Smc5/6 functions with Sgs1-Top3-Rmi1 to complete chromosome replication at natural pause sites

Abstract

Smc5/6 is essential for genome structural integrity by yet unknown mechanisms. Here we find that Smc5/6 co-localizes with the DNA crossed-strand processing complex Sgs1-Top3-Rmi1 (STR) at genomic regions known as natural pausing sites (NPSs) where it facilitates Top3 retention. Individual depletions of STR subunits and Smc5/6 cause similar accumulation of joint molecules (JMs) composed of reversed forks, double Holliday Junctions and hemicatenanes, indicative of Smc5/6 regulating Sgs1 and Top3 DNA processing activities. We isolate an intra-allelic suppressor of smc6-56 proficient in Top3 retention but affected in pathways that act complementarily with Sgs1 and Top3 to resolve JMs arising at replication termination. Upon replication stress, the smc6-56 suppressor requires STR and Mus81-Mms4 functions for recovery, but not Srs2 and Mph1 helicases that prevent maturation of recombination intermediates. Thus, Smc5/6 functions jointly with Top3 and STR to mediate replication completion and influences the function of other DNA crossed-strand processing enzymes at NPSs.

Pathogenic variants in SLF2 and SMC5 cause segmented chromosomes and mosaic variegated hyperploidy

“…19a, b , 15c, d ). This is in line with previous work demonstrating a role for the SMC5/6 complex in resolving recombination intermediates 41 – 44 . We also observed an increased frequency of telomeric SCEs in SLF2-mutant LCLs (Supplementary Fig.…”

Parental histone deposition on the replicated strands promotes error-free DNA damage tolerance and regulates drug resistance

“…Recombination by template switching is mediated by the formation of sister chromatid junctions composed of pseudo-double Holliday junction-like structures, which are subsequently processed by the Sgs1–Top3–Rmi1 (STR) complex ( Branzei et al 2008 ; Giannattasio et al 2014 ). To address potential defects in the formation/stability of template switch intermediates, we used a Tc-sgs1 mutant background in which Sgs1 can be conditionally depleted upon addition of tetracycline ( Agashe et al 2021 ). This strategy avoids potential growth defects of double-knockout mutants, already noted for sgs1 Δ ctf4 Δ ( Fumasoni et al 2015 ), while enabling stabilization of recombination structures arising in the course of the experiment.…”

Computed structures of core eukaryotic protein complexes

“…Our model of the Siz2 and Mms21 SUMO ligase complex (fig. S21A) suggests that both E3s could act jointly to modify DNA-associated substrates, perhaps through the DNA binding SAP domain of Siz2 ( 53 ) or involving the Mms21 (Nse2)–containing Smc5–6 complex, which modulates DNA recombination, replication, and repair ( 54 , 55 ). The Smc5–6 complex contains another RING-finger E3 ligase–like subunit, Nse1 ( 56 ), that interacts with Nse3 and Nse4.…”