RNF149 negatively regulates LPS/TLR4 signal transduction by ubiquitination-mediated CD63 degradation.

{"sci_version": "1.2", "schema": "https://corpus.bntrg.co.uk/sci/1.0/sci.schema.json", "id": "pmid:39104473", "kind": "science-record", "revision": 2, "content_hash": "sha256:875c5b58d82bffdef4f30123ef0d6ebb864ae10688217ee119e43d1ce5565339", "created_at": "2026-11-01T20:29:11Z", "updated_at": "2026-11-01T21:33:35Z", "generator": {"name": "corpus-assay-engine", "run_id": "reading-sci-2026-11-01"}, "paper": {"state": "complete", "title": "RNF149 negatively regulates LPS/TLR4 signal transduction by ubiquitination-mediated CD63 degradation.", "doi": "10.1016/j.heliyon.2024.e34350", "pmid": "39104473", "authors": ["Xiu-An Yang", "Yingying Wang", "Mingyu Gong", "Zicheng Zhao", "Fengchun Lv", "Xiaoyu Zhang", "Yan Li"], "journal": "Heliyon", "year": 2024, "abstract": "This study aims to investigate the role of RNF149 and tetraspanin CD63 in lipopolysaccharide/Toll-like receptor 4 (LPS/TLR4) signal transduction. TNF-α was assessed using enzyme-linked immunosorbent assay. The distribution of TLR4 was examined through flow cytometry after CD63 knockdown. Real-time polymerase chain reaction was used to analyze the expression of the target genes RNF149 and CD63 under different conditions. Western blotting was employed to detect gene expression, while immunoprecipitation and confocal microscopy were used to evaluate protein interactions. Transcriptome array data from stimulated monocytes (GSE7547) was obtained from GEO and subjected to bioinformatic analysis. It is suggested that CD63 may serve as a substrate of RNF149, with RNF149 capable of directly interacting with CD63. RNF149 degrades CD63 through covalent modification of CD63 at lysine 29 of the ubiquitin monomer, leading to the formation of a multiubiquitin chain. Both RNF149 and CD63 interact with TLR4, with CD63 promoting LPS/TLR4 signaling and RNF149 inhibits it. CD63 does not impact the distribution of TLR4 on the cell surface and does not directly interact with TIRAP, IRAK4, or TRAF6, but does interact with Myd88.RNF149 plays a negative regulatory role in LPS/TLR4 signal transduction by mediating ubiquitination-induced CD63 degradation.", "claims": [{"id": "claim-method", "statement": "The boxplot of characteristic genes was created using the ggboxplot function in the “ggplot 2” package, and the differences in gene expression among the different groups were compared using the wilcox.test.", "kind": "methodological", "target": null, "evidence": [{"asset": "source:pmid:39104473", "locator": "sections[12] \"Bioinformatics analysis\" paragraph 3", "quote": "The boxplot of characteristic genes was created using the ggboxplot function in the “ggplot 2” package, and the differences in gene expression among the different groups were compared using the wilcox.test."}], "state": "complete"}, {"id": "claim-data-access", "statement": "The datasets used and/or analyzed during the current study are available from the corresponding author upon reasonable request.", "kind": "methodological", "target": null, "evidence": [{"asset": "source:pmid:39104473", "locator": "sections[22] \"Data availability statement\" paragraph 2", "quote": "The datasets used and/or analyzed during the current study are available from the corresponding author upon reasonable request."}], "state": "complete"}], "gaps": []}, "experiment": {"state": "complete", "objective": "", "steps": [{"id": "read_published_estimate", "order": 1, "name": "Read the published estimate from the source", "kind": "other", "state": "complete", "blocked_on": null, "description": "The claim, method and evidence span published with the source article.", "duration_hours": null, "parameters": {}, "inputs": ["source:pmid:39104473"], "outputs": ["the published estimate and its evidence"], "instruments": [], "reagents": []}, {"id": "obtain_source_data", "order": 2, "name": "Obtain the source data from the named repository", "kind": "data-preparation", "state": "complete", "blocked_on": null, "description": "The GEO series accession resolves and its sample accessions are listed. 