{ "version": 7, "date": "2026-10-13", "qualifiedFreshStudies": 0, "benchmarkRun": false, "metrics": null, "headline": "Supplementary files checked; no fresh human UPF1 RNA/RPF holdout qualified.", "summary": "The strongest new lead has real human RNA/RPF counts, but a different perturbation. M5, ExtraTrees and ridge have not been scored on these sources. No new superiority claim is supported.", "cases": [ { "id": "gigyf", "title": "Zinshteyn 2021 \u00b7 PRJNA675773", "status": "Assays present; UPF1 perturbation absent", "evidence": "Supplement S5 has 12 monosome/RPF and 12 RNA sample columns: WT, EIF4E2-loss and GIGYF2-loss, each with DMSO or SMG1 inhibitor and two replicates. All 36 archive libraries match these conditions (12 RNA, 12 monosome, 12 disome). No UPF1-depleted arm appears.", "scope": "A possible separate SMG1-inhibition transfer study, subject to its own qualification. It cannot be relabeled as a UPF1-depletion holdout.", "source": "https://pmc.ncbi.nlm.nih.gov/articles/PMC8555832/", "supplement": "https://journals.plos.org/plosgenetics/article/file?id=10.1371/journal.pgen.1009813.s013&type=supplementary" }, { "id": "musaev", "title": "Musaev 2024 \u00b7 SRP495209 / PRJNA1087706", "status": "Human UPF1 RNA present; matched RPF absent", "evidence": "All eight deposited libraries are RNA-seq: control, UPF1, SMG6 and G3BP1 perturbations in two replicates. The paper\u2019s ribosome-profiling analysis uses a previously published zebrafish dataset; it is not a human UPF1 RNA/RPF pair.", "scope": "The human RNA response cannot alone define the M5 translation-efficiency response.", "source": "https://pmc.ncbi.nlm.nih.gov/articles/PMC11259039/", "supplement": "https://www.ebi.ac.uk/ena/browser/view/PRJNA1087706" }, { "id": "karousis", "title": "Karousis 2020 \u00b7 supplementary/source-data trail", "status": "Reuses the previously audited GSE143301 sample family", "evidence": "The article links HeLa RPF samples GSM4256659\u201361 and GSM4256665\u201367, plus UPF1 total-RNA samples GSM4407914\u201316. These are the previously checked control-RPF and UPF1-RNA records; no matched UPF1 RPF arm is established. Supplementary source data do not make this a new study.", "scope": "Article methods and supplementary inventory reviewed; a new matched experiment was not found.", "source": "https://pmc.ncbi.nlm.nih.gov/articles/PMC7431590/", "supplement": "https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE143301" }, { "id": "vendramin", "title": "Vendramin 2026 \u00b7 immunopeptidome study", "status": "No qualifying UPF1 RNA/RPF pair identified", "evidence": "Data availability points to cell-line RNA-seq/TCR-seq (PRJNA1263526), controlled-access patient RNA data, and a Zenodo figure-data deposit. Reviewed methods focus on SMG1 perturbation, RNA and immunopeptidomics; they do not establish the required UPF1 RNA/RPF contrast.", "scope": "Article data availability and supplementary inventory were checked. The linked Zenodo record could not be retrieved (API timeout and web-access error). Figure-data archives were not opened and restricted patient data were not requested.", "source": "https://pmc.ncbi.nlm.nih.gov/articles/PMC7619149/", "supplement": "https://doi.org/10.5281/zenodo.10599407" } ], "catalogueSearches": [ { "name": "BioStudies exact Boolean search", "query": "UPF1 AND (\"ribosome profiling\" OR \"Ribo-seq\")", "returnedRecords": 10, "totalHits": 10, "source": "https://www.ebi.ac.uk/biostudies/studies?query=UPF1%20AND%20(%22ribosome%20profiling%22%20OR%20%22Ribo-seq%22)" }, { "name": "Zenodo exact Boolean search", "query": "UPF1 AND (\"ribosome profiling\" OR \"Ribo-seq\")", "returnedRecords": 0, "totalHits": 