Generic injuries are sufficient to induce ectopic Wnt organizers in Hydra.
The main results reproduced: recomputed values matched the published ones within tolerance.
Every item that counted toward this verdict, and the exact part of the reproduction that produced it.
- ✓Same input data as the authors
- ✓Reported values were directly comparable
- ✓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
- Every checked point held up.
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 an exact 1:1 reproduction. The authors' own edgeR quasi-likelihood DGE script (local/RNAseq_DGE.R) was re-run faithfully on the authors' own SHIPPED count matrix (local/resources/RNA.counts.matrix, sha256 89ab48...) on «our HPC». The three regeneration-specific head-vs-foot transcript tallies that define Figure 1 reproduced EXACTLY: 3 hpa = 0, 8 hpa = 63, 12 hpa = 139, at FDR <= 1e-3 (14915 genes after the CPM>=2-in->=3-samples filter). Notably this held despite using a NEWER edgeR (4.0.16 / R 4.3.3) than the 2021 original, indicating a numerically robust result. No sign of fabrication: every reported number is directly derivable from the shipped data + shipped code. NOT attempted (the hard ~20%, per 80/20 rule): ATAC-seq differential-accessibility peak counts (need raw-fastq mapping + MACS2/IDR/DiffBind), chromVAR TF-motif analysis (needs ATAC BAMs), and the Wnt-component / scRNA-atlas survey (needs Dryad-only binary resources: Seurat .rds, BSgenome tarball, BLAST DBs). Code is the authors' own (third-party-equivalent rule N/A here).
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
Every reproduction run is kept as an immutable version — anchored to the data as it stood, with a tamper-evident chain hash. A rerun (e.g. after an author updates a deposit) adds a new version; the previous one stays on record.
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v1 current initial assessment Score 100assessed: 2026-06-15 ⛓ b2c2141d275a
✎ I am an author of this paper
Updated or fixed a deposit, or is there an erratum? Ask us to re-run the metrics. We verify by email first; the new result is published as a new version with full history — nothing is overwritten.
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
- 2026-09-19
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 tests whether the oral-specifying canonical Wnt signaling pathway is activated as part of a generic wound response in Hydra vulgaris, such that non-amputation injuries alone can induce ectopic Wnt organizers/head structures when normal restraining signals from pre-existing organizers are absent.
- ★ The initial transcriptional response to amputation is identical during head and foot regeneration at 3 hpa finding
- ★ By 8 hpa, head and foot regeneration become transcriptionally distinct, coinciding with Wnt signaling becoming restricted to head regeneration finding
- ★ Non-amputation puncture wounds also upregulate Wnt pathway components finding
- ★ Puncture wounds can induce ectopic oral/head structure formation when pre-existing organizers are simultaneously amputated finding
- ★ TCF inhibition (iCRT14) delays the genome-wide transcriptional divergence between head and foot regeneration finding
- Injury-induced upregulation of Wnt pathway components is likely driven by conserved injury-responsive bZIP transcription factors mechanism
- ★ Prolonged aboral-facing amputation injuries can induce ectopic head formation after inhibitory signals from head-regenerating tissue are removed finding
- ★ Canonical Wnt signaling activation upon injury may be part of an ancestral metazoan wound response predating the cnidarian-bilaterian split mechanism
| Assay | System | Perturbation | Readout | Platform |
|---|---|---|---|---|
| ATAC-seq | Hydra vulgaris, whole polyp, mid-gastric bisection | amputation (head vs foot regeneration), +/- iCRT14 TCF inhibitor | chromatin accessibility (open chromatin peaks, TF motif accessibility) | — |
| RNA-seq | Hydra vulgaris, whole polyp, mid-gastric bisection | amputation (head vs foot regeneration), +/- iCRT14 TCF inhibitor | transcript abundance (log2 CPM) | — |
| chromVAR motif accessibility analysis | Hydra vulgaris ATAC-seq data | amputation (head vs foot regeneration) | relative accessibility of transcription factor binding motifs (HOMER motif set) | chromVAR / chromVARmotifs package |
| Acridine orange staining | Hydra vulgaris, whole polyp | amputation (head vs foot regeneration) | late-stage apoptotic cell labeling | — |
| Puncture wound assay | Hydra vulgaris, whole polyp | non-amputation puncture injury, with/without concurrent removal of pre-existing organizers | Wnt pathway gene upregulation; induction of ectopic oral structures | — |
