Molecular Insights into the Architecture of the Human SMC5/6 Complex

Adamus, Marek; Lelkes, Edit; Potěšil, David; Ganji, Sri Ranjani; Kolesar, Peter; Zabrady, Katerina; Zdráhal, Zbyněk; Paleček, Jan

doi:10.1016/j.jmb.2020.04.024

Cited by 35 publications

(60 citation statements)

References 59 publications

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“…While Nse5/6 has been reported to bind to the head-proximal region of the Smc5/6 hexamer in S. pombe (Palecek et al ., 2006), it was shown to bind to the hinge domain in S. cerevisiae (Duan et al ., 2009b). Recent mapping experiments with human Slf1/2 showed a binding mode similar to that observed in fission yeast (Adamus et al ., 2020). While neither Nse5 nor Nse6 are essential in S. pombe they are required for viability in all other organisms tested (Aragon, 2018; Oravcova et al ., 2019).…”

supporting

confidence: 59%

“…Structural information on the Nse5/6 complex is currently limited to the disordered N-terminus of Nse6 that interacts with Rtt107 (Wan et al ., 2019). Compounding this, there are conflicting reports in the literature on the presence of α-helical HEAT repeats in Nse6 and its putative relatedness to human Slf2, a subunit of the Smc5/6 associated Slf1/Slf2 complex (Adamus et al ., 2020; Pebernard et al ., 2006; Raschle et al ., 2015; Yu et al ., 2021). Limited proteolysis of purified Nse5/6 (Fig 2B) led to the design of two N-terminally truncated Nse6 fragments (154-C and 177-C) which were easily co-purified with Nse5 indicating a stable interaction with Nse5.…”

Section: Resultsmentioning

confidence: 99%

“…In our XL-MS experiments, such contact points were indeed mapped, for example near the active sites of Smc5 and Smc6 and at a Nse5 surface located opposite of the Nse6 N-terminus (Fig 3C) (Gutierrez-Escribano et al ., 2020). An equivalent interaction has recently also been mapped between the joint regions of human Smc5/6 and the loader Slf1/2 by Yeast-Two-Hybrid assays (Adamus et al ., 2020). Whether the yeast and human interfaces indeed share a similar fold and a common origin remains to be determined due to the absence of recognizable sequence conservation in the relevant regions.…”

Section: Discussionmentioning

confidence: 99%

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Nse5/6 inhibits the Smc5/6 ATPase to facilitate DNA substrate selection

Basquin

Steigenberger

Schäfer

et al. 2021

Preprint

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Eukaryotic cells employ three SMC complexes to control DNA folding and topology. The Smc5/6 complex plays roles in DNA repair and in preventing the accumulation of deleterious DNA junctions. To elucidate how specific features of Smc5/6 govern these functions, we reconstituted the yeast holo-complex. We found that the Nse5/6 sub-complex strongly inhibited the Smc5/6 ATPase by preventing productive ATP binding. This inhibition was relieved by plasmid DNA binding but not by short linear DNA, while opposing effects were observed without Nse5/6. We uncovered two binding sites for Nse5/6 on Smc5/6, based on an Nse5/6 crystal structure and cross-linking mass spectrometry data. One binding site is located at the Smc5/6 arms and one at the heads, the latter likely exerting inhibitory effects on ATP hydrolysis. Cysteine cross-linking demonstrated that the interaction with Nse5/6 anchored the ATPase domains in a non-productive state, which was destabilized by ATP and DNA. Under similar conditions, the Nse4/3/1 module detached from the ATPase. Altogether, we show how DNA substrate selection is modulated by direct inhibition of the Smc5/6 ATPase by Nse5/6.

