Figure EV1. WRN helicase unfolds cruciform DNA structures.

1. Representative assay showing the activity of T7 Endonuclease I, which cleaves the 4‐way junction formed at the cruciform DNA site. Incubation with T7 Endonuclease I and SspI results in the production of bands at the expected positions of 607 and 2,119 bp, indicated by the green arrows. The SspI restriction site is 607 bp away from the cruciform site. A plasmid without the inverted repeats is cleaved less efficiently and produces bands of different sizes (lane 2), resulting from T7 Endonuclease I activity at other sites that spontaneously extrude in pUC19 DNA. The DNA molecules were resolved by standard electrophoresis on a 1% native agarose gel, stained with GelRed.
2. Representative cruciform unfolding assay with WRN, WRN‐K577M, and WRN‐E84A (all used at 10 nM). The assay was incubated either with or without EcoRI and the DNA species were separated on a 1% native agarose gel, stained with GelRed.