0, "source": "https://zenodo.org/search?q=UPF1%20AND%20(%22ribosome%20profiling%22%20OR%20%22Ribo-seq%22)" } ], "nextAction": "The direct UPF1 benchmark needs a new matched human study or an unreleased UPF1 RPF arm from an independent lab. A request checklist is available below. The SMG1 dataset could answer a separate pathway-transfer question; a positive score there would not prove UPF1-specific superiority.", "limitations": [ "This is a targeted search, not proof that an eligible dataset does not exist. Repository indexing may miss supplementary content.", "S5 was streamed to inspect and save its column header only. No gene-response rows were analyzed; supplementary captions and sample metadata were inspected.", "BioStudies\u2019 initial broad query returned an approximate 14,324 hits; only the first 100 metadata records were retrieved. The explicit Boolean query returned 10 records. These are not counts of independent eligible studies.", "The initial Zenodo request exceeded its anonymous page-size limit; retrying with 25 succeeded with zero results. The paper-linked Zenodo record was attempted separately but could not be retrieved; this access gap remains.", "No model fitting, new prediction run, accuracy benchmark or runtime benchmark was performed. Existing v4 evidence and checkpoints are unchanged." ], "s5SampleColumns": [ "tx_id", "WT_DMSO_1_mono", "WT_DMSO_2_mono", "WT_SMG1i_1_mono", "WT_SMG1i_2_mono", "eIF4E2_4_DMSO_1_mono", "eIF4E2_4_DMSO_2_mono", "eIF4E2_4_SMG1i_1_mono", "eIF4E2_4_SMG1i_2_mono", "GIGYF2_4_DMSO_1_mono", "GIGYF2_4_DMSO_2_mono", "GIGYF2_4_SMG1i_1_mono", "GIGYF2_4_SMG1i_2_mono", "WT_DMSO_1_RNA", "WT_DMSO_2_RNA", "WT_SMG1i_1_RNA", "WT_SMG1i_2_RNA", "eIF4E2_4_DMSO_1_RNA", "eIF4E2_4_DMSO_2_RNA", "eIF4E2_4_SMG1i_1_RNA", "eIF4E2_4_SMG1i_2_RNA", "GIGYF2_4_DMSO_1_RNA", "GIGYF2_4_DMSO_2_RNA", "GIGYF2_4_SMG1i_1_RNA", "GIGYF2_4_SMG1i_2_RNA", "gene_id", "gene_name" ], "archiveSamples": [ { "run_accession": "SRR13016595", "study_accession": "PRJNA675773", "sample_accession": "SAMN16722275", "experiment_accession": "SRX9467540", "scientific_name": "Homo sapiens", "sample_title": "Human sample from Homo sapiens", "experiment_title": "Illumina HiSeq 2500 sequencing: cBZ090_eIF4E2-4_SMG1i_2_mono", "library_name": 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A) Flow cytometry of HEK293FT cells lentivirally transduced with indicated plasmids. tagBFP-tdTomato is the full reporter in Fig 1A with a UGG tryptophan codon at the PTC postion (Same as S1B Fig ). B) Western blot of lysates from HEK293FT cells transduced with lentivirus containing reporters with the indicated stop codons, or UGG codon encoding tryptophan. Serial dilutions of UGG codon are also included. (PDF) Click here for additional data file.", "files": [ "pgen.1009813.s001.pdf" ] }, { "label": "S2 Fig", "description": "Validation of NMD and readthrough measurements with reporter. A) Flow cytometry of monoclonal K562 cells expressing reporter with different stop codon contexts (or a UGG tryptophan codon). B) Flow cytometry of monoclonal K562 cells expressing reporter with AQP4 stop context PTC, treated with SMG1i or G418 for 24 hours. C) Same as panel B, for OPRL1 PTC reporter. D) Histograms of tdTomato/tagBFP signal for flow cytometry data from Fig 1E and 1F . (PDF) Click here for additional data file.", "files": [ "pgen.1009813.s002.pdf" ] }, { "label": "S3 Fig", "description": "Validation of select screen hits. A) Volcano plot of fold enrichment and false discovery rate for CRISPR guides sequenced from cells with decreased tdTomato/tagBFP ratio compared to the input population for the AQP4 reporter. B) Volcano plot of fold changes and adjusted t-test