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supporting

confidence: 59%

Section: Resultsmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

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Nse5/6 inhibits the Smc5/6 ATPase to facilitate DNA substrate selection

Basquin

Steigenberger

Schäfer

et al. 2021

Preprint

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“…Two additional proteins, SLF1 and SLF2 (SMC5/6 localization factors) form a dimer that primarily interacts with the SMC5/6 complex during DNA repair in response to DNA lesions that block DNA replication (Räschle et al, 2015). SLF2 binds to the arms of SMC5 and SMC6 and may lock the arms together to immobilize SMC5/6 at sites of DNA damage (Adamus et al, 2020;Räschle et al, 2015).…”

Section: Introductionmentioning

confidence: 99%

SMC5/6 is required for replication fork stability and faithful chromosome segregation during neurogenesis

Atkins

et al. 2020

eLife

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Mutations of SMC5/6 components cause developmental defects, including primary microcephaly. To model neurodevelopmental defects, we engineered a mouse wherein Smc5 is conditionally knocked out (cKO) in the developing neocortex. Smc5 cKO mice exhibited neurodevelopmental defects due to neural progenitor cell (NPC) apoptosis, which led to reduction in cortical layer neurons. Smc5 cKO NPCs formed DNA bridges during mitosis and underwent chromosome missegregation. SMC5/6 depletion triggers a CHEK2-p53 DNA damage response, as concomitant deletion of the Trp53 tumor suppressor or Chek2 DNA damage checkpoint kinase rescued Smc5 cKO neurodevelopmental defects. Further assessment using Smc5 cKO and auxin-inducible degron systems demonstrated that absence of SMC5/6 leads to DNA replication stress at late-replicating regions such as pericentromeric heterochromatin regions. In summary, SMC5/6 is important for completion of DNA replication prior to entering mitosis, which ensures accurate chromosome segregation. Thus, SMC5/6 functions are critical in highly proliferative stem cells during organism development.

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“…Besides the domains that form extensive interfaces in the Nse5-6 heterodimer, Nse5-HAD and Nse6-CAD are in close proximity with the head and arm regions, respectively. Recent yeast two-hybrid findings suggest that the human Nse6 homolog binds to the Smc5/6 arm regions, indicating that this interaction is evolutionarily conserved (47). Nse5-6 was reported to associate with the fragments of Smc5 and 6 hinge in vitro (20), but their crosslink in the holo-Smc5/6 was not seen here.…”

Section: Discussionmentioning

confidence: 56%

Integrative analysis reveals unique features of the Smc5/6 complex

Ser

et al. 2021

Preprint

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Structural maintenance of chromosomes (SMC) complexes are critical chromatin modulators. In eukaryotes, the cohesin and condensin SMC complexes organize chromatin, while the Smc5/6 complex directly regulates DNA replication and repair. The molecular basis for Smc5/6’s distinct functions is currently poorly understood. Here, we report an integrative structural study of the budding yeast Smc5/6 complex using electron microscopy, cross-linking mass spectrometry, and computational modeling. We show that while the complex shares a similar overall organization with other SMC complexes, it possesses several unique features. In contrast to the reported folded-arm structures of cohesin and condensin, our data suggest that Smc5 and Smc6 arm regions do not fold back on themselves. Instead, these long filamentous regions interact with subunits uniquely acquired by the Smc5/6 complex, namely the Nse2 SUMO ligase and the Nse5-Nse6 subcomplex. Further, we show that Nse5-Nse6 subcomplex adopts a novel structure with an extensive dimerization interface and multiple domains contacting other subunits of the Smc5/6 complex. We also provide evidence that the Nse5-Nse6 module uses its SUMO-binding motifs to contribute to Nse2-mediated sumoylation. Collectively, our integrative multi-scale study identifies distinct structural features of the Smc5/6 complex and functional cooperation amongst its co-evolved unique subunits.

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Molecular Insights into the Architecture of the Human SMC5/6 Complex

Cited by 35 publications

References 59 publications

Nse5/6 inhibits the Smc5/6 ATPase to facilitate DNA substrate selection

Nse5/6 inhibits the Smc5/6 ATPase to facilitate DNA substrate selection

SMC5/6 is required for replication fork stability and faithful chromosome segregation during neurogenesis

Integrative analysis reveals unique features of the Smc5/6 complex

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