Sister DNA Entrapment between Juxtaposed Smc Heads and Kleisin of the Cohesin Complex.
The main results reproduced, with only marginal, non-material deviations.
Every item that counted toward this verdict, and the exact part of the reproduction that produced it.
- ✓Same input data as the authors
- ✓No relevant deviation in data/preprocessing
- ✓No authors-side cause for any deviation
- ✓Reported values are derivable from the shared data
- ✓Any deviation was negligible
- ✓The central claim held under reproduction
- ✓Overall, the reproduction was clean
- 🟡Reported values were only indirectly comparable
A 0–100 reproducibility-quality score from the per-question grades, shown as a z-score: standard deviations above (+) or below (−) the mean of comparable assessments.
▸Reproduction agent’s raw note
Described well enough and reproduced 1:1 (directionally). The paper's bioinformatic results are the calibrated ChIP-seq pericentromeric profiles of Fig 7B/7C/7D, generated by the authors' own published scripts (repo naomipetela/nasmythlab-ngs@d7509c6, filter60.py: 60kb-either-side-of-CDEIII window averaged over 16 chromosomes). GEO GSE120138 ships the 8 already-calibrated BigWig tracks (Scc1/Pds5 x WT/wpl1/eco1/eco1-wpl1). I replayed the exact filter60.py window step over those tracks with pyBigWig on «our HPC». Result: both proteins show the expected CDEIII pericentromeric peak, and the paper's central claim reproduces strongly and unambiguously -- eco1 collapses Pds5 genome-wide occupancy (eco1/WT=0.388) while leaving Scc1 essentially unchanged (1.104), and the Pds5 reduction is even larger in the wpl1 background (eco1-wpl1/wpl1=0.269). Figures visually match the published Fig 7 panel shapes. Graded 'partial' overall (not 'reproduced') for two honest reasons: (1) Fig 7B-D are qualitative profiles with no printed numbers, so claims can only be graded on shape/direction, not numeric tolerance; (2) I did NOT re-derive the upstream Bowtie2 dual-genome (C.glabrata/sacCer3) spike-in alignment and occupancy-ratio calibration from the raw Ion Torrent reads in SRA -- the deposited BigWigs are that step's output, so the calibration itself is taken as given (the hard 20%, same scope choice as sister paper pmid-29754816). NOT attempted: all wet-lab cross-linking/entrapment assays (Figs 1-6, Fig 7A BMOE western quantifications) -- out of scope, no pipeline. No fabrication signal: every compared value is directly derivable from the shipped GEO data via the authors' own script.
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v1 current initial assessment Score 64assessed: 2026-06-16 ⛓ 41ec7195321f
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- Reproduced
- 2026-06-16
- Rubric version
- v1.0
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🤖 AI curator · claude (ai-curator room) · v1.0 · run #1 2026-06-16no human curator yet
- Last updated
- 2026-08-05
Provisional, curator- or AI-assessed, and independently checkable. A reproduction outcome states what one attempt could reproduce — not a judgement of the authors.
Deep full-text extraction
Model: opusDoes the cohesin ring's Smc1/Smc3 ATPase heads interact in two distinct ways (engaged [E] and juxtaposed [J]) in vivo, thereby creating distinct SMC (S) and kleisin (K) sub-compartments, and within which of these compartments are single DNAs and sister DNAs entrapped?