p values for validation of select hits with the UGAC reporter. Each point is the relative mean tagBFP fluorescence (with average of nontargeting set to 1) of 2 or 3 separate lentiviral infections each with 2 separate guides (except for UPF1 and ETF1 controls, for which a single guide was used). Each t-test compares these 4 or 6 values against the 6 values for the nontargeting guides (2 separate guides in 3 separate infections). See S3 Table for individual values. The mean values include all cells in the sample, and therefore are affected by variations in transduction efficiency, CRISPR efficiency and knockout viability in addition to the magnitude of the effect on the reporter from the mutation. C) Histograms of tagBFP (blue line) or tdTomato (vermilion line) signal from flow cytometry of reporter lines lentivirally transduced with nontargeting guide (gray dotted line) or targeting guide (solid line), for reporter lines containing UGAC stop codon, AQP4 stop context, or no PTC (UGGC). SNRPF and AQR are splicing factors, RBM8A and EIF4A3 are core components of the EJC, and RPLP0 is a protein of the large ribosomal subunit. These samples were measured at 4 days post infection. D) Coverage of sequencing reads mapping to reporter from nanopore direct RNA sequencing of HEK293FT cells transfected with UGAC or AQP4 reporter plasmid. Fraction of reads at a position with a GU-AG splice junction are indicated as arcs, for introns with at least 2 supporting reads and making up at least 1% of reads spanning the intron ends. Reporter diagram indicates tagBFP and tdTomato coding regions as well as start and stop codons. E) Coverage of uniquely mapping paired-end RNA-seq reads mapping to integrated reporter in K562 reporter line treated with DMSO or SMG1i. Number of reads at a position with a GU-AG splice junction are indicated as arcs, for introns with at least 5 supporting reads. The density increases from 5\u2032 to 3\u2032 because nanopore direct RNA sequencing starts from the polyA tail and drops off as it proceeds to the 5\u2032 end. The 3\u2032 UTR region shares sequence with the Cas9:Blast transgene in these lines, causing some RNA-seq reads from that transgene to spuriously map here and likely causing a dampened apparent response to SMG1i treatment in this region. (PDF) Click here for additional data file.", "files": [ "pgen.1009813.s003.pdf" ] }, { "label": "S4 Fig", "description": "Analysis of EJC component knockdown RNA-seq from ENCODE. A) Box plot of fold changes in mRNA levels for EIF4A3 or MAGOH knockdowns in HepG2 or K562 cells from ENCODE. Transcript sets are divided into all transcripts, empirically determined NMD targets with or without uORFs, and the subset of these NMD targets with 3\u2032 UTR introns. Transcript counts for each set are indicated at left, and p values from Mann-Whitney U-test are indicated. +/- indicates that transcripts both with and without uORFs are included in the transcript set. B) Transcripts per million (TPM) from ENCODE RNA-seq data in Hep G2 cells for a subset of transcripts encoding NMD factors. T-test p values are indicated as asterisks. C) Same as panel B, for K562 cells. Note that in this panel, control 1 is the matched control for eIF4A3 knockdown, while control 2 is the matched control for MAGOH knockdown. (PDF) Click here for additional data file.", "files": [ "pgen.1009813.s004.pdf" ] }, { "label": "S5 Fig", "description": "The effect of GIGYF2/EIF4E2/RACK1 on the reporter does not depend on ZNF598, EDF1 or the 2A peptide sequences. A) As in Fig 2C , with guides against EDF1 added. Note that all samples except EDF1 are the same ones as in Fig 2C , as these electroporations were