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| Status |
Public on Jul 08, 2024 |
| Title |
SLAM/SAP signaling regulates discrete γδ T cell developmental checkpoints and shapes the innate-like γδ TCR repertoire |
| Organism |
Mus musculus |
| Experiment type |
Other
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| Summary |
During thymic development, most γδ T cells acquire innate-like characteristics that are critical for their function in tumor surveillance, infectious disease, and tissue repair. The mechanisms, however, that regulate γδ T cell developmental programming remain unclear. Recently, we demonstrated that the SLAM-SAP signaling pathway regulates the development and function of multiple innate-like γδ T cell subsets. Here, we used a single-cell proteogenomics approach to identify SAP-dependent developmental checkpoints and to define the SAP-dependent γδ TCR repertoire. SAP deficiency resulted in both a significant loss of an immatureGzma+Blk+Etv5+Tox2+γδT17 precursor population, and a significant increase inCd4+Cd8+Rorc+Ptcra+Rag1+thymic γδ T cells. SAP-dependent diversion of embryonic day 17 thymic γδ T cell clonotypes into the αβ T cell developmental pathway was associated with a decreased frequency of mature clonotypes in neonatal thymus, and an altered γδ TCR repertoire in the periphery. Finally, we identify TRGV4/TRAV13-4(DV7)-expressing T cells as a novel, SAP-dependent Vγ4 γδT1 subset. Together, the data suggest that SAP-dependent γδ/αβ T cell lineage commitment regulates γδ T cell developmental programming and shapes the γδ TCR repertoire.
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| Overall design |
Single-cell CITE-Seq and TCR repertoire profiling of γδ T cells across various developmental stages and mouse models shows the influence of SLAM/SAP signaling on discrete γδ T cell developmental checkpoints and the shaping of the innate-like γδ TCR repertoire. Thymus single-cell CITE-seq with TCR repertoire analysis includes embryonic day 17 (E17) B6 and B6.Sh2d1a-/- (SAP-/-) mice with four biological replicates each, along with nine-day-old (D9) neonatal B6 and B6.SAP-/- mice with three biological replicates each. Additionally, the dataset encompasses thymus samples from six-week-old B6 and B6.SAP-/-mice with three replicates per group. Following thymic analysis, single-cell RNAseq with TCR repertoire analysis of adult lung γδ T cellsinvolved pooled cells from three 13-week-old B6 and B6.SAP-/- littermates per group. These experiments generated three libraries per experiments: RNA, VDJ, and cell surface protein (CSP). The cell surface protein (CSP) libraries encompassed both Antibody-derived tags (ADTs)and hashtag oligonucleotides (HTOs). This comprehensive approach sheds light on the molecular mechanisms underlying γδ T cell development and the impact of SLAM/SAP signaling across different developmental stages and mouse models.
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ADT/HTO barcode sequence hashtagged sample ADT sample
TotalSeq-C0212 anti-mouse CD24 TATATCTTTGCCGCA E17thy, D9thy, W6thy
TotalSeq-C0073 anti-mouse CD44 TGGCTTCAGGTCCTA E17thy