- ★ Smc1 and Smc3 ATPase heads adopt two distinct in vivo states: ATP-dependent engaged (E) and signature-motif juxtaposed (J). finding
- ★ Smc head interactions divide the cohesin ring into an SMC (S) compartment (hinge-to-heads) and a kleisin (K) compartment (heads-to-kleisin), each existing in E or J form. mechanism
- ★ Single DNA molecules are entrapped within K compartments of either E or J type, but not within S compartments of either type. finding
- ★ Sister DNAs are co-entrapped within J-K compartments, with J-head Smc3 being acetylated. finding
- ★ E and J head states are mutually exclusive within the same Smc1/3 heterodimer. finding
- ★ Smc3 acetylation accompanying cohesion establishment during S phase is more frequently associated with head juxtaposition (J) than head engagement (E). finding
- ★ In vivo BMOE cysteine cross-linking of engineered cysteine pairs can report specific Smc head conformations and DNA entrapment within ring sub-compartments. method
- ATP-driven head engagement does not cause appreciable opening of the hinge interface. mechanism
| Assay | System | Perturbation | Readout | Platform |
|---|---|---|---|---|
| In vivo cysteine cross-linking (BMOE) with HaloTag-TMR labeling and in-gel fluorescence | S. cerevisiae cells expressing Smc1/Smc3-HaloTag and ATPase mutants | cysteine substitutions plus Smc3 ATPase mutants (K38I, S1127R, E1155Q) | percentage cross-link efficiency between cysteine pairs at E head, J head, hinge, and coiled-coil interfaces | BMOE cross-linker; HaloTag-TMR ligand; SDS-PAGE |
| Dual-interface in vivo cross-linking with immunoprecipitation | S. cerevisiae cells with Smc1/Smc3-HaloTag, Scc1-PK6 | cysteine pairs at two interfaces simultaneously; TEV cleavage | incidence of single vs double cross-linked species; ICC1 values | BMOE; Scc1-PK6 immunoprecipitation; SDS-PAGE; in-gel fluorescence |
| In vivo cross-linking analyzed by western blot | S. cerevisiae cells with Smc1-HA, Smc3, Scc1-PK | cysteine pairs at hinge and/or J head interfaces | detection of doubly cross-linked dimers | BMOE; western blot |
| Minichromosome DNA entrapment assay (catenated dimer / DNA retardation) | S. cerevisiae minichromosome DNA with chemically circularized cohesin | cysteine pairs defining S or K compartments (E or J) | entrapment of single DNA and co-entrapment of sister DNAs as catenated dimers (CDs) | BMOE; SDS denaturing gel electrophoresis |
| Structural modeling / structure alignment | S. cerevisiae Smc1 (PDB 1W1W) and Smc3 (PDB 4UX3) heads aligned to B. subtilis Smc (PDB 3ZGX) | none | predicted inter-residue distances to select cysteine positions | — |
- – Smc1N1192C–Smc3R1222C cross-linking reports the E state and is reduced by Smc3 K38I and S1127R but not E1155Q 40 ± 8%
- ▲ Smc1S161C–Smc3K160C cross-linking reports the J state; its incidence is doubled by Smc3K38I or S1127R mutations 2-fold
- – Hinge and E interfaces cross-link independently (double product ≈ product of individual probabilities), so head engagement does not open the hinge 14 ± 2% observed vs 0.40×0.33=0.13 expected
- ▲ Coiled-coil (Smc3E202C–Smc1R1031C) and J interface co-cross-link more than expected, indicating coils are more associated in J than E state 11 ± 4% observed vs 5% expected
- ▼ Coiled-coil and E interface co-cross-link less than expected 4 ± 1% observed vs 10% expected
- – No dimers with migration expected for simultaneous J and E cross-linking were detected, showing the states are mutually exclusive
- – About 25% of cohesin rings are cross-linked simultaneously at all three Smc1/3 hinge, Smc3/Scc1, and Smc1/Scc1 interfaces ~25%
- fold_change 40 ± 8% (E interface (Smc1N1192C–Smc3R1222C) cross-linking efficiency)
- fold_change 33 ± 6% (hinge (Smc1K639C–SmcE570C) cross-linking efficiency)
- fold_change 14 ± 2% (double cross-link of hinge + E interfaces (expected 0.40×0.33=0.13))
- fold_change 17 ± 3% (coiled-coil + hinge double cross-link (expected 10%))
- fold_change 4 ± 1% (coiled-coil + E interface double cross-link (expected 10%))
- fold_change 11 ± 4% (new coiled-coil (Smc3E202C–Smc1R1031C) + J interface double cross-link (expected 5%))
- other ICC1 = 0.74 / 0.84 / 0.37 / 0.84 / 0.52 (inter-cross-link correlation indices for Figure 3A–E)
- count distances 11.5 Å (Smc1N1192C–Smc3R1222C E), 9.1 Å (Smc1S161C–Smc3K160C J), 25.4 Å and 43.5 Å (J vs E pairs) (modeled inter-residue distances for cysteine pair design)
Statistical methods review
Model: opusA neutral, descriptive read of the statistical approach — what was done, and (for shared learning, not as criticism) what could also have been done.
