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GEO help: Mouse over screen elements for information. |
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| Status |
Public on May 05, 2023 |
| Title |
Genome-wide CRISPR screens decode cancer cell pathways that trigger gamma-delta T cell detection and killing |
| Organism |
synthetic construct |
| Experiment type |
Other
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| Summary |
target tumours broadly, independent of patient-specific neoantigens or HLA background. Gamma-delta T cells can sense conserved cell stress signals prevalent in transformed cells, although the mechanisms behind the targeting of stressed target cells remain poorly characterized. Vg9Vd2 T cells – the most abundant subset of human gamma-delta T cells – recognize a protein complex containing butyrophilin 2A1 (BTN2A1) and BTN3A1, a widely expressed cell surface protein that is activated by phosphoantigens abundantly produced by tumour cells. Here, we layered genome-wide CRISPR screens in target cancer cells to identify pathways that regulate: (1) killing by gamma-delta T cells during co-culture, and (2) BTN3A cell surface expression. In addition to confirming the importance of the phosphoantigen-producing mevalonate pathway, the screens revealed previously unappreciated multilayered regulation of BTN3A abundance on the cell surface and triggering of gamma-delta T cells through transcription, post-translational modifications, membrane trafficking. Notably, IRF1 and ZNF217 were discovered to be positive and negative transcriptional regulators that directly occupy the promoters of BTN3A1/2/3 genes. In addition, diverse genetic perturbations and inhibitors disrupting metabolic pathways in the cancer cells, particularly ATP-producing processes, were found to alter BTN3A levels. This induction of BTN3A, as well as BTN2A1, during metabolic crises was dependent on AMP-activated protein kinase (AMPK). Finally, small molecule activation of AMPK in a cell line model and in patient-derived tumour organoids led to increased expression of the BTN2A1-BTN3A complex and increased Vg9Vd2 TCR-mediated killing. This AMPK-dependent mechanism of metabolic stress-induced ligand upregulation deepens our understanding of gamma-delta T cell stress surveillance and suggests new avenues to enhance gamma-delta T cell anti-cancer activity.
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| Overall design |
Genome-wide CRISPR KO in Daudi-Cas9 cells, followed by FACS sorting the top (high) and bottom (low) 25% of cells according to BTN3A expression.
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| Contributor(s) |
Mamedov M, Vedova S, Freimer JW, Sahu AD, Ramesh A, Arce MM, Meringa AD, Ota M, Chen PA, Hanspers K, Nguyen VQ, Takeshima KA, Pritchard JK, Kuball J, Sebestyen Z, Adams EJ, Marson A |
| Citation(s) |
37648854 |
| Submission date |
Dec 30, 2021 |
| Last update date |
Sep 08, 2023 |
| Contact name |
Alexander Marson |
| E-mail(s) |
alexander.marson@ucsf.edu
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| Organization name |
Gladstone-UCSF Institute of Genomic Immunology
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| Street address |
1650 Owens St
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| City |
San Francisco |
| State/province |
CA |
| ZIP/Postal code |
94158 |
| Country |
USA |
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| Platforms (1) |
| GPL26526 |
Illumina NovaSeq 6000 (synthetic construct) |
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| Samples (6)
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| Relations |
| BioProject |
PRJNA793280 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE192827_BTN3A_screen_counts.txt.gz |
2.4 Mb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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