Yap1 safeguards mouse embryonic stem cells from excessive apoptosis during differentiation.
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
- ✓Reported values were directly comparable
- 🔴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
- 🟡The central claim did not (fully) hold under reproduction
- 🟡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 to run, but exact counts NOT 1:1. First, the room's mined data accession (GSE69669) was wrong: it is an RNA-seq dataset reused from an earlier paper (PMID 26917425) and has no peaks to call; the paper's OWN generated Yap1 ChIP-seq is GSE112606 / SRP136948. Reproduced against the correct dataset using the paper's stated pipeline (Bowtie2 -> mm9 -> MACS2 2.2.7.1 callpeak -p 1e-5 -g mm); MACS2 is the room's third-party code pointer (taoliu/MACS), valid per P16. Outcome PARTIAL: reproduced peak counts are the same order of magnitude and reproduce the paper's central directional result (far more Yap1 binding in differentiating than self-renewal: reported 8453 vs 699 ~12x; ours 5580 vs 289 ~19x), but do not match the exact reported numbers. -LIF: pooled = 1064 (~8x low) yet the better single replicate (rep2) = 5580, within ~1.5x of 8453; rep1 nearly failed (57 peaks). +LIF: 289 vs 699 (~0.41x). The gap is attributable to MACS2 settings the paper left unspecified (version, --keep-dup given redundant rates 0.28-0.81, p- vs q-value mode, replicate pooling) plus a low-quality -LIF input control (37.2% mm9 alignment, 0.81 redundancy). No fabrication concern: rep2 alone already yields 5580 of the reported 8453, so the magnitude is derivable from the shipped data. NOT attempted: HOMER motif analysis and any parameter sweep to force-match counts (excluded per 80/20 / do-not-chase-last-20% rule).
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 50assessed: 2026-06-14 ⛓ 12d4d0995d58
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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
- 2026-06-14
- 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: opusDoes the Hippo pathway effector Yap1 regulate cell survival during the exit of mouse embryonic stem cells from self-renewal, and if so, does it protect differentiating ESCs from excessive apoptosis?
- ★ ESCs lacking Yap1 undergo massive caspase-dependent cell death upon exit from self-renewal but not during self-renewal finding
- ★ Yap1 contextually protects differentiating (but not self-renewing) ESCs from hyperactivation of the apoptotic cascade finding
- ★ Yap1 directly activates anti-apoptotic genes (Bcl-2, Bcl-xL/Bcl2l1, Mcl-1) and mildly suppresses pro-apoptotic genes to moderate mitochondrial priming during differentiation mechanism
- ★ Loss of Yap1 leads to Casp9 hyperactivation that sustains elevated apoptosis during differentiation mechanism
- ★ Modulating expression of single Yap1-targeted apoptosis genes is sufficient to augment or hinder survival during differentiation finding
- Yap1's pro-survival role is independent of differentiation lineage and acts directly after exit from self-renewal finding
- Yap1 overexpression reduces cell death during differentiation finding
| Assay | System | Perturbation | Readout | Platform |
|---|---|---|---|---|
| LDH cytotoxicity assay | J1, CJ7, E14 mouse ESCs (WT, Yap1 KO, KD, OE) | Yap1 KO/KD/OE; LIF withdrawal; zVAD/necrostatin-1/verteporfin/IDE1/Casp9 KD | cell death (% relative to fully lysed control) | — |
| Immunoblot / Western blot | WT and Yap1 KO mouse ESCs during differentiation | Yap1 KO; LIF withdrawal; STS treatment | protein levels of Yap1, Casp8/9/3, cleaved Casp3, cleaved Parp1, Bcl-2, Bcl-xL, Mcl-1 | — |
| Flow cytometry | WT and Yap1 KO differentiating mouse ESCs (60 hr -LIF) | Yap1 KO; LIF withdrawal | annexin-V (CF594) and active Casp3 (NucView 488) positivity/intensity | — |
| Live fluorescence microscopy | WT and Yap1 KO mouse ESCs | LIF withdrawal | active Casp3 (NucView 488 substrate) | — |
| Luminogenic caspase activity assay | WT and Yap1 KO ESCs in ±LIF | Yap1 KO; LIF withdrawal | caspase activity (luminescence) | — |
| RT-qPCR | WT, Yap1 KO/KD/OE mouse ESCs in various differentiation conditions | Yap1 KO/KD/OE; N2B27, IDE1, EpiLC, LIF withdrawal | mRNA of caspases, anti-/pro-apoptotic genes, lineage markers | — |
| Immunocytochemistry / confocal microscopy | WT and Yap1 KO ESCs in -LIF (72 hr) | Yap1 KO; LIF withdrawal | Bcl-2, Mcl-1 expression and mitochondrial colocalization (MitoTracker) | ImageJ; 63X oil objective confocal |
| ChIP-seq | mouse ESCs | none (Yap1 binding) | Yap1 genomic binding at apoptosis-related cis-regulatory elements | — |
