Single cell RNA-seq of human CD8+ T cells binding the IAV HLA-A24/PB1 498-505 tetramer
Ontology highlight
ABSTRACT:
INSTRUMENT(S): NextSeq 500
ORGANISM(S): Homo sapiens
SUBMITTER: Katherine Kedzierska
PROVIDER: E-MTAB-10109 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
Similar Datasets
Project description:The aim of the experiment was to assess the differentiation and activation status of influenza-specific CD8+ T cells (A2/PB1 413 and A2/BHA 543) from an individual naturally infected with influenza B (PCR-confirmed). Single tetramer+ CD8+ T cells from tetramer-enriched PBMCs were sorted from baseline (~3 months before infection), at 14 days after infections, at 3 months after infection and at 1.5 years after infection. Libraries were prepared using a modified Smart-seq2 protocol as detailed in the methods section of the manuscript.
2019-02-18 | E-MTAB-7606 | biostudies-arrayexpress
Project description:Roots adaptation to drought stress was analyzed using transcriptome and metabolomics profiles in two wild emmer wheat (Triticum turgidum ssp. dicoccoides) genotypes: Y12-3 (drought resistance) and A24-39 (drought susceptible). Roots samples of Y12-3 and A24-39 genotypes grown under well-watered (control) and water-stressed (7 days of withholding water) were collected for RNA extraction and hybridization on Affymetrix wheat microarrays chip.
2013-01-01 | E-GEOD-25940 | biostudies-arrayexpress
Project description:T cells were separately isolated from MLN and PP from two naive mice, and T cells binding to a cholera toxin B subunit specific MHC II tetramer from mice orally immunized with cholera toxin 7 days or boosted 5 days with a second dose of cholera toxin. Cells were purified using flow cytometric cell sorting and 3´ (n=5) or 5´ (n=5) libraries were constructed using the 10X workflow. From 5´libraries TCR libraries were also constructed and sequenced.
2022-06-22 | E-MTAB-11805 | biostudies-arrayexpress
Project description:The ability to detect and isolate bGL1-22/LGL1âspecific human type II NKT cells allowed us to compare the global gene expression profiles of these cells with type I NKT cells using microarray analysis. Principal component analysis revealed that the gene expression profile signature for bGL1-22 and LGL1-specific T cells both before and after activation with anti-CD3/CD28 beads is distinct from that of type I NKT cells. RNA from CD1d tetramer-sorted b-GL122/LGL1âspecific T cells or iNKT cells was amplified, labeled, and hybridized on the Affymetrix Human Genome U133 Plus 2.0 microarray chips. The data were analyzed with GeneSpring GX 12.5 (Agilent Technologies)
2016-02-16 | E-GEOD-77909 | biostudies-arrayexpress
Project description:To characterize the consequences of systemic IL-27R agonism via IL-27 overexpression or IL-27 blockade (αIL-27) on anti-tumor CTL, we performed scRNA-seq of M86 neoantigen-specific CD8+ T cells from MC38 tumors and dLN.
2024-12-05 | E-MTAB-14601 | biostudies-arrayexpress
Project description:Background Clinical success of T cell receptor (TCR) gene therapy has previously been demonstrated for NY-ESO-1 TCR gene therapy. To increase numbers of cancer patients that can be treated with TCR gene therapy we aimed to identify a novel set of high-affinity cancer specific TCRs targeting different cancer testis (CT) antigens in prevalent HLA class I alleles. Methods In this study, we selected based on publicly available gene expression databases the most promising CT genes to target. From these selected genes we identified by HLA peptidomics the naturally processed and presented HLA class I peptides. With these peptide-HLA tetramers were generated, and by single cell sorting CT specific CD8+ T cells were selected, and expanded from the allo-HLA repertoire of healthy donors. By several functional assays high avidity CT-specific T cell clones with safe recognition pattern were selected. To evaluate the potential for clinical application in TCR gene therapy, TCRs were sequenced and transferred into peripheral blood derived CD8+ T cells. Results In total we identified, 7 novel CT-specific TCRs that effectively target MAGE-A1, MAGE-A3, MAGE-A6 and MAGE-A9 in the context of human leukocyte antigen(HLA) -A1, -A2, -A3, -B7, -B35 and -C7. TCR gene transfer into CD8⁺ T cells resulted in efficient cytokine production and cytotoxicity of variety of different tumor types without detectable cross-reactivity. In addition, major in vivo antitumor effects of MAGE-A1 specific TCR engineered CD8⁺ T cells was observed in an orthotopic xenograft model for established multiple myeloma, in which bone marrow located tumor cells were completely eradicated after T cell injection. Conclusion The identification of 7 novel CT-specific TCRs, reactive against CT antigens presented in a variety of different HLA class I alleles, allows selection of therapeutic TCRs for an increased number of cancer patients, and will improve development of personalized TCR gene therapy.
