Duplex DNA and BLM regulate gate opening by the human TopoIIIα-RMI1-RMI2 complex
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Topoisomerase IIIα is a type 1A topoisomerase that forms a complex with RMI1 and RMI2 called TRR in human cells. TRR plays an essential role in resolving DNA replication and recombination intermediates, often alongside the helicase BLM. While the TRR catalytic cycle is known to involve a protein-mediated single-stranded (ss)DNA gate, the detailed mechanism is not fully understood. Here, we probe the catalytic steps of TRR using optical tweezers and fluorescence microscopy. We demonstrate that TRR forms an open gate in ssDNA of 8.5 ± 3.8 nm, and directly visualize binding of a second ssDNA or double-stranded (ds)DNA molecule to the open TRR-ssDNA gate, followed by catenation in each case. Strikingly, dsDNA binding increases the gate size (by ~16%), while BLM alters the mechanical flexibility of the gate. These findings reveal an unexpected plasticity of the TRR-ssDNA gate size and suggest that TRR-mediated transfer of dsDNA may be more relevant in vivo than previously believed.
Originalsprog | Engelsk |
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Artikelnummer | 584 |
Tidsskrift | Nature Communications |
Vol/bind | 13 |
Udgave nummer | 1 |
ISSN | 2041-1723 |
DOI | |
Status | Udgivet - 2022 |
Bibliografisk note
Funding Information:
We are grateful to Daan Vorselen for generating the step-fitting algorithm, Dian Spak-man for helpful feedback, and Giulia Bergamaschi for help with the figures. This work was supported by a Chemical Sciences Top grant from the Netherlands Organization for Scientific Research (NWO) (714.015.002, G.J.L.W, E.J.G.P, G.A.K), an NWO ENW grant (OCENW.GROOT.2019.012, G.J.L.W.), the NovoNordisk Foundation (Chromocapture; NNF18OC0034948: G.J.L.W., K.S., A.H.B., and I.D.H) and the Danish National Research Foundation (DNRF115) and Nordea Foundation (K.S., A.H.B., and I.D.H.).
Publisher Copyright:
© 2022, The Author(s).
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