Gene Regulatory Interactions at Lamina-Associated Domains.
Provisional — an automated or curator check raised a specific concern and points reviewers here. This is NOT a final assessment and not a determination about the authors.
The main result did not reproduce in this reproduction attempt. Where our recomputation produced values that differ from the published ones, those discrepancies are listed below. This is a single automated attempt — not peer review and not a finding of error or misconduct — and differences can also arise from data access, undocumented parameters or the computing environment. The verdict can be contested via “report an error”.
Every item that counted toward this verdict, and the exact part of the reproduction that produced it.
- ✓Reported values were directly comparable
- ✓The central claim held under reproduction
- 🟡Could not use the authors’ exact input data
- 🟡A deviation arose in the data or preprocessing
- 🔴A deviation was attributed to the published material
- 🟡Reported values were not (fully) derivable from the shared data
- 🟡The deviation was non-trivial in magnitude
- 🟡Overall, the reproduction showed a material discrepancy
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 for the UPSTREAM pipeline but NOT for the cLAD-overlap counts. In-scope = Fig 2a-d pipeline-derived counts (liftOver hg19->hg38 + BEDtools 2.29.2 intersections on the authors' OWN deposited processed files: ECHi-C connections GSE140782, LMNA/C EDD peaks + H3K4me1 broadPeak GSE221288). EXACT 1:1: total ECHi-C baits = 18,710 (C1) - the bait set + liftOver reproduce perfectly. CENTRAL QUALITATIVE FINDING reproduces: K4 baits are ~2x more connected than nonK4 baits (reproduced ratio 24.25/13.28 = 1.83 vs paper 27.5/14.7 = 1.87; nonK4 mean within ~10pct). NOT reproduced: the absolute cLAD-overlap counts C2-C6 (768/225/543/6108/7988). Our constitutive-cLAD set rebuilt from the deposited EDD _peaks.bed files (merge replicates per day, intersect D0+D1+D3) gives 1203 baits-in-cLAD, not 768; the union variant gives 3572. The paper's 'cLAD coverage' definition (EDD parameters / coverage threshold / how replicates+timepoints are combined into the final cLAD set) is underspecified in the deposited files, so the exact cLAD input cannot be uniquely reconstructed from public data - this is the hard 20pct, intentionally not chased per brief. NO fabrication signal: the paper's numbers are internally consistent (225+543=768; 7988/543=14.7114.7) and our reproduced K4/nonK4 ratio matches, corroborating the biology while the absolute counts hinge on their internal cLAD definition. NOT attempted (out of scope): re-calling peaks from raw FASTQ (Bowtie2->MACS2/EDD), ChromHMM 15-state, deeptools profiles, FISH, lamin-knockdown qPCR, WiggleTools 1kb-bin track (no single reported scalar). Brief's listed code (WiggleTools) and accession (GSE109924, the authors' prior dataset) were corrected to the actual analysis tools (BEDtools/liftOver) and data (GSE221288 + GSE140782) per P16.
These records describe the outcome of reproduction attempts carried out autonomously by brainbox using large language models (LLMs). They are not peer review, not an audit, and not a determination of error or misconduct by any author. A verdict reflects what one attempt could or could not reproduce — which may depend on data access, undocumented parameters, the computing environment, or the depth of effort — and not a judgement of the people who did the work. We can be wrong, and we correct mistakes quickly: every record carries a “report an error” button.
Assessment versions
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v1 current initial assessment Score 31assessed: 2026-06-15 ⛓ ddaaebd310ed
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Provenance — full disclosure
When this reproduction was carried out, which methodology version was used, and by whom — so the record can be audited and checked independently.
- Reproduced
- 2026-06-15
- Rubric version
- v1.0
- Assessed by
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🤖 AI curator · claude (ai-curator room) · v1.0 · run #1 2026-06-15no 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: sonnetThe paper investigates how the ~10% of genes within lamina-associated domains (LADs) that remain transcriptionally active are regulated, and specifically whether these genes and their enhancers can form chromatin interactions with regulatory elements inside and outside LADs despite the generally repressive nuclear lamina environment.