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TNF-α was assessed using enzyme-linked immunosorbent assay. The distribution of TLR4 was examined throu"], "order_basis": "paper_text_order:228", "id": "assay:1"}, {"role": "data_generation", "definition_id": "flow_cytometry", "definition_version": "99e1195c7d6f", "definition_digest": null, "required_operations": [], "support_state": "partial", "source_hashes": {"assay_lexicon": "99e1195c7d6f"}, "confidence": 0.93, "evidence": ["he distribution of TLR4 was examined through flow cytometry after CD63 knockdown. Real-time polymerase c"], "order_basis": "paper_text_order:309", "id": "assay:2"}, {"role": "data_generation", "definition_id": "transfection", "definition_version": "99e1195c7d6f", "definition_digest": null, "required_operations": [], "support_state": "partial", "source_hashes": {"assay_lexicon": "99e1195c7d6f"}, "confidence": 0.99, "evidence": ["s examined through flow cytometry after CD63 knockdown. Real-time polymerase chain reaction was use", "Materials and methods Cell culture and transfection The human embryonic kidney Line 293T (293T),", "endotoxin-free DMEM. The Lipofectamine® 2000 Transfection Reagent was used for plasmid transfection, a", "00 Transfection Reagent was used for plasmid transfection, and the siRNA Transfection Reagent was used"], "order_basis": "paper_text_order:335", "id": "assay:3"}, {"role": "data_generation", "definition_id": "western_blot", "definition_version": "99e1195c7d6f", "definition_digest": null, "required_operations": [], "support_state": "partial", "source_hashes": {"assay_lexicon": "99e1195c7d6f"}, "confidence": 0.93, "evidence": ["RNF149 and CD63 under different conditions. Western blotting was employed to detect gene expression, whil"], "order_basis": "paper_text_order:482", "id": "assay:4"}, {"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": ["ne expression, while immunoprecipitation and confocal microscopy were used to evaluate protein interactions.", "sion, while immunoprecipitation and confocal microscopy were used to evaluate protein interactions."], "order_basis": "paper_text_order:568", "id": "assay:5"}, {"role": "data_generation", "definition_id": "cell_culture", "definition_version": "99e1195c7d6f", "definition_digest": null, "required_operations": [], "support_state": "partial", "source_hashes": {"assay_lexicon": "99e1195c7d6f"}, "confidence": 0.99, "evidence": ["ells were used in this study. The cells were cultured in high-glucose Dulbecco's modified Eagle's med", "d to the culture medium. Raw264.7 cells were cultured in endotoxin-free DMEM. The Lipofectamine® 2000", "293T (293T), human cervical cancer cell line HeLa, and mouse Raw264.7 cells were used in this", "ng the manufacturer's instructions. Briefly, HeLa cells intended for immunofluorescence coloca"], "order_basis": "paper_text_order:1537", "id": "assay:6"}, {"role": "data_generation", "definition_id": "animal_model", "definition_version": "99e1195c7d6f", "definition_digest": null, "required_operations": [], "support_state": "partial", "source_hashes": {"assay_lexicon": "99e1195c7d6f"}, "confidence": 0.69, "evidence": ["), human cervical cancer cell line HeLa, and mouse Raw264.7 cells were used in this study. The"], "order_basis": "paper_text_order:1547", "id": "assay:7"}, {"role": "data_generation", "definition_id": "ihc", "definition_version": "99e1195c7d6f", "definition_digest": null, "required_operations": [], "support_state": "partial", "source_hashes": {"assay_lexicon": "99e1195c7d6f"}, "confidence": 0.93, "evidence": ["structions. Briefly, HeLa cells intended for immunofluorescence colocalization were seeded at a density of 3"], "order_basis": "paper_text_order:2243", "id": "assay:8"}]}