. R. W. Deibler, J. K. Mann, D. W. L. Sumners et al., Hin-mediated DNA knotting and recombining promote replicon dysfunction and mutation, BMC Mol. Biol. 8, 44 (2007).
  39. J. Portugal and A. Rodríguez-Campos, T7 RNA polymerase cannot transcribe through a highly knotted DNA template, Nucleic Acids Res. 24, 4890 (1996).
  40. A. Valdés, J. Segura, S. Dyson , B. Martínez-García, and J. Roca, DNA knots occur in intracellular chromatin, Nucleic Acids Res. 46, 650 (2018).
  41. A. Suma and C. Micheletti, Pore translocation of knotted DNA rings, Proc. Natl. Acad. Sci. U.S.A. 114, E2991 (2017).
  42. C. Micheletti and E. Orlandini, Knotting and unknotting dynamics of DNA strands in nanochannels, ACS Macro Lett. 3, 876 (2014).
  43. L. Coronel, A. Suma, and C. Micheletti, Dynamics of supercoiled DNA with complex knots: Large-scale rearrangements and persistent multi-strand interlocking, Nucleic Acids Res. 46, 7533 (2018).
  44. A. R. Klotz, B. W. Soh, and P. S. Doyle, Motion of knots in DNA stretched by elongational fields, Phys. Rev. Lett. 120, 188003 (2018).
  45. R. K. Sharma, I. Agrawal, L. Dai, P. Doyle, and S. Garaj, DNA knot malleability in single-digit nanopores, Nano Lett. 21, 3772 (2021).
  46. See Supplemental Material at http://link.aps.org/supplemental/10.1103/spyg-kl86 for methods and supporting data.
  47. E. Uehara, L. Coronel, C. Micheletti, and T. Deguchi, Bimodality in the knotting probability of semiflexible rings suggested by mapping with self-avoiding polygons, React. Funct. Polym. 134, 141 (2019).
  48. L. Tubiana, A. Rosa, F. Fragiacomo, and C. Micheletti, Spontaneous knotting and unknotting of flexible linear polymers: Equilibrium and kinetic aspects, Macromolecules 46, 3669 (2013).
  49. S. Amin, A. Khorshid, L. Zeng, P. Zimny, and W. Reisner, A nanofluidic knot factory based on compression of single DNA in nanochannels, Nat. Commun. 9, 1506 (2018).
  50. A. Khorshid, P. Zimny, D. Tétreault-La Roche, G. Massarelli, T. Sakaue, and W. Reisner, Dynamic compression of single nanochannel confined DNA via a nanodozer assay, Phys. Rev. Lett. 113, 268104 (2014).
  51. A. Rosa, M. Di Ventra, and C. Micheletti, Topological jamming of spontaneously knotted polyelectrolyte chains driven through a nanopore, Phys. Rev. Lett. 109, 118301 (2012).
  52. Y. A. Fosado, D. Michieletto, C. A. Brackley, and D. Marenduzzo, Nonequilibrium dynamics and action at a distance in transcriptionally driven DNA supercoiling, Proc. Natl. Acad. Sci. U.S.A. 118, e1905215118 (2021).
  53. N. Laohakunakorn, V. V. Thacker, M. Muthukumar, and U. F. Keyser, Electroosmotic flow reversal outside glass nanopores, Nano Lett. 15, 695 (2015).
  54. N. Laohakunakorn and U. F. Keyser, Electroosmotic flow rectification in conical nanopores, Nanotechnology 26, 275202 (2015).
  55. G. Chirico and J. Langowski, Kinetics of DNA supercoiling studied by Brownian dynamics simulation, Biopolymers 34, 415 (1994).
  56. C. A. Brackley, A. N. Morozov, and D. Marenduzzo, Models for twistable elastic polymers in Brownian dynamics, and their implementation for LAMMPS, J. Chem. Phys. 140, 135103 (2014).
  57. J. Wilson, K. Sarthak, W. Si, L. Gao, and A. Aksimentiev, Rapid and accurate determination of nanopore ionic current using a steric exclusion model, ACS Sens. 4, 634 (2019).
  58. A. Choudhary, C. Maffeo, and A. Aksimentiev, Multi-resolution simulation of DNA transport through large synthetic nanostructures, Phys. Chem. Chem. Phys. 24, 2706 (2022).
  59. A. Valdés, L. Coronel, B. Martínez-García, J. Segura, S. Dyson, O. Díaz-Ingelmo, C. Micheletti, and J. Roca, Transcriptional supercoiling boosts topoisomerase II-mediated knotting of intracellular DNA, Nucleic Acids Res. 47, 6946 (2019).

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Phys. Rev. X 15, 031041– Published 12 August, 2025
  • Received 3 July 2024
  • Revised 8 May 2025
  • Accepted 20 May 2025
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Published by the American Physical Society under the terms of the Creative Commons Attribution 4.0 International license. Further distribution of this work must maintain attribution to the author(s) and the published article’s title, journal citation, and DOI.

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