- ★ Active genes in constitutive LADs (cLADs) reside in euchromatic regions with low LMNB1 and LMNA/C enrichment, i.e., are locally detached from the nuclear lamina finding
- ★ H3K4me1-marked enhancer baits in cLADs (K4 baits) show higher ECHi-C connection density than non-H3K4me1 baits (nonK4 baits) finding
- ★ The majority of ECHi-C enhancer connections originating from cLAD baits (both K4 and nonK4) occur within cLADs rather than outside finding
- ★ K4 baits preferentially connect to active/weak enhancer chromatin states, whereas nonK4 baits mainly connect to H3K9me3 heterochromatin finding
- ★ Dual-color FISH shows gene-enhancer spatial distances change during adipogenic differentiation in a manner correlated with gene expression changes finding
- ★ Knock-down of LMNA/C, but not LMNB1, is sufficient to activate the silenced IL5RA gene located adjacent to an active cLAD region finding
- Integration of public ECHi-C data with in-house ChIP-seq, RNA-seq, ATAC-seq, chromatin state modeling, FISH, and lamin knock-down to probe LAD gene regulation method
- A 15-state ChromHMM chromatin state model of adipose stem cells (ASCs) built from H3K4me1/H3K4me3/H3K27me3/H3K27ac/H3K9me3/H3K36me3 ChIP-seq resource
| Assay | System | Perturbation | Readout | Platform |
|---|---|---|---|---|
| ChIP-seq | human adipose stem cells (ASCs) | adipogenic differentiation time course (D0,D1,D3) | genome-wide enrichment of LMNA/C, H3K4me3, H3K4me1, H3K27ac, H3K9me3, H3K27me3, H3K36me3 | Nextseq 500 / Novaseq (Illumina) |
| RNA-seq | human ASCs | adipogenic differentiation (D0-D9) | gene expression (transcript normalized read counts) | GEO GSE185066 |
| ATAC-seq (reprocessed public data) | human adipose tissue, gluteofemoral fat, 10 donors | none | chromatin accessibility profile at TSS | GEO GSE143450 |
| Enhancer-capture Hi-C (ECHi-C, reprocessed public data) | human mesenchymal stem cells | adipogenic differentiation | enhancer bait-target chromatin contacts | GEO GSE140782 |
| Fluorescence in situ hybridization (FISH) | human ASCs | adipogenic differentiation (D0, D1, D3) | gene-enhancer 3D spatial distance and distance to nuclear periphery | DeltaVision imaging; NEMO analysis |
| siRNA knock-down + RT-qPCR | human ASCs | LMNA/C, LMNB1, or combined knock-down | expression of IL5RA, TRNT1, CRBN, CNTN4 | IQ SYBR green (Bio-Rad) |
| Immunoblotting (Western blot) | human ASCs | siRNA knock-down of LMNA/C/LMNB1 | LMNB1, LMNA/C, γ-Tubulin protein levels | Odyssey/Image Lab (Bio-Rad) |
| ChromHMM chromatin state modeling | human ASCs | none | 15-state genome segmentation from 6 histone marks | ChromHMM (Baum-Welch) |
- ▼ H3K4me1-high cLAD regions show significantly lower LMNA/C enrichment than the rest of the LAD p < 10^-4
- ▲ K4 baits have higher mean ECHi-C connection density than nonK4 baits 27.5 vs 14.7 targets/bait
- – Over 70% of K4 bait targets localize within cLADs >70%
- – K4 baits connect mostly to enhancer states (60%) vs H3K9me3 heterochromatin (30%); nonK4 baits connect mostly to heterochromatin (66.5%) vs enhancers (28%)
- ▲ CMKLR1 (downregulated during differentiation) shows increasing distance to its distal enhancer near the nuclear periphery
- ▼ OTUD1 (upregulated during differentiation) shows increasing gene-enhancer proximity (decreasing distance)
- ▲ IL5RA expression is induced by D3 after knock-down of LMNA/C or LMNA/C+LMNB1, but not LMNB1 alone
- – 244 expressed genes identified within adipogenic cLADs 244 genes
- pvalue p < 10^-4 (LMNA/C enrichment: H3K4me1-high cLAD regions vs rest of LAD, unpaired two-tailed t-test with Welch's correction)
- count 768 baits (ECHi-C baits (out of 18,710 total) overlapping adipogenic cLADs)
- count 225 K4 baits; 543 nonK4 baits (cLAD baits split by overlap with H3K4me1 peaks)
- mean 27.5 targets/bait (K4) vs 14.7 targets/bait (nonK4) (ECHi-C connection density within cLADs)
- count 6108 targets from K4 baits; 7988 targets from nonK4 baits (ECHi-C connections identified)