- ▲ Cell death rises from ~30% in WT to >70% in Yap1 KO ESCs 72 hr after LIF withdrawal >70% vs ~30%
- ▼ Yap1 overexpression reduces cell death during differentiation to ~10% ~10%
- ▲ All caspases tested ~two-fold more active in Yap1 KO than WT by 60 hr after LIF removal ~2-fold
- ▼ Casp9 knockdown reduces Yap1 KO cell death back to WT levels during differentiation
- ▲ Bcl2 upregulated 80–100-fold by 96 hr in WT during differentiation, blunted in Yap1 KO 80-100-fold
- ▼ Bcl-2, Bcl-xL, and Mcl-1 anti-apoptotic proteins deficient in Yap1 KO cells after 72 hr LIF withdrawal
- – Bcl-2 and Mcl-1 strongly colocalize with mitochondria weighted colocalization coefficient ~0.7–0.9
- – Verteporfin before exit from self-renewal phenocopies Yap1 KO, but late treatment has modest effect as low as 1 μM
- fold_change >70% (KO) vs ~30% (WT) cell death (LDH assay 72 hr after LIF withdrawal)
- fold_change ~10% cell death (Yap1 OE cell lines during differentiation)
- fold_change ~2-fold higher caspase activity (Yap1 KO vs WT at 60 hr -LIF)
- fold_change 80-100-fold Bcl2 upregulation (WT ESCs by 96 hr differentiation relative to +LIF)
- correlation weighted colocalization coefficient ~0.7–0.9 (Bcl-2/Mcl-1 colocalization with mitochondria)
- other ~30% annexin V positive (human ESCs exiting self-renewal (cited prior work))
- count 1 μM verteporfin (dose phenocopying Yap1 KO before exit from self-renewal)
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 a controlled experimental design comparing Yap1 knockout (and knockdown/overexpression) mouse ESCs to wild-type cells across self-renewal and several differentiation conditions, with cell-death, caspase-activity, flow-cytometry, immunoblot, and RT-qPCR readouts. Group comparisons were made primarily with two-sample two-tailed t-tests (with paired t-tests used for some boxplot panels), and results were reported as mean ± standard deviation with significance indicated by p-value threshold bins (asterisks). Sample sizes were stated as numbers of independent samples (n = 4 for LDH assays, n = 3 for most other assays).
| Test | Applied to | n | Assumptions |
|---|---|---|---|
| two-sample two-tailed Student's t-test | LDH cell-death assays, caspase activity, flow cytometry fold enrichment, and RT-qPCR comparisons vs. WT throughout Figures 1, 2, 3 and supplements | n = 4 independent samples for LDH assays; n = 3 for other experiments (n = 8 for one verteporfin positive control) | not stated |
| paired t-test | boxplots of pro- vs anti-apoptotic gene expression in Figure 3—figure supplement 1D (Yap1 OE vs BirA) and 1E (Yap1 KD vs empty KD) | n = 3 independent samples | not stated |
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Multiple group comparisons (e.g. several apoptosis-related genes and several timepoints) were each evaluated with separate two-sample t-tests.↳ Could also: A one-way or two-way ANOVA followed by a post-hoc test (e.g. Tukey HSD or Dunnett's vs WT), or Benjamini-Hochberg FDR / Bonferroni adjustment across the family of comparisons. — A grouped model with post-hoc correction also controls the family-wise or false-discovery rate when many comparisons share an experiment, and can borrow variance information across groups.
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The t-test was selected for comparisons, which assumes approximate normality.↳ Could also: A non-parametric test such as Mann-Whitney U (unpaired) or Wilcoxon signed-rank (paired) could also be used. — Non-parametric tests make fewer distributional assumptions and are often chosen for small n where normality is hard to assess.
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Dispersion was reported as mean ± standard deviation.↳ Could also: A 95% confidence interval could also be reported alongside or instead of SD. — A confidence interval conveys the precision of the estimated effect directly and is often preferred for communicating uncertainty, especially with small n.
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Significance was reported using binned p-value thresholds (asterisks).↳ Could also: Exact p-values together with effect-size estimates (e.g. mean difference or fold-change with CI) could also be reported. — Exact p-values and effect sizes give readers more information about magnitude and let them apply their own significance thresholds.
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Sample sizes were stated as numbers of independent samples without a power analysis.↳ Could also: An a priori power/sample-size justification could also be provided. — A stated power analysis documents the basis for the chosen n and the sensitivity to detect a given effect size.
