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| Status |
Public on Jan 11, 2023 |
| Title |
Time-resolved assessment of single-cell protein secretion by sequencing |
| Organism |
Homo sapiens |
| Experiment type |
Expression profiling by high throughput sequencing
Other
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| Summary |
Secreted proteins play critical roles in cellular communication. Methods enabling concurrent measurement of cellular protein secretion, phenotypes and transcriptomes are still unavailable. Here, we describe Time-Resolved Assessment of Protein Secretion from single cells by sequencing (TRAPS-seq). Released proteins are trapped onto cell surface and probed by oligonucleotide-barcoded antibodies before simultaneously sequenced with transcriptomes in single cells. TRAPS-seq unravels the phenotypic and transcriptional determinants of the secretion of pleiotropic Th1 cytokines (IFN-γ, IL-2 and TNF-α) in activated T cells. We further demonstrate the use of TRAPS-seq to track dynamic secretion of multiple cytokines over time, uncovering unique molecular signatures that govern the preservation or transition of single-cell cytokine secretions. Our results revealed that early central memory T cells with CD45RA expression (TCMRA) are important in both the production and maintenance of polyfunctional cytokines. TRAPS-seq presents a unique tool for seamless integration of secretomics measurement with multi-omics profiling in single cells.
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| Overall design |
For end-timepoint measurement of cytokine secretion by TRAPS-seq, human lymphocytes with depletion of CD19+ cells were enriched using Moflo Astrios cell sorter (Beckman Coulter) and kept in incubator overnight before stimulation. Cells were activated by anti-CD3/CD28 Dynabeads (Gibco, 11161D) at 2:1 bead-to-cell ratio for indicated time periods (0 hour, 1 hours, 6 hours and 16 hours) in 96-well U-bottom plate. Post beads removal, cells were subjected to TRAPS-seq. For secretion tracing, total T cells were enriched by immunomagnetic negative selection (STEMCELL Technologies, 19051) and loaded onto the nanowell array dropwise at 40,000 cells per array. The unloaded cells were gently rinsed off and 20 µl anti-CD3/CD28 Dynabeads (Gibco, 11161D) were loaded. The cells were activated in open array for 10 hours before subjected to TRAPS-seq-powered on-array secretion tracing.
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| Web link |
https://www.nature.com/articles/s41592-023-01841-y
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| Contributor(s) |
Wu T, Cheow L |
| Citation(s) |
37037998 |
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| Submission date |
Apr 12, 2022 |
| Last update date |
Apr 11, 2023 |
| Contact name |
Tongjin WU |
| Organization name |
National University of Singapore
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| Department |
Institute for Health Innovation and Technology
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| Street address |
MD6, 14 Medical Drive
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| City |
Singapore |
| ZIP/Postal code |
117599 |
| Country |
Singapore |
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| Platforms (1) |
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| Samples (8)
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| GSM6042322 |
CD19+ cells-depleted primary lymphocytes, mRNA-derived cDNA |
| GSM6042323 |
CD19+ cells-depleted primary lymphocytes, HTO-derived cDNA |
| GSM6042324 |
CD19+ cells-depleted primary lymphocytes, surface protein marker-derived cDNA |
| GSM6042325 |
CD19+ cells-depleted primary lymphocytes, cytokine-derived cDNA |
| GSM6042326 |
Primary T cells, mRNA-derived cDNA |
| GSM6042327 |
Primary T cells, HTO-derived cDNA |
| GSM6042328 |
Primary T cells, surface protein marker-derived cDNA |
| GSM6042329 |
Primary T cells, cytokine-derived cDNA |
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| Relations |
| BioProject |
PRJNA826085 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE200690_RAW.tar |
85.8 Mb |
(http)(custom) |
TAR (of MTX, TSV) |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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