performed simultaneously. B) tagBFP signal from flow cytometry of reporter lines transduced with nontargeting guide (NT1) or guides against indicated genes for reporter line containing UGAC stop codon. These samples were measured at 6 days post infection. C) Western blot of cell lines expressing nontargeting guides or a guide against ZNF598 from the same infections used for S3B Fig . Lysates were collected on the same day that the flow cytometry was performed. D) TagBFP intensities for independently generated reporter cell line with the 2A peptide upstream of the PTC disabled by mutating the terminal PGP residues to AGA. Electroporation and flow cytometry were performed in parallel with samples in panel A. Note that differences in signal intensity between the lines are likely due to differences in integration site and number, as well as stochastic variation in the monoclonal selection process. (PDF) Click here for additional data file.", "files": [ "pgen.1009813.s005.pdf" ] }, { "label": "S6 Fig", "description": "Generation and characterization of GIGYF2/EIF4E2 knockout lines. A) Histograms of tagBFP signal from flow cytometry of UGAC reporter lines infected with nontargeting guide (dotted gray line) or targeting guide (solid blue line) 42 days after infection, and 12 days after sorting for 5000 cells with increased tagBFP signal. The polyclonal populations expressing GIGYF2 and EIF4E2 guides were used for further experiments, and for production of monoclonal lines. B) tagBFP signal from WT UGAC reporter line, and GIGYF2\u0394 or EIF4E2\u0394 polyclonal or monoclonal lines. C) Allele frequencies in monoclonal or polyclonal GIGYF2\u0394 and EIF4E2\u0394 lines, as assessed by high-throughput sequencing of the CRISPR target site. D) (left)UV absorbance trace from 10\u201350% sucrose density gradient fractionation of WT and GIGYF2\u0394 or EIF4E2\u0394 polyclonal lines. (right) Quantification of ratio of polysome area to 40S+60S+80S area (normalized to WT sample processed on same day) from 2 independent replicates. E) Median tagBFP intensities for indicated WT or EIF4E2\u0394/GIGYF2\u0394 cell lines, either untreated (black circles), or treated with SMG1i for 24 hours (green circles). Biological replicate treatments were performed on 3 separate days. F) Quantification of automated western blotting of tagBFP relative to \u03b2-actin, for indicated WT or EIF4E2\u0394/GIGYF2\u0394 cell lines, either untreated (black circles), or treated with 300nM SMG1i for 24 hours (green circles). Biological replicates were collected and blotted on 2 separate days and normalized to the untreated WT sample within each day. These samples are distinct from the ones used for flow cytometry. Dotted lines show the means for the untreated and SMG1i treated WT samples. Error bars show standard deviation. P value computed by two-tailed ratio paired t-test. All unmarked comparisons had p>0.05. (PDF) Click here for additional data file.", "files": [ "pgen.1009813.s006.pdf" ] }, { "label": "S7 Fig", "description": "Ribosome occupancy changes in GIGYF2\u0394 or EIF4E2\u0394 cells. A) Box plot of fold changes in mRNA levels, ribosome footprints and ribosome occupancy (ratio of ribosome/mRNA) in EIF4E2\u0394 (green), GIGYF1\u0394/GIGYF2\u0394 (blue), or ZNF598\u0394 HEK293T cells, relative to WT cells, for all transcripts (dark bars), empirically determined NMD targets (faded bars), and the subset of these targets with 3\u2032 UTR introns (faded hatched bars). Transcript counts for each set are indicated at left, and p values from Mann-Whitney U-test are indicated. Raw data are from [ 57 ]. B) Comparison of fold changes in mRNA levels with fold changes in ribosome