High-resolution quantitative proteome analysis reveals substantial differences between phagosomes of RAW 264.7 and bone marrow derived macrophages.

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The RAW 264.7 cell line is commonly used for experiments in the macrophage and phagocytosis field. However, little is known how its functions compare to primary macrophages. Here, we have performed an in-depth proteomics characterization of phagosomes from RAW 264.7 and bone marrow derived macrophages by quantifying more than 2500 phagosomal proteins. Our data indicate that there are significant differences for a large number of proteins including important receptors such as mannose receptor 1 and Siglec-1. Moreover, bone marrow derived macrophages phagosomes mature considerably faster by fusion with endosomes and the lysosome which we validated using fluorogenic phagocytic assays. We provide a valuable resource for researcher in the field and recommend careful use of the RAW 264.7 cell line when studying phagosome functions. All MS data have been deposited in the ProteomeXchange with identifier PXD001293 (http://proteomecentral.proteomexchange.org/dataset/PXD001293).", "claims": [{"id": "claim-method", "statement": "Error bars represent standard deviation, pair‐wise t‐test comparison with ***p < 0.0001 and **p < 0.001.", "kind": "methodological", "target": null, "evidence": [{"asset": "source:pmid:25504905", "locator": "sections[6] \"fig\" paragraph 1", "quote": "Error bars represent standard deviation, pair‐wise t‐test comparison with ***p < 0.0001 and **p < 0.001."}], "state": "complete"}, {"id": "claim-data-access", "statement": "All MS data have been deposited in the ProteomeXchange with identifier PXD001293 (http://proteomecentral.proteomexchange.org/dataset/PXD001293).", "kind": "methodological", "target": null, "evidence": [{"asset": "source:pmid:25504905", "locator": "sections[0] \"Abstract\" paragraph 1", "quote": "All MS data have been deposited in the ProteomeXchange with identifier PXD001293 (http://proteomecentral.proteomexchange.org/dataset/PXD001293)."}], "state": "complete"}, {"id": "claim-target-result", "statement": "RAW 264.7 and bone marrow-derived macrophage phagosomes differ substantially in protein composition, with bone marrow-derived macrophage phagosomes maturing faster; proteomic differences were quantified by label-free quantitation using MaxQuant v1.5.0.12, with enrichment defined as twofold and p < 0.05, and the source MS data are available through ProteomeXchange/PRIDE dataset PXD001293 at http://proteomecentral.proteomexchange.org/dataset/PXD001293.", "kind": "quantitative", "target": {"value": 0.05, "unit": "p-value", "tolerance": null, "source": "A total of 290 and 226 proteins were significantly enriched (twofold, p < 0.05) on BMDM and RAW 264.7 phagosomes, respectively."}, "evidence": [{"asset": "source:pmid:25504905", "locator": "sections[0] \"Abstract\" paragraph 1", "quote": "Our data indicate that there are significant differences for a large number of proteins including important receptors such as mannose receptor 1 and Siglec‐1."}, {"asset": "source:pmid:25504905", "locator": "sections[7] \"p\" paragraph 1", "quote": "Data were analyzed by label‐free quantitation using MaxQuant v1.5.0.12 21 and searched against a murine Uniprot‐Trembl database (51 372 entries; downloaded February 19 2014) and a list of common contaminants."}, {"asset": "source:pmid:25504905", "locator": "sections[7] \"p\" paragraph 1", "quote": "A total of 290 and 226 proteins were significantly enriched (twofold, p < 0.05) on BMDM and RAW 264.7 phagosomes, respectively."}, {"asset": "source:pmid:25504905", "locator": "sections[0] \"Abstract\" paragraph 1", "quote": "All MS data have been deposited in the ProteomeXchange with identifier PXD001293 (http://proteomecentral.proteomexchange.org/dataset/PXD001293)."}], "state": "complete"}], "gaps": []}, "experiment": {"state": "complete", "objective": "RAW 264.7 and bone marrow-derived macrophage phagosomes differ substantially in protein composition, with bone marrow-derived macrophage phagosomes maturing faster; proteomic differences were quantified by label-free quantitation using MaxQuant v1.5.0.12, with enrichment defined as twofold and p < 0.05, and the source MS data are available through ProteomeXchange/PRIDE dataset PXD001293 at http://proteomecentral.proteomexchange.org/dataset/PXD001293.", "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:25504905"], "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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