TotalSeq-C1063 anti-mouse CD45RB TTGTATCTCCCTTGG E17thy
TotalSeq-C0209 anti-mouse Vg1 TCGTTTAACCAGCCT E17thy
TotalSeq-C0211 anti-mouse Vg4 AAGCTGCACCGTAAT E17thy, D9thy, W6thy, Lung
TotalSeq-C0930 anti-mouse SLAMF6 CGATTCTTTGCGAGT E17thy, D9thy, W6thy
TotalSeq-C0203 anti-mouse SLAMF1 CAACGCCTAGAAACC E17thy, D9thy, W6thy
TotalSeq-C0077 anti-mouse CD73 ACACTTAACGTCTGG E17thy, D9thy, W6thy
TotalSeq-C0301 anti-mouse hashtag 1 ACCCACCAGTAAGAC E17thy_B6-1, D9thy_B6-1, W6thy_B6-1
TotalSeq-C0302 anti-mouse hashtag 2 GGTCGAGAGCATTCA E17thy_B6-2, D9thy_B6-2, W6thy_B6-2
TotalSeq-C0303 anti-mouse hashtag 3 CTTGCCGCATGTCAT E17thy_B6-3, D9thy_B6-3, W6thy_B6-3, Lung_B6
TotalSeq-C0304 anti-mouse hashtag 4 AAAGCATTCTTCACG E17thy_B6-4, D9thy_SAPKO-1, W6thy_SAPKO-1, Lung_SAPKO
TotalSeq-C0305 anti-mouse hashtag 5 CTTTGTCTTTGTGAG E17thy_SAPKO-1, D9thy_SAPKO-2, W6thy_SAPKO-2
TotalSeq-C0306 anti-mouse hashtag 6 TATGCTGCCACGGTA E17thy_SAPKO-2, D9thy_SAPKO-3, W6thy_SAPKO-3
TotalSeq-C0307 anti-mouse hashtag 7 GAGTCTGCCAGTATC E17thy_SAPKO-3
TotalSeq-C0308 anti-mouse hashtag 8 TATAGAACGCCAGGC E17thy_SAPKO-4, D9thy_ADTcontrol, W6thy_ADTcontrol
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| Contributor(s) |
Mistri SK, Hilton BM, Horrigan KJ, Andretta ES, Savard R, Dienz O, Hampel KJ, Gerrard DL, Rose JT, Sidiropoulos N, Majumdar D, Boyson JE |
| Citation(s) |
39656519 |
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| Submission date |
Mar 20, 2024 |
| Last update date |
Jan 02, 2025 |
| Contact name |
Jonathan E Boyson |
| E-mail(s) |
jboyson@med.uvm.edu
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| Organization name |
University of Vermont
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| Department |
Surgery
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| Street address |
89 Beaumont Ave.
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| City |
Burlington |
| State/province |
Vermont |
| ZIP/Postal code |
05405 |
| Country |
USA |
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| Platforms (2) |
| GPL16417 |
Illumina MiSeq (Mus musculus) |
| GPL18480 |
Illumina HiSeq 1500 (Mus musculus) |
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| Samples (12)
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| Relations |
| BioProject |
PRJNA1090116 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE262064_D9thy_Feature_reference_ADT.csv.gz |
243 b |
(ftp)(http) |
CSV |
| GSE262064_D9thy_Feature_reference_HTO.csv.gz |
260 b |
(ftp)(http) |
CSV |
| GSE262064_D9thy_RNA_ADT_barcodes.tsv.gz |
25.7 Kb |
(ftp)(http) |
TSV |
| GSE262064_D9thy_RNA_ADT_features.tsv.gz |
284.2 Kb |
(ftp)(http) |
TSV |
| GSE262064_D9thy_RNA_ADT_filtered_feature_bc_matrix.h5 |
16.5 Mb |
(ftp)(http) |
H5 |
| GSE262064_D9thy_RNA_ADT_matrix.mtx.gz |
43.0 Mb |
(ftp)(http) |
MTX |
| GSE262064_D9thy_RNA_HTO_barcodes.tsv.gz |
25.7 Kb |
(ftp)(http) |
TSV |
| GSE262064_D9thy_RNA_HTO_features.tsv.gz |
284.2 Kb |
(ftp)(http) |
TSV |
| GSE262064_D9thy_RNA_HTO_filtered_feature_bc_matrix.h5 |
16.5 Mb |
(ftp)(http) |
H5 |
| GSE262064_D9thy_RNA_HTO_matrix.mtx.gz |
43.1 Mb |
(ftp)(http) |
