Single-cell gene expression analysis of in vitro expanded CD4 T cells responding to HHV-6B and HCMV epitopes
Ontology highlight
ABSTRACT:
INSTRUMENT(S): Illumina HiSeq 4000
ORGANISM(S): Homo sapiens
SUBMITTER: Aniuska Becerra
PROVIDER: E-MTAB-16174 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:Assessment of the functional differences of HHV-6B-specific and HCMV-specific CD4 T cells at RNA level. The peripheral blood mononuclear cells samples were collected from normal subjects expressing HLA-DRB1*03:01. The HHV-6B-specific or HCMV-specific T cells were expanded in vitro using known HLA-DRB1*03:01 epitopes derived from HHV-6B (6 epitopes) or HCMV (1 epitope) and isolated after 12-15 days of culture using the corresponding HLA-DRB1*03:01 tetramers.
2025-11-20 | E-MTAB-16144 | biostudies-arrayexpress
Project description:Infection or reactivation with human cytomegalovirus (HCMV) is still a major cause of morbidity and mortality in patients undergoing solid organ transplantation or allogeneic stem cell transplantation (alloSCT). Recent studies have shown that protection against latent HCMV infection relies on a much broader antigen spectrum than previously expected. The identification of novel immunogenic HCMV-derived peptides that facilitate HCMV control is therefore essential to improve immune monitoring of HCMV-specific T cells and HCMV vaccine development. In particular, only a few HCMV-derived peptides have been described for less common HLA alleles, such as HLA-A*03:01 and HLA-B*15:01, limiting the implementation of these techniques for a substantial number of patients. Here, we identified novel HCMV-derived HLA-A*03:01- and HLA-B*15:01-restricted immunogenic peptides by an innovative combined in vitro and in silico approach. We utilized the computational tools Peptide-PRISM (1) and PRICE (2) to analyze mass spectrometric (MS) and ribosome sequencing (ribo-seq) datasets, respectively. To refine our in silico approach, we utilized machine learning to rate the identified peptide candidates based on their translational activity, binding affinity and positioning within the small open reading frames (sORFs), with the goal of assessing their likelihood to be presented on HLA-A*03:01 and HLA-B*15:01 molecules. After identifying the highest-scoring 49 canonical and 49 cryptic potentially immunogenic peptides for each HLA allele, we assessed their immunogenicity by screening HCMV-seropositive and -seronegative healthy donors, as well as alloSCT patients for their peptide-specific T-cell responses. We used a stimulation procedure in two consecutive steps: peripheral blood cells (PBMCs) were stimulated starting with peptide pools and subsequently with potentially reactive single peptides. In vitro T-cell stimulation resulted in the direct identification of three canonical and one cryptic HLA-A*03-restricted immunogenic peptides as well as five canonical and one cryptic HLA-B*15-restricted immunogenic peptide, with a specific IFNγ+/CD8+ T cell response of ≥ 0.02%. Highest T-cell responses were identified against two HLA-A*03-restricted and three HLA-B*15-restricted canonical peptides with up to 3.42% of IFNγ+/CD8+ T cells in patients 180 days post alloSCT. In conclusion, we identified specific T-cell responses against eight canonical and two cryptic HLA-A*03:01- and HLA-B*15:01-restricted peptides in healthy individuals and post-transplanted patients and which were classified as immunogenic. Of these, six peptides are novel HCMV-derived, immunogenic peptides according to the UniProt database, broadening the spectrum of immunogenic HCMV-derived peptides for personalized immune monitoring and vaccine development.