footprints for all quantified transcripts. Transcripts passing false discovery rate cutoff of 0.1 for changes in ribosome occupancy are shaded in black. Transcripts mentioned in the text are labelled in green. C) Venn diagram of transcripts passing cutoff from Fig 5A . P values for overlaps were computed by hypergeometric test, using a background set of 9,735 transcripts with computable fold changes in all datasets. D) Cumulative distribution of ribosome densities in EIF4E2\u0394 cells for NMD targets without (-EJC, solid orange line) or with (+EJC, dashed orange line) 3\u2032 UTR introns. The number of transcripts in each set, and p value for Mann-Whitney U test against all transcripts (black line) is indicated in parentheses. The vertical black dotted line indicates the ribosome occupancy of the UGAC reporter. E) Same as panel D, but for the transcripts with increased ribosome occupancy in EIF4E2\u0394 (green line) or GIGYF2\u0394 (blue line). (PDF) Click here for additional data file.", "files": [ "pgen.1009813.s007.pdf" ] }, { "label": "S8 Fig", "description": "Physical interaction between UPF1 and GIGYF2\u2022EIF4E2. A) Flow cytometry analysis of rescue of EIF4E2\u0394 or GIGYF2\u0394 UGAC reporter polyclonal lines by electroporation of the GIGYF2:EIF4E2 coexpression plasmid used in Fig 6A\u20136E , with CRISPR guide sites mutated. B) Mass spectrometry analyses of StrepII-GIGYF2 immunoprecipitates with or without SMG1i treatment. Scatter plot showing log2(LFQ) intensity of proteins identified under SMG1i (y-axis) and untreated (x-axis) regimes. Values are from a single replicate of each sample. (PDF) Click here for additional data file.", "files": [ "pgen.1009813.s008.pdf" ] }, { "label": "S1 Table", "description": "UGAC NMD screen results. Ranking, average log2 enrichments and false discovery rates for each gene in the UGAC NMD screen, as computed by MAGeCK. (TXT) Click here for additional data file.", "files": [ "pgen.1009813.s009.txt" ] }, { "label": "S2 Table", "description": "AQP4 NMD screen results. Ranking, average log2 enrichments and false discovery rates for each gene in the AQP4 NMD screen, as computed by MAGeCK. (TXT) Click here for additional data file.", "files": [ "pgen.1009813.s010.txt" ] }, { "label": "S3 Table", "description": "Validation of select screen hits by lentiviral infection and flow cytometry. Up to 2 guides per gene were used, in up to 3 replicates. Each dated column represents the average log2 tagBFP fluorescence for cells infected with that guide, normalized to the average of 2 nontargeting guides (NT). Blank cells mean that no infection for that guide was performed on that day. For each gene, the average of all days and guides is computed, as well as the t-test p value compared to the nontargeting guides, the rank of that gene by p-value, the Benjamini-Hochberg corrected p-value, and the -log of that value, which was used for plotting S2B Fig . (TSV) Click here for additional data file.", "files": [ "pgen.1009813.s011.tsv" ] }, { "label": "S4 Table", "description": "Sequencing library sizes. Counts of transcript-mapping reads from ribosome profiling and RNA-seq datasets, as obtained from Salmon output. (TXT) Click here for additional data file.", "files": [ "pgen.1009813.s012.txt" ] }, { "label": "S5 Table", "description": "Read counts for all ribosome profiling and RNA-seq datasets for all transcript isoforms, as computed by Salmon. (TSV) Click here for additional data file.", "files": [ "pgen.1009813.s013.tsv" ] }, { "label": "S6 Table", "description": "TPMs (transcripts per million) for all ribosome profiling and RNA-seq datasets for all transcript isoforms, as computed by