- count 244 expressed genes (expressed genes within adipogenic cLADs)
- other >70% of K4 bait targets within cLADs (localization of K4 bait ECHi-C connections)
- other K4 baits: 60% enhancer, 30% Het, 9.5% ReprPc; nonK4 baits: 28% enhancer, 66.5% Het, 5% ReprPc (chromatin state distribution of ECHi-C bait targets)
Statistical methods review
Model: sonnetA neutral, descriptive read of the statistical approach — what was done, and (for shared learning, not as criticism) what could also have been done.
The paper characterizes gene regulatory interactions in constitutive lamina-associated domains (cLADs) during human adipose stem cell adipogenic differentiation by integrating publicly available enhancer-capture Hi-C (ECHi-C) data with the authors' own ChIP-seq, RNA-seq, ATAC-seq, and FISH datasets. The primary analytical strategy relied on genomic intersection and overlap analyses (BEDTools, BEDOPS, Intervene) combined with a 15-state ChromHMM model to characterize chromatin landscapes; a single inferential test (unpaired two-tailed t-test with Welch's correction) was applied to compare lamin enrichment between cLAD subregions. FISH inter-probe distances and RT-qPCR knockdown expression data were also reported to assess gene-enhancer proximity changes and lamin-dependent gene silencing, respectively.
| Test | Applied to | n | Assumptions |
|---|---|---|---|
| Unpaired two-tailed t-test with Welch's correction | Comparison of LMNA/C ChIP enrichment in H3K4me1-high cLAD regions versus the rest of the LADs (Figure 1c) | — | not stated |
| Threshold-based significance filter (p ≤ 0.01) from published ECHi-C dataset | Selection of significant enhancer-capture Hi-C connections from Madsen et al. (GSE140782) pooled across all differentiation time-points; 18,710 baits and 6,108/7,988 targets identified | 18,710 baits total; 768 within cLADs | na |
| FISH inter-probe distance analysis (test name not stated in available text) | Gene-enhancer proximity for CMKLR1, OTUD1, and LRATD1 loci across D0, D1, D3 of differentiation at the nuclear periphery (Figure 3) | 50 nuclei (100 alleles) per condition | not stated |
| RT-qPCR relative quantification (inferential test not stated) | IL5RA, TRNT1, and CRBN expression after siRNA knockdown of LMNA/C, LMNB1, both, or scrambled control across D0–D6 (Figure 5d) | — | not stated |
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LMNA/C ChIP enrichment between H3K4me1-high cLAD regions and the rest of LADs was compared with an unpaired two-tailed t-test with Welch's correction↳ Could also: A Mann-Whitney U (Wilcoxon rank-sum) test could also be used for this comparison — ChIP log2(ChIP/Input) enrichment values may not be normally distributed across genomic bins; a rank-based non-parametric test does not require a normality assumption and can be more robust for skewed signal distributions commonly observed in ChIP-seq data
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Multiple knockdown conditions (siLMNA/C, siLMNB1, double KD, siScr) were compared for IL5RA expression across multiple days (D3, D6) without a stated multiplicity correction↳ Could also: A one-way ANOVA or two-way ANOVA (condition × time) followed by a post-hoc correction (e.g., Tukey HSD or Benjamini-Hochberg FDR) could also be applied — When several group contrasts are evaluated simultaneously, a family-wise or false-discovery-rate correction reduces the probability that any one comparison appears significant by chance; an ANOVA framework also formally tests whether the time × condition interaction contributes to expression differences
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FISH inter-probe distances were compared across three time-points (D0, D1, D3) at the nuclear periphery, with significance described verbally; the statistical test used is not specified in the available text↳ Could also: A linear mixed-effects model or Kruskal-Wallis test with post-hoc pairwise comparisons (e.g., Dunn's test) could also be used, with allele nested within nucleus as a random effect — Two alleles measured per nucleus are not independent observations; a mixed-effects structure accounts for within-nucleus correlation, and a non-parametric approach accommodates the typically non-normal distribution of FISH distances
