Abstract
Kinetoplast DNA (kDNA) is a two-dimensional Olympic-ring-like network of mutually linked DNA minicircles found in certain parasites called trypanosomes. Understanding the self-assembly and replication of this structure are not only major open questions in biology but can also inform the design of synthetic topological materials. Here, we report the first high-resolution, single-molecule study of kDNA network topology using AFM and steered molecular dynamics simulations. We map out the DNA density within the network and the distribution of linking number and valence of the minicircles. We also characterize the DNA hubs that surround the network and show that they cause a buckling transition akin to that of a 2D elastic thermal sheet in the bulk. Intriguingly, we observe a broad distribution of density and valence of the minicircles, indicating heterogeneous network structure and individualism of different kDNA structures. Finally, we estimate the 2D Young modulus of the network to be orders of magnitude smaller than that of other 2D materials. Our findings explain outstanding questions in the field and offer single-molecule insights into the properties of a unique topological material.
- Received 2 September 2022
- Revised 10 December 2022
- Accepted 27 February 2023
DOI:https://doi.org/10.1103/PhysRevX.13.021010
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.
Published by the American Physical Society
Physics Subject Headings (PhySH)
Popular Summary
Trypanosomes are unique single-cell parasites: Their mitochondrial genome is made of thousands of DNA minicircles interlinked to form a giant 2D Olympic-ring-like network, known as a “kinetoplast DNA” (kDNA). This type of DNA organization is seen nowhere else in nature, and several questions on its evolution, self-assembly, and replication are far from answered. In this work, we combine high-resolution atomic force microscopy, quantitative image analysis, and molecular dynamics simulations to characterize the structure and topology of kDNAs at single-molecule resolution.
We show that the minicircles’ valence (the number of DNA rings linked to any one ring) has a broad distribution, with a mean around 3. We argue that the valence is controlled in vivo to drive the formation of a connected structure that preserves the integrity of the genome during replication yet avoids redundant constraints and a “topologically frustrated” rigid network. Our simulations explain that kDNAs undergo a buckling transition when extracted from the parasite, akin to the buckling of thermal elastic sheets. Finally, we estimate the bending rigidity and Young’s moduli of kDNAs to be thousands of times smaller than lipid vesicles. This ultrasoft nature of kDNAs may motivate the design of synthetic 2D interlocking ring networks, perhaps made of synthetic polycatenanes or DNA plasmids.
Our work sheds light on the structure and topology of a fascinating and topologically complex self-assembled genome at the single molecule scale and could inspire the creation of ultrasoft topological materials.