2023-01-23 | PXD038862 | Pride
Project description:2×10^7 HL60 cells were treated with 10 uM ATRA for 2 and 5 days, and conducted Hi-C to study the chromosomal architecture. Cells were fixed by 1% formaldehyde (v/v) for 15 min at room temperature (RT) with slowly rotation. Cells were lysed by 550 μl lysis buffer (500 μl 10 mM Tris-HCl pH 8.0, 10 mM NaCl, 0.2% Igepal CA-630 and 50 μl protease inhibitors) using a homogenizer, and harvested the chromatin at 5,000 rpm and washed twice. The pellet was added 1% SDS and incubated at 65 oC for 10 min, then quenched by Triton X-100. The chromatin was digested by 600 U MboI (NEB, Ipswich, USA) at 37 oC overnight. The next day, enzymes were inactivated by 10% SDS at 65 oC for 30 min, and the restriction fragment overhangs were filled in using biotin-14-dCTP and Klenow at 37 oC for 45 min, and the ligation was conducted in 7.61 ml ligation mix (745 μl 10% Triton X-100, 745 μl 10x ligation buffer (500 mM Tris-HCl pH 7.5, 100 mM MgCl2, 100 mM DTT), 80 μl 10 mg/ml BSA, 80 μl 100 mM ATP and 5.96 ml water) for 4 h at 16°C. The blunt-end Hi-C ligation was sonicated and pull down by streptavidin beads, then purified by ethanol. Hi-C library was prepared for paired ending sequencing using NEBNext® UltraTM DNA Library Prep Kit for Illuina® (NEB). For regular 3C ligation, after enzymes inactivation, 10 μl 1 U/μl T4 DNA ligase was added to incubate at 16°C overnight. The interaction was detected by qPCR using the specific 3C primers (Table 1) followed by DNA purification. Hi-C was conducted in PE-150. An iterative method for Hi-C mapping with initial length 30bp, if it fails to map uniquely, the next round of mapping for additional 20 bp continued, this procedure lasts until the full read length reaches 150bp. The Hi-C reads were mapped to the human reference genome (assembly GRCh38) using bowtie2 (v2.2.9). The reads mapped to the same restriction fragment, the reads less than 500bp and PCR duplicates were all removed. Hi-C contact maps were normalized using ICE.
2020-05-07 | E-MTAB-8547 | biostudies-arrayexpress
Project description:Human cytomegalovirus infection (CMV) can stimulate robust human leukocyte antigen (HLA)-E restricted CD8 T cell responses. These T cells recognize a peptide from UL40, which differs by as little as a single methyl group from self-peptides that also bind HLA-E, challenging their capacity to avoid self-reactivity. We showed in one donor 2 distinct populations of UL40/HLA-E T cells, with vastly different T cell receptor (TCR) affinities for the UL40/HLA-E complex. However, paradoxically, lower cytokine responses were observed from UL40/HLA-E T cells bearing TCRs with high affinity for HLA-E. To identify why these T cells bearing high affinity T cell receptors were less responsive to antigens, we performed single cell RNAseq analysis. These 2 distinct populations of T cells were single-cell sorted into 96-well plates prior to single cell RNA-seq analysis.
2021-05-21 | E-MTAB-10305 | biostudies-arrayexpress
Project description:HLA-E molecules can present self and pathogen-derived peptides to both NK-cells and T-cells. T-cells that recognize HLA-E peptides via their T-cell receptor (TCR) are termed donor-unrestricted T-cells due to restricted allelic variation of HLA-E. The composition and repertoire of HLA-E TCRs is not known so far. We performed TCR sequencing on CD8+ T-cells from 21 individuals recognizing HLA-E tetramers (TM) folded with 2 Mtb HLA-E restricted peptides. We sorted HLA-E Mtb TM+ and TMCD8+ T-cells directly ex vivo and performed bulk RNA-sequencing and single cell TCR sequencing. The identified TCR repertoire was diverse and showed no conservation between and within individuals. TCRs selected from our single cell TCR sequencing data could be activated upon HLA-E/peptide stimulation, although not robust, reflecting potentially weak interactions between HLA-E peptide complexes and TCRs. Thus, HLA-E Mtb specific T-cells have a highly diverse TCR repertoire.
2024-02-04 | GSE236154 | GEO
Project description:Thirty female BALB/c mice of 6-8 weeks of age were purchased from Charles River (Montreal, QC, Canada) and divided randomly into control (n=15) and treated (n=15) groups. Mice were maintained on an egg free diet in the pathogen-free conditions at the University of Guelph Central Animal Facility. Animals were maintained between 2224°C and at a relative humidity of 4050%. The light-to-dark cycle was maintained at 12 h from 7 AM to 7 PM. All of the animal experiments were approved by the University of Guelph Animal Care Committee according the Canadian Council on Animal Care (CCAC) guidelines.Mice were exposed to intra-gastric gavage (i.g.) twice every week for 5 weeks with OVM (1 mg/100 μl) and 10 μg of cholera toxin adjuvant. An amino acid (Sigma, St. Louis, MO) solution of 10 μg/μl (100 μl per mouse with 10 μg of cholera toxin) was used as a negative control. The amino acids were mixed accordingly to match the OVM amino acid composition. Five weeks after the first sensitization, mice were challenged by i.p. injection with 1 mg of OVM in 50 μl of sterile water and aluminum hydroxide (50 μl, 2% Alhydrogel) adjuvant (Superfos Biosector, Denmark), to induce an immediate type-1 hypersensitivity reaction. The anaphylactic responses were visualized and scored depending on the response of the individual mice in each of the treated and control groups. After 40 min, mice were euthanized with CO2 and partial spleen tissue samples were collected in RNAlater solution for microarray experiment (Ambion Inc., TX, USA). Spleen samples in RNAlater solution were kept at 4oC overnight prior to long term storage at -80oC.
2010-07-15 | E-MEXP-2796 | biostudies-arrayexpress