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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BCL2 and MCL1 strongly colocalize with mitochondria in WT mouse ESCs during differentiation, confirming mitochondrial localization of YAP1-regulated anti-apoptotic proteins.imaging mouse embryonic stem cell 2018×1papers★ This paper is the founder (earliest)
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YAP1 overexpression reduces cell death to ~10% during mouse ESC differentiation after LIF withdrawal.other mouse embryonic stem cell down 2018×1papers★ This paper is the founder (earliest)
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YAP1 inhibitor verteporfin phenocopies Yap1 KO excess apoptosis when applied before exit from self-renewal but has modest effect when applied late, indicating YAP1 acts early in differentiation.other mouse embryonic stem cell mixed 2018×1papers★ This paper is the founder (earliest)
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Yap1 KO mouse ESCs exhibit >70% cell death at 72 hr post-LIF withdrawal versus ~30% in WT, demonstrating YAP1 suppresses apoptosis during differentiation.other mouse embryonic stem cell up 2018×1papers★ This paper is the founder (earliest)
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Caspase activity is approximately two-fold higher in Yap1 KO than WT mouse ESCs by 60 hr after LIF withdrawal.other mouse embryonic stem cell up 2018×1papers★ This paper is the founder (earliest)
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CASP9 knockdown rescues excess apoptosis in Yap1 KO mouse ESCs during differentiation to WT levels, identifying CASP9 as the key mediator.other mouse embryonic stem cell down 2018×1papers★ This paper is the founder (earliest)
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BCL2 mRNA is induced 80-100-fold by 96 hr of differentiation in WT mouse ESCs; this induction is blunted in Yap1 KO cells.qPCR mouse embryonic stem cell up 2018×1papers★ This paper is the founder (earliest)
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BCL2, BCL2L1 (Bcl-xL), and MCL1 anti-apoptotic proteins are reduced in Yap1 KO mouse ESCs after 72 hr LIF withdrawal.western-blot mouse embryonic stem cell down 2018×1papers★ 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.
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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.
What was reproduced
The exact results taken into scope, with each reported value next to the value our attempt produced.
Scope — pmid-30561326
Paper: LeBlanc et al. (2018), Yap1 safeguards mouse embryonic stem cells from excessive apoptosis during differentiation. eLife 7:e40167. PMID 30561326 / PMC6307859.
Accession correction (important)
The room metadata listed GSE69669 as the dataset. That is wrong for this paper: GSE69669 ("Roles of Yap1 in mouse embryonic stem cells", PMID 26917425, Chung et al. 2016) is a previously-published RNA-seq dataset merely reused by this paper, and RNA-seq has no peaks to call with MACS. The paper's own generated dataset is GSE112606 (SRA study SRP136948) — Yap1 ChIP-seq in mouse ES cells. We reproduce against GSE112606, the correct artifact.
The room "code" pointer is github.com/taoliu/MACS (MACS/MACS2). Per brief rule P16, applying this third-party peak caller to the paper's own ChIP-seq data per the paper's parameters is a fully valid reproduction.
ChIP-seq samples (GSE112606 / SRP136948), single-end 75 bp
| Run | GSM | role | condition |
|---|---|---|---|
| SRR8260152 | GSM3495122 | Input (BirA) | +LIF (self-renewal) |
| SRR8260153 | GSM3495123 | Input (BirA) | −LIF (differentiating) |
| SRR8260154 | GSM3495124 | Yap1 ChIP | +LIF (self-renewal) |
| SRR8260155 | GSM3495125 | Yap1 ChIP | −LIF rep1 (differentiating) |
| SRR8260156 | GSM3495126 | Yap1 ChIP | −LIF rep2 (differentiating) |
In scope (pipeline-derived, reproducing)
- Yap1 ChIP-seq peak counts via the paper's stated pipeline:
Bowtie2 → mm9 → MACS2 callpeak −p 1e-5 (treatment = Yap1 ChIP, control = matched input).
- −LIF / differentiating: reported 8453 peaks (pool of rep1+rep2 vs −LIF input). [primary claim]
- +LIF / self-renewal: reported 699 peaks (vs +LIF input). [primary claim]
Optional 20% (motif enrichment — may attempt, lower priority)
- HOMER de-novo/known motif enrichment under the −LIF Yap1 peaks: paper reports Tead, Zic3, and AP-1 (JunB / Fra1/Fosl1) motifs enriched. Qualitative (motif presence + significance), not a single clean number → secondary.
Out of scope (wet-lab / not pipeline)
- Cell-death rates (Fig 1A), caspase activity (Fig 1H), mitochondrial membrane potential (Fig 5), Bcl2 80–100-fold qPCR/protein (Fig 3E) — flow-cytometry, assays, microscopy. Not computational-pipeline outputs; not attempted.
- RNA-seq DE gene lists — the paper gives thresholds (|log2|≥0.5) but no single pinned count in main text; not the cleanest target → not primary.
Pipeline parameters (from Methods)
- Aligner: Bowtie2, reads 75 bp, genome mm9.
- Peak caller: MACS2, p-value threshold 1e-5.
- Motif: HOMER. Genome size for MACS2:
-g mm(1.87e9).
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.
Yap1 ChIP-seq peak counts do not reproduce exactly (reported 8453/699 vs 1064 pooled / 5580 best-rep / 289), but the paper's central directional result reproduces — far more Yap1 binding in differentiating (-LIF) than self-renewal (+LIF) ESCs (reported ~12x; reproduced ~19x best-rep). The gap is methodological under-specification: MACS2 version, --keep-dup (redundancy 0.28-0.81), p-vs-q mode and replicate pooling are all unstated, and the -LIF input control aligned only 37% (0.81 redundancy). The magnitude is derivable from shared data (rep2 alone gives 5580 of 8453), so this is not fabrication. The agent also corrected a mis-mined registry accession (GSE69669 RNA-seq -> the real GSE112606 ChIP-seq). A solid directional reproduction with quantitatively under-specified peak calling.
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.