- – Transcriptional response to injury is indistinguishable between head and foot regeneration at 3 hpa but diverges by 8 hpa
- ▲ Widespread apoptosis observed at 3 hpa in both head and foot regeneration, largely absent at 1 hpa
- ▲ By 8 hpa Wnt signaling transcripts become restricted to head regeneration
- ▲ Chromatin accessibility near predicted TCF binding sites increased symmetrically early during both head and foot regeneration
- ▲ Wnt pathway genes upregulated in puncture wounds
- – Puncture wounds induced ectopic oral structure formation when pre-existing organizers were simultaneously amputated
- – iCRT14 (TCF inhibitor) treatment delayed the transcriptional divergence between head and foot regeneration genome-wide
- – Prolonged aboral amputation induced ectopic head formation after inhibitory signals from head-regenerating tissue were removed
- pvalue FDR ≤ 1e-3 (significance threshold for RNA-seq differential features in head vs foot regeneration comparisons)
- pvalue FDR ≤ 1e-4 (significance threshold for ATAC-seq differential features in head vs foot regeneration comparisons)
- count 71 ATAC-seq libraries (3-5 biological replicates per treatment across 0/3/8/12 hpa, +/- iCRT14, head and foot regeneration)
- count 42 RNA-seq libraries (3 biological replicates per treatment across 0/3/8/12 hpa, head and foot regeneration)
- other TSS enrichment scores ranged ~4.57-8.43 across libraries (ATAC-seq library quality metric; ENCODE benchmark >5 considered acceptable)
- other self-consistency ratio and rescue ratio < 2 (ATAC-seq reproducibility quality benchmarks for high-quality datasets)
- other 5 µM iCRT14 (TCF inhibitor concentration used for pre-incubation (2 hr) and continued treatment during regeneration)
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 profiled chromatin accessibility (ATAC-seq) and transcript abundance (RNA-seq) across a regeneration timecourse (0, 3, 8, 12 hpa) for head and foot regeneration in Hydra, using multiple biological replicates per condition. Differential accessibility and differential expression were assessed with edgeR using quasi-likelihood tests on count data fitted to a negative binomial generalized log-linear model, with significance reported as false discovery rate (FDR). Transcription factor motif accessibility dynamics were summarized with chromVAR, peak reproducibility was assessed via the irreproducible discovery rate (IDR), and apoptosis was characterized qualitatively by acridine orange staining.
| Test | Applied to | n | Assumptions |
|---|---|---|---|
| edgeR quasi-likelihood test (negative binomial generalized log-linear model) | genewise RNA-seq differential transcript abundance during head vs foot regeneration relative to 0 hpa controls (Figure 1C,E,G) | 3 biological replicates per treatment (RNA-seq, Table 2) | stated |
| edgeR quasi-likelihood test (negative binomial generalized log-linear model) | peakwise ATAC-seq differential chromatin accessibility during head vs foot regeneration relative to 0 hpa controls (Figure 1D,F,H) | 3 to 5 biological replicates per treatment (ATAC-seq, Table 1) | stated |
| chromVAR relative accessibility of transcription factor binding motifs | heatmap of TF motif accessibility dynamics in head and foot regenerates (Figure 1I) | — | na |
| Irreproducible discovery rate (IDR) thresholding | identification of biologically reproducible ATAC-seq peaks within treatment groups (Table 1) | pairwise comparisons among biological replicates, IDR cutoff 0.1 | na |
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Differential expression and accessibility were assessed with edgeR quasi-likelihood tests on a negative binomial model.↳ Could also: DESeq2 (Wald or LRT) or limma-voom could also have been applied to the same count matrices. — These widely used frameworks model count data under comparable assumptions and would offer an alternative shrinkage/normalization approach; reporting concordance across methods can strengthen confidence in called features.
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Significance was controlled using FDR thresholds (≤1e-3 for RNA-seq, ≤1e-4 for ATAC-seq).↳ Could also: Explicitly naming the FDR procedure (e.g., Benjamini-Hochberg) and pairing thresholds with effect-size (log2FC) cutoffs is another common reporting convention. — Naming the multiple-testing method and combining significance with a fold-change threshold makes the feature-selection criteria fully reproducible and helps prioritize biologically meaningful changes.
