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Targeted RNASeq using a custom panel of metabolic genes

ABSTRACT: Purpose: The purpose of this experiment was to determine the time point following activation at which greatest transcriptional differences in metabolic genes were observed between primary human CD4+ T cells treated with metformin + 2-deoxyglucose combination regimen and cells treated with vehicle. Methods: Targeted RNA-Seq data was generated from primary human CD4+ T cell mRNA using a Qiagen custom designed panel of metabolic genes and the Illumina TruSeq stranded mRNA prep. Results: We obtained differential gene expression between metformin + 2-deoxyglucose and vehicle-treated cells using the DESeq2 app in Illumina Basespace for a set of metabolic genes during different time points following activation, and found the greatest changes at 24hr.

ORGANISM(S): Homo sapiens

PROVIDER: GSE144352 | GEO | 2020/06/15

REPOSITORIES: GEO

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Project description:Purpose: To elucidate the potential immune-regulatory mechanism of TRAIL of activated T cells in EAE, we analyzed gene expression profiles of splenic CD4+ T cells from EAE mice treated with TRAIL by RNA sequencing and transcriptome analysis. Methods: Splenic CD4+ T cell mRNA profiles of control or EAE mice treated with vehicle or TRAIL were generated by deep sequencing using Illumina Solexa. Library construction of all samples were used by Agilent's SureSelect Strand Specific RNA Library Preparation Kit for 75SE (Single-End or Paired-End) sequencing on Solexa platform. The sequence was directly determined using sequencing-by-synthesis technology via the TruSeq SBS Kit. Raw sequences were obtained from the Illumina Pipeline software bcl2fastq v2.0 and expected to generate 20M (million reads or Gb) per sample. Results: By comparing mRNA exression of splenic CD4+ T cells from EAE mice treated with vehicle or TRAIL, there were 244 genes significantly differentially expressed. By analyzing these 244 significant genes categorized by a Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis, the most significantly enriched category for CD4+ T cells was “cell cycle” (multiple of enrichment: 3.13, p = 1.64 × 10−5) followed by “TCR signaling pathway” (multiple of enrichment: 2.80, p = 8.25 × 10−4). In addition, significant genes in the“TCR signaling pathway” tended to be downregulated while those in “cell cycle”tended to be upregulated in a volcano plot analysis. Consistent with the volcano plot results, by using unsupervised hierarchical clustering, the heatmap showed that significant TCR signaling pathway-associated genes were downregulated, while significant cell cycle-associated genes were upregulated in splenic CD4+ T cells from EAE mice treated with TRAIL Conclusions: Our study demonstrated the gene transcription pattern from CD4+ T cells of TRAIL-treated EAE mice were involved in distinct TCR signaling and cell cycle pathways.

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Targeted RNASeq using a custom panel of metabolic genes

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