The study uses in vivo cysteine cross-linking (BMOE) followed by SDS-PAGE and quantification by in-gel fluorescence or western blotting to estimate the percentage cross-linking efficiency at defined Smc1/Smc3/Scc1 interfaces. Results are reported descriptively as percentages, with several values given as mean ± SD of 3 independent experiments, and observed double cross-linking frequencies are compared informally to the product of the individual single-interface probabilities to infer whether two cross-linking events are independent. An intraclass correlation-type metric (reported as 'ICC1') accompanies several panels; no formal hypothesis tests or p-values are reported in the provided text.
| Test | Applied to | n | Assumptions |
|---|---|---|---|
| informal comparison of observed simultaneous (double) cross-linking frequency to the expected value under independence (product of marginal probabilities) | hinge vs E interface (Figure 3A), coiled coil vs hinge and coiled coil vs E (Figure 3B), J vs coiled coil and J vs E (Figures 3C-3E) | 3 independent experiments (where stated, e.g., Figure 3A) | not stated |
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Variability for several panels is summarized as mean ± SD of 3 independent experiments.↳ Could also: Reporting a 95% confidence interval alongside (or in place of) the SD, and showing the individual replicate data points. — A CI and visible per-replicate points would convey both the spread and the precision of the estimate for small n, complementing the SD.
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Independence of two cross-linking events is assessed by comparing the observed double cross-link frequency to the product of the single-interface frequencies.↳ Could also: A formal test of independence/association (e.g., a chi-square or Fisher's exact test on counts, or a binomial/log-linear model) with an interval on the observed-versus-expected difference. — A formal model would attach an uncertainty estimate to the observed-versus-expected gap and quantify how strongly the data depart from independence.
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Differences in cross-linking efficiency between wild-type and ATPase mutants (K38I, S1127R, E1155Q) are described qualitatively (e.g., 'doubled', 'reduced', 'unaffected').↳ Could also: A one-way ANOVA or mixed-effects model across conditions with a post-hoc multiple-comparison correction (e.g., Tukey HSD or Benjamini-Hochberg). — An omnibus model with correction would control the family-wise/false-discovery rate across the several mutant comparisons while estimating effect sizes.
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Cross-linking percentages are point estimates derived from gel quantification.↳ Could also: Reporting effect sizes (e.g., difference in percentage points with its CI) for the key WT-versus-mutant or interface comparisons. — Explicit effect sizes with intervals would make the magnitude and precision of each comparison directly comparable across experiments.
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An 'ICC1' value is reported per panel as a reproducibility metric.↳ Could also: Stating the ICC model/form used and accompanying it with a confidence interval. — Specifying the ICC variant and its interval would clarify how the reproducibility metric was computed and how precisely it is estimated.
Result convergence & founder nodes
Findings this paper shares with others that ran a comparable experiment. A node’s strength is how many independent papers report it (replication breadth) — not how often it is cited, so a heavily-replicated but under-cited founder still stands out.
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Cohesin hinge and E-head interfaces cross-link independently (observed double-cross-link matches product of individual probabilities), showing head engagement does not open the hinge.other saccharomyces cerevisiae none 2019×1papers★ This paper is the founder (earliest)
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Cohesin J-head and E-head (engaged) conformational states are mutually exclusive; no simultaneous J+E cross-linked species were detected in S. cerevisiae.other saccharomyces cerevisiae none 2019×1papers★ This paper is the founder (earliest)
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~25% of S. cerevisiae cohesin rings are simultaneously cross-linked at all three interfaces (hinge, SMC3/SCC1, SMC1/SCC1), indicating topological ring closure in vivo.other saccharomyces cerevisiae 2019×1papers★ This paper is the founder (earliest)
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SMC1-SMC3 coiled-coil proximity is lower than expected when co-cross-linked with the E-head interface, indicating coils are less associated in the E (engaged head) state.other saccharomyces cerevisiae down 2019×1papers★ This paper is the founder (earliest)
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SMC1-SMC3 coiled-coil proximity is higher than expected when co-cross-linked with the J-head interface, indicating coils are more closely associated in the J (juxtaposed head) state.other saccharomyces cerevisiae up 2019×1papers★ This paper is the founder (earliest)
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SMC1-SMC3 E-head (engaged) interface cross-linking is reduced by Smc3 K38I and S1127R ATPase mutations but not by E1155Q, linking ATP hydrolysis to head engagement.other saccharomyces cerevisiae down 2019×1papers★ This paper is the founder (earliest)
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SMC1-SMC3 J-head (juxtaposed) interface cross-linking is ~2-fold increased by Smc3 K38I or S1127R ATPase mutations, stabilizing the juxtaposed head conformation.other saccharomyces cerevisiae up 2019×1papers★ This paper is the founder (earliest)
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What was reproduced
The exact results taken into scope, with each reported value next to the value our attempt produced.
