PT  - JOURNAL ARTICLE
AU  - Etheridge, Thomas J.
AU  - Villahermosa, Desiree
AU  - Irmisch, Anja
AU  - Watson, Adam T.
AU  - Herbert, Alex
AU  - Dang, Hung Q.
AU  - Osborne, Mark A.
AU  - Oliver, Antony W.
AU  - Carr, Antony M.
AU  - Murray, Johanne M.
TI  - Single-molecule live cell imaging of the Smc5/6 DNA repair complex
AID  - 10.1101/2020.06.19.148106
DP  - 2020 Jan 01
TA  - bioRxiv
PG  - 2020.06.19.148106
4099  - http://biorxiv.org/content/early/2020/06/20/2020.06.19.148106.short
4100  - http://biorxiv.org/content/early/2020/06/20/2020.06.19.148106.full
AB  - The Smc5/6 complex is involved in various DNA transactions and is best known for ensuring the fidelity of homologous recombination. We exploit single-molecule tracking in live fission yeast to investigate Smc5/6 chromatin association. We show that Smc5/6 is chromatin associated in unchallenged cells and this depends on the non-SMC protein Nse6. We define a minimum of two Nse6-dependent sub-pathways, one of which requires the BRCT-domain protein Brc1. Using defined mutants in genes encoding the core Smc5/6 complex subunits we show that the Nse3 double-stranded DNA binding activity and the two arginine fingers of the two Smc5/6 ATPase binding sites are critical for chromatin association. Interestingly, disrupting the ssDNA binding activity at the hinge region does not prevent chromatin association. However, unlike a mutant attenuating chromatin association, a mutant that disrupts ssDNA binding results in highly elevated levels of gross chromosomal rearrangements during replication restart. This is consistent with a downstream function for ssDNA binding in regulating homologous recombination.Competing Interest StatementThe authors have declared no competing interest.