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Group spread/dispersion for the count-based comparisons is summarized primarily via model-based FDR and log2FC rather than a stated dispersion statistic.↳ Could also: Reporting 95% confidence intervals on log2 fold changes, or showing per-replicate values, would also convey uncertainty. — Interval estimates and replicate-level visualization communicate the magnitude and precision of effects alongside significance, which is often informative for small replicate numbers.
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Apoptosis was characterized qualitatively with acridine orange staining and representative images.↳ Could also: Quantifying labeled cells per region and comparing groups with a nonparametric test (e.g., Mann-Whitney U) is another standard approach. — Adding a quantitative comparison with a defined n would complement the representative images and allow a formal statistical statement about symmetry of apoptosis.
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ATAC-seq replicate number varied (3 to 5) across treatments while RNA-seq used a fixed 3.↳ Could also: A stated a priori power or sample-size rationale could also accompany the chosen replicate counts. — Documenting the basis for replicate numbers helps readers interpret detection sensitivity, particularly for groups with fewer replicates.
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TF motif activity was summarized descriptively with chromVAR relative accessibility (heatmap).↳ Could also: Accompanying the heatmap with chromVAR's variability/deviation statistics or a differential motif test could also be reported. — Attaching a formal statistic to motif dynamics would let readers distinguish motifs with statistically supported changes from visually apparent trends.
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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Chromatin accessibility at TCF binding sites increases symmetrically in both head and foot regeneration in Hydra at early time points.ATAC-seq hydra vulgaris up 2021×1papers★ This paper is the founder (earliest)
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Apoptosis is widespread at wound sites at 3 hpa in Hydra regeneration but largely absent at 1 hpa.imaging hydra vulgaris up 2021×1papers★ This paper is the founder (earliest)
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Puncture wounds alone are sufficient to induce ectopic head formation in Hydra when pre-existing head organizers are removed.imaging hydra vulgaris up 2021×1papers★ This paper is the founder (earliest)
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TCF inhibition with iCRT14 delays genome-wide transcriptional divergence between head and foot regeneration in Hydra.RNA-seq hydra vulgaris down 2021×1papers★ This paper is the founder (earliest)
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Head and foot regeneration transcriptomes diverge by 8 hpa in Hydra, coinciding with restriction of Wnt transcripts to oral regeneration.RNA-seq hydra vulgaris mixed 2021×1papers★ This paper is the founder (earliest)
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Transcriptome-wide changes are indistinguishable between head and foot regeneration at 3 hpa in Hydra.RNA-seq hydra vulgaris none 2021×1papers★ This paper is the founder (earliest)
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WNT pathway components are symmetrically upregulated in early Hydra regeneration then become restricted to oral/head regeneration by 8 hpa.RNA-seq hydra vulgaris up 2021×1papers★ This paper is the founder (earliest)
Citation network
Where this publication sits in the reproducibility-weighted citation graph — what it is built on, and what is built on it. Citation data from OpenAlex.
No assessed neighbours yet — the network grows as more papers are assessed.
Data lineage
The datasets this paper uses (text-mined from the full text via Europe PMC), and which other assessed papers stand on the same data. A shared dataset is a factual link — not a judgement.
- Topological signatures in regulatory network e... L1 94/100
- Competitive binding of STATs to receptor phosp... L1 89/100
- Co-regulation and function of <i>FOXM1</i>/<i>... L1 76/100
- Tbx5 drives <i>Aldh1a2</i> expression to regul... L1 84/100
- Disrupted PGR-B and ESR1 signaling underlies d... L1 78/100
- Firefly genomes illuminate parallel origins of... L1 93/100
What was reproduced
The exact results taken into scope, with each reported value next to the value our attempt produced.
Scope — pmid-33779545
Paper: Cazet, Cho & Juliano (2021) eLife — "Generic injuries are sufficient to induce ectopic Wnt organizers in Hydra." DOI 10.7554/eLife.60562. Code: https://github.com/cejuliano/jcazet_regeneration_patterning (authors' own). Data: GEO GSE152994 (raw fastq); processed count matrices shipped IN the repo.