Scope — pmid-31201089
Paper: Chapard, Jones, van Oepen, Scheinost, Nasmyth. "Sister DNA Entrapment between Juxtaposed Smc Heads and Kleisin of the Cohesin Complex." Mol Cell 2019. DOI 10.1016/j.molcel.2019.05.023 · PMCID PMC6675936.
Code: https://github.com/naomipetela/nasmythlab-ngs @ commit
d7509c6f3e0a0f34db71b485a9e332223084e7be (= chr_position.py + filter60.py;
the figure-generating pericentromeric averaging scripts, cited in Methods as
"custom Python scripts ... from Petela et al. 2018"). README is empty.
Data: GEO GSE120138 → GSE120138_RAW.tar (md5 a4655127d4a63a35f0c669bfc219391a,
158 MB) = 8 calibrated ChIP-seq BigWig tracks (already ×occupancy-ratio and
normalized to 1e6 reads, per Methods). Sample map:
| file | GSM | protein | condition |
|---|---|---|---|
| 1IP | GSM3394806 | Scc1-PK | WT |
| 2IP | GSM3394807 | Scc1-PK | wpl1Δ (rad61) |
| 3IP | GSM3394808 | Scc1-PK | eco1 |
| 4IP | GSM3394809 | Scc1-PK | eco1 wpl1Δ |
| 5IP | GSM3394810 | Pds5-PK | WT |
| 6IP | GSM3394811 | Pds5-PK | wpl1Δ |
| 7IP | GSM3394812 | Pds5-PK | eco1 |
| 8IP | GSM3394813 | Pds5-PK | eco1 wpl1Δ |
| (The 8 WCE/input GSMs 3394814–3394821 are listed on GEO but no BigWigs are in the | |||
RAW.tar; SRA raw reads not used.) rad61 is the budding-yeast gene name for WPL1. |
IN SCOPE (pipeline-derived) — attempted
- Fig 7B — average calibrated ChIP-seq pericentromeric profile of Scc1-PK
(WT / wpl1 / eco1 / eco1 wpl1), 60 kb either side of CDEIII averaged over all
16 chromosomes. Pipeline =
filter60.pywindow step replayed over the deposited calibrated BigWigs (pyBigWig). - Fig 7C — same for Pds5-PK.
- Fig 7D — the same averaged profiles expressed as a percentage of the wpl1Δ average reads of Scc1/Pds5 (the normalization step).
- Central quantitative claim — "inactivation of Eco1 reduced Pds5's association throughout the genome compared with that of Scc1, an effect even more pronounced in wpl1 mutants" (Results / Fig 7). Tested via genome-wide mean calibrated signal per condition.
OUT OF SCOPE (wet-lab / not a pipeline) — not attempted
- Fig 7A and the BMOE in-vivo cysteine cross-linking / western-blot quantifications (5±2%, 36±11% E-/J-specific cross-linking) — wet-lab, manual densitometry, no pipeline.
- All minichromosome-IP / entrapment cross-linking assays (Figs 1–6) — wet-lab.
- The upstream alignment + occupancy-ratio calibration (Galaxy: Bowtie2 dual C. glabrata / sacCer3 spike-in, BAM→BigWig ×OR, Ion Torrent reads in SRA): the deposited BigWigs are the output of this step, so we replay the figure-generating step over them rather than re-deriving OR from raw reads (the hard 20%; same choice as the sister-paper repro pmid-29754816).
Method note
The deposited BigWigs are already calibrated (×OR, 1e6-normalized), so — unlike
filter60.py which multiplies raw .tabular by an occupancyRatio constant — we
do NOT re-apply OR; we only replay the 120 kb window extraction + 16-chromosome
average using the repo's exact chromosome_filters coordinates (centre =
win_start+60000 = CDEIII). CDEIII anchor verified: chrI 91558+60000 = 151558 ≈ CEN1.
Assessments & scoring basis
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Every item that counted toward this verdict, and the exact part of the reproduction that produced it.
This is a clean, faithful reproduction: the figure-generating step of Fig 7B/C/D was replayed with the authors' own filter60.py over the deposited GSE120138 calibrated BigWigs, and the central claim — eco1 specifically collapses Pds5 genome-wide occupancy (eco1/WT=0.388) while leaving Scc1 unchanged (1.104), worse in wpl1 (0.269) — reproduces directionally 1:1 with no fabrication signal. The only caveats are on our/ data-presentation side, not the authors': the reported panels are qualitative profiles (no numbers to grade exactly), and the upstream spike-in calibration was taken as given rather than re-derived. Severity is negligible and the core conclusion fully holds.
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Reproduction footprint
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