A framework for TRIM21-mediated protein depletion in early mouse embryos: recapitulation of Tead4 null phenotype over three days.

{"sci_version": "1.2", "schema": "https://corpus.bntrg.co.uk/sci/1.0/sci.schema.json", "id": "pmid:31638890", "kind": "science-record", "revision": 7, "content_hash": "sha256:3b740e32075ee3749c0d4bc51c686abbda9dbf35b3338bec32f70ab1717eb3fc", "created_at": "2026-11-01T15:40:50Z", "updated_at": "2026-11-01T22:00:56Z", "generator": {"name": "corpus-assay-engine", "run_id": "reading-sci-2026-11-01"}, "paper": {"state": "complete", "title": "A framework for TRIM21-mediated protein depletion in early mouse embryos: recapitulation of Tead4 null phenotype over three days.", "doi": "10.1186/s12864-019-6106-2", "pmid": "31638890", "authors": ["Steffen Israel", "Ellen Casser", "Hannes C A Drexler", "Georg Fuellen", "Michele Boiani"], "journal": "BMC genomics", "year": 2019, "abstract": "BACKGROUND: While DNA and RNA methods are routine to disrupt the expression of specific genes, complete understanding of developmental processes requires also protein methods, because: oocytes and early embryos accumulate proteins and these are not directly affected by DNA and RNA methods. When proteins in the oocyte encounter a specific antibody and the TRIpartite Motiv-containing 21 (TRIM21) ubiquitin-protein ligase, they can be committed to degradation in the proteasome, producing a transient functional knock-out that reveals the role of the protein. However, there are doubts about whether this targeted proteolysis could be successfully used to study mammalian development, because duration of the transient effect is unknown, and also because amounts of reagents delivered must be adequate in relation to the amount of target protein, which is unknown, too.\nRESULTS: We show that the mouse egg contains up to 1E-02 picomoles/protein, as estimated by mass spectrometry using the intensity-based absolute quantification (iBAQ) algorithm. However, the egg can only accommodate ≈1E-04 picomoles of antibody or TRIM21 without incurring toxic effects. Within this framework, we demonstrate that TRIM21-mediated protein depletion efficiently disrupts the embryonic process of trophectoderm formation, which critically depends on the TEA domain family member 4 (Tead4) gene. TEAD4 depletion starting at the 1-cell stage lasts for 3 days prior to a return of gene and protein expression to baseline. This time period is long enough to result in a phenotype entirely consistent with that of the published null mutation and RNA interference studies: significant underexpression of trophectodermal genes Cdx2 and Gata3 and strongly impaired ability of embryos to cavitate and implant in the uterus. Omics data are available via ProteomeXchange (PXD012613) and GEO (GSE124844).\nCONCLUSIONS: TRIM21-mediated protein depletion can be an effective means to disrupt gene function in mouse development, provided the target gene is chosen carefully and the method is tuned accurately. The knowledge gathered in this study provides the basic know-how (prerequisites, requirements, limitations) to expedite the protein depletion of other genes besides Tead4.", "claims": [{"id": "claim-data-access", "statement": "Availability of data and materialsThe mass spectrometry proteomics data have been deposited to the ProteomeXchange Consortium via the PRIDE partner repository [39, 40] with the data set identifier PXD012613. The microarray data generated and analyzed in this article have been deposited in the NCBI’s Gene Expression Omnibus and are accessible through GEO Series accession number GSE124844. Summary tables of PXD012613 and GSE124844 are provided in supplementary material.", "kind": "methodological", "target": null, "evidence": [{"asset": "source:pmid:31638890", "locator": "sections[30] \"notes\" paragraph 1", "quote": "Availability of data and materialsThe mass spectrometry proteomics data have been deposited to the ProteomeXchange Consortium via the PRIDE partner repository [39, 40] with the data set identifier PXD012613. The microarray data generated and analyzed in this article have been deposited in the NCBI’s Gene Expression Omnibus and are accessible through GEO Series accession number GSE124844. 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RESULTS: We show that the mouse egg contains up to 1E-02 picomoles/protein,", "effective means to disrupt gene function in mouse development, provided the target gene is cho", "nce with regulations on research animals All mice were maintained in individually ventilated cages"], "order_basis": "paper_text_order:14", "id": "assay:1"}, {"role": "data_generation", "definition_id": "mass_spectrometry", "definition_version": "99e1195c7d6f", "definition_digest": null, "required_operations": [], "support_state": "partial", "source_hashes": {"assay_lexicon": "99e1195c7d6f"}, "confidence": 0.99, "evidence": ["to 1E-02 picomoles/protein, as estimated by mass spectrometry using the intensity-based absolute quantific", "mples were analyzed by liquid chromatography-mass spectrometry (LC-MS/MS), either on a LTQ Orbitrap Velos o", "described in our previous work [23, 65]. Raw mass spectrometry data were deposited to the PRIDE repository", "by liquid chromatography-mass spectrometry (LC-MS/MS), either on a LTQ Orbitrap Velos or a Q-Exac"], "order_basis": "paper_text_order:1048", "id": "assay:2"}, {"role": "data_generation", "definition_id": "qpcr", "definition_version": "99e1195c7d6f", "definition_digest": null, "required_operations": [], "support_state": "partial", "source_hashes": {"assay_lexicon": "99e1195c7d6f"}, "confidence": 0.99, "evidence": ["for each target gene. PrimeTime®Predesigned qPCR Assay (6-FAM/ZEN/IBFQ) from Integrated DNA T", "neAmp® PCR System 9700 (Applied Biosystems). Real-time quantitative PCR reactions were performed on a 7900 HT FAST R", "R reactions were performed on a 7900 HT FAST Realtime PCR System (Applied Biosystems). The cDNA equiva"], "order_basis": "paper_text_order:7939", "id": "assay:3"}, {"role": "data_generation", "definition_id": "microarray", "definition_version": "99e1195c7d6f", "definition_digest": null, "required_operations": [], "support_state": "partial", "source_hashes": {"assay_lexicon": "99e1195c7d6f"}, "confidence": 0.99, "evidence": ["Transcriptome Analysis Console was used for microarray data analysis. The robust multiarray averagi", "and the statistical program JMP v.13 (SAS). Microarray data analysis was performed in-house using t", "ene expression profiling was performed using Affymetrix GeneChip® Mouse Transcriptome Array 1.0 (Aff", "rix GeneChip® Mouse Transcriptome Array 1.0 (Affymetrix United Kingdom Ltd., High Wycombe, UK) conta"], "order_basis": "paper_text_order:9100", "id": "assay:4"}, {"role": "data_generation", "definition_id": "ihc", "definition_version": "99e1195c7d6f", "definition_digest": null, "required_operations": [], "support_state": "partial", "source_hashes": {"assay_lexicon": "99e1195c7d6f"}, "confidence": 0.99, "evidence": ["ermined by applying an analysis of variance. Immunofluorescence analysis of GFP antibody and TEAD4/CDX2 expr", "ession Embryo were analyzed by performing an immunostaining followed by confocal microscopy imaging. The"], "order_basis": "paper_text_order:10105", "id": "assay:5"}, {"role": "data_generation", "definition_id": "cell_imaging", "definition_version": "99e1195c7d6f", "definition_digest": null, "required_operations": [], "support_state": "partial", "source_hashes": {"assay_lexicon": "99e1195c7d6f"}, "confidence": 0.99, "evidence": ["by performing an immunostaining followed by confocal microscopy imaging. The following primary antibodies we", "rming an immunostaining followed by confocal microscopy imaging. 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