biostudies-arrayexpressChristian CasarHomo sapienshttps://www.ebi.ac.uk/biostudies/studies/E-MTAB-13405NK cells are increasingly recognized for their modulation of adaptive T cell responses; however the mechanisms by which NK cells modulate immune responses in human are unclear. Here we report that NKp44+ NK cells regulate CD8+ T cell expansion in an HLA-DP haplotype-dependent process. HLA-DP expression was significantly upregulated on CD8+ effector T cells, in particular in HCMV+ individuals. NK cells were activated in vitro through NKp44 by HLA-DP+ CD8+ T cells expressing NKp44-binding HLA-DP haplotypes. In individuals homozygous for non-NKp44-binding HLA-DP haplotypes, larger frequencies of HLA-DP+ CD8+ T cells were observed, and these specifically included hyper-expanded CD8+ T cell clones that were not observed in individuals encoding for NKp44-binding HLA-DP haplotypes. These data identify a pathway by which NKp44+ NK cells can edit CD8+ T cell effector populations in an HLA-DP haplotype-dependent process and prevents the generation of hyper-expanded T cell clones, which have been suggested to have increased potential for autoimmunity.biostudies-arrayexpressSequencing - The scRNA libraries were sequenced on an Illumina NextSeq or HiSeq 4000 to a minimum sequencing depth of 25,000 reads per cell using read lengths of 26 bp read 1, 8 bp i7 index, 98 bp read 2.Sample Collection - Peripheral blood mononuclear cells (PBMCs) were isolated from healthy individuals blood using density gradient centrifugation (Capricorn Scientific). Freshly isolated PBMCs were immediately used for experiments or cryopreserved for subsequent analysis in heat inactivated fetal bovine serum (FBS) (Capricorn Scientific) supplemented with 10% (v/v) dimethyl sulfoxide (DMSO) (Sigma-Aldrich) and stored in liquid nitrogen.Library Construction - 25.000 cells were used for GEM generation through the 10X Chromium Controller using the 10x V3 B Chip GEM generation kit (10x Genomics, USA).Nucleic Acid Extraction - Freshly isolated PBMCs from three healthy individuals were stained using surface markers antibodies and barcoded using antibodies for CITE-Seq using TotalSeq A antibodies. (BioLegend, USA) (Key Resources Table). CD8+ IL-7R+ HLA-DP-, IL-7R+ HLA-DP+, IL-7R- HLA-DP+ cells were FACS-sorted, using a FACSAria fusion (BD Biosciences). FACS-sorted cells were washed once with autoMACS Running Buffer (Miltenyi Biotec) and resuspended in PBS.OrganizationMINSEQE ScoreAssays and DataProcessed DataAdditional FilesMAGE-TAB FilesSequence Alignment - The sequencing reads were process by cellranger (version 4.0.0, 10x Genomics, USA) and aligned against the reference genome provided by 10X (refdata-gex-GRCh38-2020-A).Data Transformation - The cells from the 3 donors were demultiplexed into the CD8+ CD127+ HLA-DP-, CD8+ CD127+ HLA-DP+, and CD8+ CD127- HLA-DP+ populations based on their cell hashing hash tag oligo (HTO, see sample_to_patient_populations.txt file) data with HTODemux implemented in the Seurat package (version 4.2.0). The HTODemux function first performs K-medoid clustering (K = #samples + 1) on the normalized HTO expression data. Then for each CD45-tag the cluster with the lowest average expression was determined and a negative binomial distribution was fit to this cluster. Based on these distributions and a quantile of 0.99 each cell was classified into being positive or negative for each CD45-tag. Cells positive for more than one tag were classified as doublets and discarded. The following filters were then applied to the cells of each demultiplexed sample individually Genes not observed in at least 1 % of all cells were dropped. Low quality or damaged cells were excluded using a combination of multiple sample dependent quality measures: minimum UMI count, minimum and maximum number of expressed genes and mitochondrial transcript percentage. Additionally, we filtered out doublet cell candidates using Scrublet (version 0.2.3). To normalize the UMI counts we used Seurat’s logNormalize method and identified the 2000 most highly variable genes (HVG) based on variance stabilizing transformation. To perform joint analysis on all samples we applied Seurat’s canonical correlation analysis data integration approach using 2,000 genes that appear in the HVG set of the maximum number of samples as anchors.MetabolomicsUnknownTranscriptomicsGenomicsProteomicsIllumina HiSeq 4000RNA-seq of coding RNA from single cellsHomo sapiensChristian CasarfalseNKp44/HLA-DP-dependent regulation of CD8 T cell clonality by NK cellsNK cells are increasingly recognized for their modulation of adaptive T cell responses; however the mechanisms by which NK cells modulate immune responses in human are unclear. Here we report that NKp44+ NK cells regulate CD8+ T cell expansion in an HLA-DP haplotype-dependent process. HLA-DP expression was significantly upregulated on CD8+ effector T cells, in particular in HCMV+ individuals. NK cells were activated in vitro through NKp44 by HLA-DP+ CD8+ T cells expressing NKp44-binding HLA-DP haplotypes. In individuals homozygous for non-NKp44-binding HLA-DP haplotypes, larger frequencies of HLA-DP+ CD8+ T cells were observed, and these specifically included hyper-expanded CD8+ T cell clones that were not observed in individuals encoding for NKp44-binding HLA-DP haplotypes. These data identify a pathway by which NKp44+ NK cells can edit CD8+ T cell effector populations in an HLA-DP haplotype-dependent process and prevents the generation of hyper-expanded T cell clones, which have been suggested to have increased potential for autoimmunity.2025-10-10T00:00:00Z2025-10-10T09:19:52.064Z2024-04-26T08:41:41.324ZE-MTAB-13405EFO_0002944EFO_0004170EFO_0005684EFO_0004917EFO_0005518EFO_0003816EFO_0004184