2024-06-16 | PXD043022 | Pride
Project description:Few non-classical HLA-E restricted HIV-specific epitopes have been described, and even less is known about the functional profile of responding CD8 T cells (CD8s). This study evaluates the functional characteristics of CD8s targeting the Gag epitope (KAFSPEVIPMF or KF11) based on their restriction by either HLA-E (E-CD8s) or HLA-B57 (B57-CD8s). CD8s from 8 people with HIV (PWH) were cocultured with KF11 peptide presented by cell lines expressing HLA-B*57:01, HLA-E*01:01 or E*01:03. CD8s were analyzed through single-cell (sc) RNA and TCR sequencing. Additionally, supernatants were analyzed for soluble proteomics using a Luminex assay. B57-CD8s secreted higher levels of cytotoxic cytokines such as IFNγ, while E-CD8s produced more chemotactic cytokines, including RANTES, CXCL10 (IP-10), and IL27confirmed through scRNAseq. Despite distinct cytokine profiles, TCR clonotypes stimulated by KF11 were cross-restricted by HLA-B*57 and HLA-E*01/03. In vitro T cell reporter assays clearly demonstrated this cross-restriction. A TRAV5-containing metaclonotype cluster was seen in PWH with lower viral loads. These findings demonstrate that HIV-specific CD8s in PWH exhibit cross HLA-B*57 and HLA-E*01/03 restriction, resulting in functionally distinct immune responses that may contribute to HIV control.
2025-12-11 | GSE306022 | GEO
Project description:Few non-classical HLA-E restricted HIV-specific epitopes have been described, and even less is known about the functional profile of responding CD8 T cells (CD8s). This study evaluates the functional characteristics of CD8s targeting the Gag epitope (KAFSPEVIPMF or KF11) based on their restriction by either HLA-E (E-CD8s) or HLA-B57 (B57-CD8s). CD8s from 8 people with HIV (PWH) were cocultured with KF11 peptide presented by cell lines expressing HLA-B*57:01, HLA-E*01:01 or E*01:03. CD8s were analyzed through single-cell (sc) RNA and TCR sequencing. Additionally, supernatants were analyzed for soluble proteomics using a Luminex assay. B57-CD8s secreted higher levels of cytotoxic cytokines such as IFNγ, while E-CD8s produced more chemotactic cytokines, including RANTES, CXCL10 (IP-10), and IL27confirmed through scRNAseq. Despite distinct cytokine profiles, TCR clonotypes stimulated by KF11 were cross-restricted by HLA-B*57 and HLA-E*01/03. In vitro T cell reporter assays clearly demonstrated this cross-restriction. A TRAV5-containing metaclonotype cluster was seen in PWH with lower viral loads. These findings demonstrate that HIV-specific CD8s in PWH exhibit cross HLA-B*57 and HLA-E*01/03 restriction, resulting in functionally distinct immune responses that may contribute to HIV control.
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2024-06-20 | E-MTAB-13654 | biostudies-arrayexpress
Project description:Human CD4+ T lymphocytes play an important role in inducing potent immune responses. They are activated and stimulated by peptides presented in Major Histocompatibility Complex Class II molecules. These MHC class II molecules typically present peptides of between 12 and 20 amino acids in length. The region that interacts with the MHC molecule, designated as the binding core, is highly conserved in the residues which anchor the peptide to the molecule. In addition, as these peptides are the product of proteolytic cleavages, certain conserved residues may be expected at the N- and C-termini outside the binding core. To study whether similar conserved residues are present in different types of cells , the ligandomes of HLA-DRB1*03:01 derived from two different cell types (dendritic cells and EBV-transformed B-cells) were identified with mass spectrometry and the binding core and N- and C-terminal residues were analysed using the frequencies of the amino acids in the human proteome. Similar binding motifs were found as well as comparable conservations in the N- and C-terminal residues. Furthermore, these terminal conservations of the HLA-DRB1*03:01 ligandome were compared to the N- and C-terminal conservations of the HLA-DRB1*04:01 ligandome acquired from dendritic cells. Again, comparable conservations were evident with only minor differences. Taken all together, these analyses show that there are conservations in the terminal residues of peptides, presumably the result of the activity of proteases involved in antigen processing.
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Project description:HLA DRB1*15:01 is overrepresentated in Parkinson's disease patients and binds with high affinity to the ⍺-synuclein peptide, 32-46. Immunization of humanized mice expressing HLA DRB1*15:01 with ⍺-syn32-46 induces enteric phenotypes similar to those of prodromal Parkinson's disease. We collected the small intestines from HLA mice immunized with either Complete Freund's Adjuvant (CFA) with ⍺-syn32-46 or CFA alone, sorted the CD45+ immune cells, then performed single-cell RNA sequencing to evaluate gene expression at single cell resolution.
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2023-08-17 | GSE231873 | GEO
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2020-01-13 | GSE131281 | GEO