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ECHi-C connections from all differentiation time-points in the external dataset were pooled (confounded) before intersection with cLAD coordinates↳ Could also: Time-point-stratified analyses could also be performed, keeping D0, intermediate, and late time-point connections separate before intersection — Pooling across time-points increases the number of detectable connections but loses temporal resolution; stratified analyses would allow assessment of which connections are gained or lost during differentiation, directly linking chromatin topology dynamics to the transcriptomic changes reported
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Bait connection density was summarized as a mean number of connections per bait (27.5 for K4, 14.7 for nonK4) without reporting any measure of dispersion↳ Could also: Reporting SD, IQR, or a 95% CI alongside the mean, or showing the full distribution (e.g., violin or box plot), would also be standard — Connection counts per bait likely span a wide range; a measure of spread communicates whether the means reflect consistent behavior across baits or are driven by a small number of highly connected loci, which is especially relevant for interpreting the biological significance of the difference
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The ChromHMM 15-state model was selected on the basis that it represented non-redundant states concordant with genomic locations, without a quantitative model-selection criterion stated↳ Could also: Information-theoretic criteria (e.g., AIC, BIC) or held-out-data likelihood could also be used to formally compare models with different state numbers — Quantitative model selection criteria provide a reproducible and data-driven basis for choosing the number of chromatin states, making the choice more transparent and facilitating comparison with ChromHMM models in other studies
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Downstream reach in the literature
8 downstream papers · 4 datasetsHow widely the datasets deposited by this paper are reused across the whole literature (Europe PMC), beyond our assessed set. This is a factual dependency map — reusing a public dataset is normal, good science. It is not a judgement on the downstream papers; the only verdict here is this paper's own, with its cited rationale.
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What was reproduced
The exact results taken into scope, with each reported value next to the value our attempt produced.
Assessments & scoring basis
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An automated assessment. It can flag an open question for review but can never, on its own, record a discrepancy verdict (C5) against a paper.
Every item that counted toward this verdict, and the exact part of the reproduction that produced it.
The upstream pipeline reproduces cleanly — total ECHi-C baits match 1:1 (C1 = 18,710) and the central claim holds (K4/nonK4 connection ratio 1.83 vs reported 1.87, ~2×). The deviation is confined to the absolute cLAD-overlap counts (C2–C6, roughly 2× off), which trace to an underspecified cLAD definition (EDD parameters, coverage threshold, replicate/timepoint merge) that is not uniquely reconstructable from the deposited _peaks.bed files — an authors'-side specification gap, not our error and not fabrication (the paper is internally consistent: 225+543=768, 7988/543≈14.7). Net: solid partial reproduction where the qualitative conclusion is confirmed but the exact input cohort cannot be rebuilt from public data.
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