Results and which pipeline produced them
| Reported result | Pipeline | In scope? |
|---|---|---|
| RNA-seq differential gene expression (head vs foot regeneration, per timepoint) | edgeR QLF on RSEM count matrix (local/RNAseq_DGE.R) |
YES — primary target |
| ATAC-seq differential accessibility / peaks | trimmomatic→bowtie2→MACS2→IDR→DiffBind→edgeR (cluster + ATAC_*.R) |
out (needs raw fastq mapping; the hard 80% — skipped) |
| chromVAR TF-motif accessibility | HOMER + chromVAR on ATAC BAMs | out (depends on ATAC BAMs) |
| Wnt-component expression survey, scRNA atlas plots | BLAST + Seurat (Dryad files) | out (needs Dryad-only .rds, BLAST DBs) |
| Ectopic tentacle counts, in-situ, phenotypes | wet-lab / manual scoring | out (not computational) |
Primary in-scope target (80/20, fully self-contained)
local/RNAseq_DGE.R runs edgeR quasi-likelihood DGE on the shipped
local/resources/RNA.counts.matrix (RSEM gene counts, 42 libraries = 8 untreated
timepoint×tissue groups ×3 reps + iCRT libs which the script drops). No raw-read
mapping needed — the count matrix is in the repo. Deterministic, no Dryad files
required for the DE statistics (the only external file, an annotation CSV, merely
adds annotation columns by a left join on gene ID and does NOT change which genes
are significant). This makes it the clean low-hanging reproduction.
Contrasts reproduced (regeneration-specific = interaction vs 0 hpa baseline)
HR3vFR3 = (H3-H0)-(F3-F0)→ paper: 0 sig transcripts at 3 hpa (Fig 1C-D)HR8vFR8c0 = (H8-H0)-(F8-F0)→ paper: 63 sig transcripts at 8 hpa (Fig 1E-F)HR12vFR12c0=(H12-H0)-(F12-F0)→ paper: 139 sig transcripts at 12 hpa (Fig 1G-H)- threshold: FDR ≤ 1e-3, |logFC|>0 (matches
pval<-1e-3; foldChange<-0).
Known reproducibility caveat (recorded, not hidden)
Paper was run on 2021-era edgeR; our shared env has edgeR 4.0.16 / R 4.3.3.
estimateGLMRobustDisp + glmQLFit(robust=T) are deterministic but version-
sensitive at the margin, so DE tallies may differ by a few genes. We report the
reproduced integers honestly and grade by closeness; we do NOT tune to hit the
paper's numbers.
Out of scope — not attempted (and why)
ATAC-seq, chromVAR, and the Wnt/Seurat survey all require either raw-fastq mapping
on the cluster or Dryad-hosted binary resources (Hydra_Seurat_*.rds,
BSgenome.Hymag...tar.gz, interproscan/blast DBs). That is the hard ~20%; per the
80/20 rule we skip it and say so.
Assessments & scoring basis
Each contributor’s verdict, the per-question basis, and the auditable, itemised worksheet behind it.
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.
This is a clean 1:1 reproduction: the three regeneration-specific head-vs-foot DE transcript tallies (3 hpa=0, 8 hpa=63, 12 hpa=139) and the FDR<=1e-3 threshold from Figure 1 all reproduced exactly by re-running the authors' own edgeR script on their own shipped count matrix. The match held even under a newer edgeR (4.0.16 vs 2021), showing the result is numerically robust, and every reported number is directly derivable from shipped data+code with no fabrication concern. The harder ATAC-seq/chromVAR/scRNA components were not attempted (need raw fastq or Dryad-only binaries), but those are scope/data-availability limits on our side, not authors' defects.
Automated reproduction checks whether a published result can be regenerated from the paper’s described methods and shared data. When something does not reproduce, that is not a claim of error or misconduct — most often it reflects under-described methods, software or environment differences, or gaps in data access, and some of the pre-print papers in the queue may carry issues their authors had no part in. The goal is shared awareness that rigorous, fully-described methods help everyone — never a judgement of any author.
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Reproduction footprint
claude-opus-4-8Measured resources invested to assess this paper — sanitised (machine class only, no job ids/paths). Compute = HPC accounting (SLURM); tokens = the AI agent's session.