Gene expressions of CD4+ T cells in each developmental stages
Ontology highlight
ABSTRACT:
ORGANISM(S): Homo sapiens
SUBMITTER: Masaru Takeshita
PROVIDER: E-GEOD-61697 | biostudies-arrayexpress |
REPOSITORIES: biostudies-arrayexpress
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Project description:Although it is recognised that CD8+ T memory stem cells (Tscm) are heterogeneous in phenotype and function, very few studies have sought to systematically examine these characteristics, particularly in human samples, as the population as a whole is rare and antigen-specific cells even more so. For each T cell subsets investigated, we have performed optimized RNAseq for small number of cells by adapting the Smartseq2 protocol from scRNAseq. The mini-bulk number of cells was chosen to reduce heterogeneity but also maintain an adequate transcriptomics signal. Adapted from Smartseq2 protocol from scRNAseq described by Picelli et al, cell lysis buffer was prepared by adding 1 μl RNase inhibitor (Clontech, Mountain View, USA) to 19 μl Triton X‐100 solution (0.2% v/v). CD8 T cell subpopulations (TN, TSCM, TCM and TEM together with TSCM CXCR3hi/lo and CD122hi/lo) were bulk sorted (50 cells/sample, with replicates) directly into 96-well PCR plates containing 2 μl lysis buffer, 1 μl dNTP mix (10 mM) and 1 μl oligo‐dT primer at 5 μM. Plates were stored at -80 °C until ready to perform the assay. Reverse-transcription and PCR amplifications were performed as described, with the exception of reducing the IS PCR primer to a 50 nM final concentration and increasing the number of cycles to 28. Sequencing libraries were prepared using the Nextera XT Library Preparation Kit (Illumina; San Diego, USA) and sequencing was performed on Illumina NextSeq500 sequencing platforms with 2×150bp paired-end read output, the read depth was approximately 1.3 million reads per sample.
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Project description:Differentiation of CD4+T-cells into effector subsets is a critical component of the adaptive immune system and an incorrect response can lead to autoimmunity or immune deficiency. Cellular differentiation including T-cell differentiation is accompanied by large-scale epigenetic remodeling, including changes in DNA methylation at key regulators of T-cell differentiation. The TET family of enzymes were recently shown to be able to catalyse methylated cytosine (5mC) into 5-hydroxymethylcytosine (5hmC) enabling a pathway of active removal of DNA methylation. Here, we characterize 5hmC, 5mC and transcriptional dynamics during human CD4+T-cell polarisation in a time series approach and relate these changes to profiles in ex-vivo CD4+memory subsets. We observed large-scale remodelling during early CD4+T-cell differentiation which was predictive of subsequent changes during late time points, these changes were also related to disease associated regions which we show can act as functional regulatory elements. This dataset was designed to assess how gene expression changes over time during human CD4+T-cell polarization towards Th1 and Th2. Gene expression was assessed in relationship to 5hmC and DNA methylation levels and changes (see data series), we observed characteristic gene expression for the specific time points and stimuli or cell type and the expression was correlated with gene body 5hmC as well as anticorrelated with promoter DNA methylation levels. This submission contains data from transcription profiling by array of human CD4+T-cells, Th1/Th2 polarized time-series and primary memory subsets. It is part of series containing 5hmC and DNA methylation profiling of the same samples. See related experiments E-MTAB-4685, E-MTAB-4686, E-MTAB-4688, E-MTAB-4689.
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Project description:Expression data from human naïve (TN), stem cell memory (TSCM), central memory (TCM) and effector memory (TEM) CD8+ T cells
| PRJNA131239 | ENA