Quantitative Interactomics of Lck-TurboID in Living Human T Cells Unveils T Cell Receptor Stimulation-Induced Proximal Lck Interactors.

{"sci_version": "1.2", "schema": "https://corpus.bntrg.co.uk/sci/1.0/sci.schema.json", "id": "pmid:33185455", "kind": "science-record", "revision": 4, "content_hash": "sha256:6a224cb4d0cd4ba9ba4ffc3215cb200f41babec818d3569be9f011dc794b54aa", "created_at": "2026-11-01T17:34:18Z", "updated_at": "2026-11-01T22:02:30Z", "generator": {"name": "corpus-assay-engine", "run_id": "reading-sci-2026-11-01"}, "paper": {"state": "complete", "title": "Quantitative Interactomics of Lck-TurboID in Living Human T Cells Unveils T Cell Receptor Stimulation-Induced Proximal Lck Interactors.", "doi": "10.1021/acs.jproteome.0c00616", "pmid": "33185455", "authors": ["Xien Yu Chua", "Timothy Aballo", "William Elnemer", "Melanie Tran", "Arthur Salomon"], "journal": "Journal of proteome research", "year": 2021, "abstract": "While Lck has been widely recognized to play a pivotal role in the initiation of the T cell receptor (TCR) signaling pathway, an understanding of the precise regulation of Lck in T cells upon TCR activation remains elusive. Investigation of protein-protein interaction (PPI) using proximity labeling techniques such as TurboID has the potential to provide valuable molecular insights into Lck regulatory networks. By expressing Lck-TurboID in Jurkat T cells, we have uncovered a dynamic, short-range Lck protein interaction network upon 30 min of TCR stimulation. In this novel application of TurboID, we detected 27 early signaling-induced Lck-proximal interactors in living T cells, including known and novel Lck interactors, validating the discovery power of this tool. Our results revealed previously unappreciated Lck PPI which may be associated with cytoskeletal rearrangement, ubiquitination of TCR signaling proteins, activation of the mitogen-activated protein kinase cascade, coalescence of the LAT signalosome, and formation of the immunological synapse. In this study, we demonstrated for the first time in immune cells and for the kinase Lck that TurboID can be utilized to unveil PPI dynamics in living cells at a time scale consistent with early TCR signaling. Data are available via ProteomeXchange with identifier PXD020759.", "claims": [{"id": "claim-method", "statement": "Missing protein intensity values were imputed from the normal distribution of each replicate (width 0.3; downshift 1.8) before p values were obtained using a two-sample unpaired student’s t-test. Subsequently, p values were corrected to q-values using permutation-based FDR multiple hypothesis testing correction.", "kind": "methodological", "target": null, "evidence": [{"asset": "source:pmid:33185455", "locator": "sections[10] \"Data Analysis\" paragraph 2", "quote": "Missing protein intensity values were imputed from the normal distribution of each replicate (width 0.3; downshift 1.8) before p values were obtained using a two-sample unpaired student’s t-test. Subsequently, p values were corrected to q-values using permutation-based FDR multiple hypothesis testing correction."}], "state": "complete"}, {"id": "claim-data-access", "statement": "The mass spectrometry proteomic data have been deposited to the ProteomeXchange Consortium (http://proteomecentral.proteomexchange.org) via the PRIDE partner repository 75 with the dataset identifier PXD020759.", "kind": "methodological", "target": null, "evidence": [{"asset": "source:pmid:33185455", "locator": "sections[27] \"Data Availability\" paragraph 2", "quote": "The mass spectrometry proteomic data have been deposited to the ProteomeXchange Consortium (http://proteomecentral.proteomexchange.org) via the PRIDE partner repository 75 with the dataset identifier PXD020759."}], "state": "complete"}, {"id": "claim-target-result", "statement": "We detected 27 early signaling-induced Lck-proximal interactors in living T cells, including known and novel Lck interactors, validating the discovery power of this tool.", "kind": "qualitative", "target": null, "evidence": [{"asset": "source:pmid:33185455", "locator": "sections[0] \"Abstract\" paragraph 1", "quote": "In this novel application of TurboID, we detected 27 early signaling-induced Lck-proximal interactors in living T cells, including known and novel Lck interactors, validating the discovery power of this tool."}], "state": "complete"}], "gaps": []}, "experiment": {"state": "complete", "objective": "We detected 27 early signaling-induced Lck-proximal interactors in living T cells, including known and novel Lck interactors, validating the discovery power of this tool.", "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:33185455"], "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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Jurkat cells were transfected with the construct via electroporation (260", "rated using PX458-Lck as indicated above via CRISPR/Cas9 targeted gene knockout 20. Jurkat cells", "to confirm that the genomic DNA targeted by CRISPR/Cas9 resulted in the disruption of the codin", "up Fig S1-s7, table s1-s5, file s1Figure S1: CRISPR knockout of Lck disrupts of coding frames in"], "order_basis": "paper_text_order:1945", "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": 0.99, "evidence": ["le cell fluorescence-activated cell sorting (FACS). Expanded single clones were screened for t", "e RPMI media for 48 hours before single cell fluorescence-activated cell sorting (FACS). Expanded single clones were screened", "rs before single cell fluorescence-activated cell sorting (FACS). Expanded single clones were screened"], "order_basis": "paper_text_order:2155", "id": "assay:2"}, {"role": "data_generation", "definition_id": "western_blot", "definition_version": "99e1195c7d6f", "definition_digest": null, "required_operations": [], "support_state": "partial", "source_hashes": {"assay_lexicon": "99e1195c7d6f"}, "confidence": 0.99, "evidence": ["ed for the loss of Lck protein expression by Western blot and subsequently sequenced to confirm that t", "ere verified by accessing Lck expression via Western blotting. J.Lck.WT was generated similarly using pEF6", "ed TCR signaling proteins.File S1: Uncropped Western blot images. Table S1-S5Table S1: List of primers"], "order_basis": "paper_text_order:2278", "id": "assay:3"}, {"role": "analysis", "definition_id": "network_analysis", "definition_version": "f2132e96ae6c7a4e", "definition_digest": null, "required_operations": ["build_graph", "centrality_scores", "pathway_enrich"], "support_state": "supported", "source_hashes": {"dsl_catalogue": "f2132e96ae6c7a4e"}, "confidence": 0.7, "evidence": ["protein-protein interaction"], "order_basis": "consumes_earlier_output", "id": "assay:4"}]}