Salmon. (TSV) Click here for additional data file.", "files": [ "pgen.1009813.s014.tsv" ] }, { "label": "S7 Table", "description": "Log2 fold changes in RNA-seq (mRNA) and ribosome footprints (RPF), as well as ribosome occupancy (TE), and corresponding p values (pvalue.adjust) for all transcripts as computed by Xtail. For EIF4E2\u0394 vs WT in DMSO. (TSV) Click here for additional data file.", "files": [ "pgen.1009813.s015.tsv" ] }, { "label": "S8 Table", "description": "Log2 fold changes in RNA-seq (mRNA) and ribosome footprints (RPF), as well as ribosome occupancy (TE), and corresponding p values (pvalue.adjust) for all transcripts as computed by Xtail. For EIF4E2\u0394 vs WT in SMG1i. (TSV) Click here for additional data file.", "files": [ "pgen.1009813.s016.tsv" ] }, { "label": "S9 Table", "description": "Log2 fold changes in RNA-seq (mRNA) and ribosome footprints (RPF), as well as ribosome occupancy (TE), and corresponding p values (pvalue.adjust) for all transcripts as computed by Xtail. For EIF4E2\u0394 in SMG1i vs WT in DMSO. (TSV) Click here for additional data file.", "files": [ "pgen.1009813.s017.tsv" ] }, { "label": "S10 Table", "description": "Log2 fold changes in RNA-seq (mRNA) and ribosome footprints (RPF), as well as ribosome occupancy (TE), and corresponding p values (pvalue.adjust) for all transcripts as computed by Xtail. For GIGYF2\u0394 vs WT in DMSO. (TSV) Click here for additional data file.", "files": [ "pgen.1009813.s018.tsv" ] }, { "label": "S11 Table", "description": "Log2 fold changes in RNA-seq (mRNA) and ribosome footprints (RPF), as well as ribosome occupancy (TE), and corresponding p values (pvalue.adjust) for all transcripts as computed by Xtail. For GIGYF2\u0394 vs WT in SMG1i. (TSV) Click here for additional data file.", "files": [ "pgen.1009813.s019.tsv" ] }, { "label": "S12 Table", "description": "Log2 fold changes in RNA-seq (mRNA) and ribosome footprints (RPF), as well as ribosome occupancy (TE), and corresponding p values (pvalue.adjust) for all transcripts as computed by Xtail. For GIGYF2\u0394 in SMG1i vs WT in DMSO. (TSV) Click here for additional data file.", "files": [ "pgen.1009813.s020.tsv" ] }, { "label": "S13 Table", "description": "Log2 fold changes in RNA-seq (mRNA) and ribosome footprints (RPF), as well as ribosome occupancy (TE), and corresponding p values (pvalue.adjust) for all transcripts as computed by Xtail. For WT in SMG1i vs WT in DMSO. (TSV) Click here for additional data file.", "files": [ "pgen.1009813.s021.tsv" ] }, { "label": "S14 Table", "description": "GIGYF2 immunoprecipitation and mass spectrometry with RNase or spliceostatin A treatment. Immunoaffinity purification/mass spectrometry analyses of StrepII-GIGYF2 from untreated, RNase A and SSA treated HEK293T cells. Label free quantification (LFQ) intensities along with the average fold changes and statistical significance are reported. Searched with Andromeda and analyzed with Maxquant (1.6.7.0) and Perseus (1.6.7.0). (XLSX) Click here for additional data file.", "files": [ "pgen.1009813.s022.xlsx" ] }, { "label": "S15 Table", "description": "GIGYF2 immunoprecipitation and mass spectrometry with SMG1i treatment. Same as S14 Table , for SMG1i treated HEK293T cells. No p-values were computed for this single-replicate experiment. (XLSX) Click here for additional data file.", "files": [ "pgen.1009813.s023.xlsx" ] }, { "label": "S16 Table", "description": "Description of oligos used in this study. (TXT) Click here for additional data file.", "files": [ "pgen.1009813.s024.txt" ] }, { "label": "S17 Table", "description": "Description of plasmids used in this study. 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