MTX |
| GSE262064_D9thy_VDJ_all_contig_annotations.csv.gz |
352.8 Kb |
(ftp)(http) |
CSV |
| GSE262064_D9thy_custom_processed_VDJ_TRGV_TRDV_paired.xlsx |
1.1 Mb |
(ftp)(http) |
XLSX |
| GSE262064_E17thy_Feature_reference_ADT.csv.gz |
293 b |
(ftp)(http) |
CSV |
| GSE262064_E17thy_Feature_reference_HTO.csv.gz |
276 b |
(ftp)(http) |
CSV |
| GSE262064_E17thy_RNA_ADT_barcodes.tsv.gz |
31.4 Kb |
(ftp)(http) |
TSV |
| GSE262064_E17thy_RNA_ADT_features.tsv.gz |
284.2 Kb |
(ftp)(http) |
TSV |
| GSE262064_E17thy_RNA_ADT_filtered_feature_bc_matrix.h5 |
34.6 Mb |
(ftp)(http) |
H5 |
| GSE262064_E17thy_RNA_ADT_matrix.mtx.gz |
94.3 Mb |
(ftp)(http) |
MTX |
| GSE262064_E17thy_RNA_HTO_barcodes.tsv.gz |
31.4 Kb |
(ftp)(http) |
TSV |
| GSE262064_E17thy_RNA_HTO_features.tsv.gz |
284.2 Kb |
(ftp)(http) |
TSV |
| GSE262064_E17thy_RNA_HTO_filtered_feature_bc_matrix.h5 |
34.5 Mb |
(ftp)(http) |
H5 |
| GSE262064_E17thy_RNA_HTO_matrix.mtx.gz |
94.3 Mb |
(ftp)(http) |
MTX |
| GSE262064_E17thy_VDJ_all_contig_annotations.csv.gz |
444.5 Kb |
(ftp)(http) |
CSV |
| GSE262064_E17thy_custom_processed_VDJ_TRGV_TRDV_paired.xlsx |
1.4 Mb |
(ftp)(http) |
XLSX |
| GSE262064_Lung_Feature_reference_ADT.csv.gz |
164 b |
(ftp)(http) |
CSV |
| GSE262064_Lung_Feature_reference_HTO.csv.gz |
182 b |
(ftp)(http) |
CSV |
| GSE262064_Lung_RNA_ADT_barcodes.tsv.gz |
22.7 Kb |
(ftp)(http) |
TSV |
| GSE262064_Lung_RNA_ADT_features.tsv.gz |
284.1 Kb |
(ftp)(http) |
TSV |
| GSE262064_Lung_RNA_ADT_filtered_feature_bc_matrix.h5 |
8.9 Mb |
(ftp)(http) |
H5 |
| GSE262064_Lung_RNA_ADT_matrix.mtx.gz |
23.2 Mb |
(ftp)(http) |
MTX |
| GSE262064_Lung_RNA_HTO_barcodes.tsv.gz |
22.7 Kb |
(ftp)(http) |
TSV |
| GSE262064_Lung_RNA_HTO_features.tsv.gz |
284.1 Kb |
(ftp)(http) |
TSV |
| GSE262064_Lung_RNA_HTO_filtered_feature_bc_matrix.h5 |
8.9 Mb |
(ftp)(http) |
H5 |
| GSE262064_Lung_RNA_HTO_matrix.mtx.gz |
23.2 Mb |
(ftp)(http) |
MTX |
| GSE262064_Lung_VDJ_all_contig_annotations.csv.gz |
394.7 Kb |
(ftp)(http) |
CSV |
| GSE262064_Lung_custom_processed_VDJ_TRGV_TRDV_paired.xlsx |
996.9 Kb |
(ftp)(http) |
XLSX |
| GSE262064_W6thy_Feature_reference_ADT.csv.gz |
225 b |
(ftp)(http) |
CSV |
| GSE262064_W6thy_Feature_reference_HTO.csv.gz |
260 b |
(ftp)(http) |
CSV |
| GSE262064_W6thy_RNA_ADT_barcodes.tsv.gz |
35.9 Kb |
(ftp)(http) |
TSV |
| GSE262064_W6thy_RNA_ADT_features.tsv.gz |
284.1 Kb |
(ftp)(http) |
TSV |
| GSE262064_W6thy_RNA_ADT_filtered_feature_bc_matrix.h5 |
20.1 Mb |
(ftp)(http) |
H5 |
| GSE262064_W6thy_RNA_ADT_matrix.mtx.gz |
52.6 Mb |
(ftp)(http) |
MTX |
| GSE262064_W6thy_RNA_HTO_barcodes.tsv.gz |
35.8 Kb |
(ftp)(http) |
TSV |
| GSE262064_W6thy_RNA_HTO_features.tsv.gz |
284.2 Kb |
(ftp)(http) |
TSV |
| GSE262064_W6thy_RNA_HTO_filtered_feature_bc_matrix.h5 |
20.1 Mb |
(ftp)(http) |
H5 |
| GSE262064_W6thy_RNA_HTO_matrix.mtx.gz |
52.6 Mb |
(ftp)(http) |
MTX |
| GSE262064_W6thy_VDJ_all_contig_annotations.csv.gz |
887.0 Kb |
(ftp)(http) |
CSV |
| GSE262064_W6thy_custom_processed_VDJ_TRGV_TRDV_paired.xlsx |
1.6 Mb |
(ftp)(http) |
XLSX |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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