{"database":"biostudies-literature","file_versions":[],"scores":null,"additional":{"submitter":["Kunnath-Velayudhan S"],"funding":["NIAID NIH HHS","NCI NIH HHS","NIGMS NIH HHS"],"pagination":["2596-2606"],"full_dataset_link":["https://www.ebi.ac.uk/biostudies/studies/S-EPMC5605459"],"repository":["biostudies-literature"],"omics_type":["Unknown"],"volume":["199(7)"],"pubmed_abstract":["Analysis of Ag-specific CD4+ T cells in mycobacterial infections at the transcriptome level is informative but technically challenging. Although several methods exist for identifying Ag-specific T cells, including intracellular cytokine staining, cell surface cytokine-capture assays, and staining with peptide:MHC class II multimers, all of these have significant technical constraints that limit their usefulness. Measurement of activation-induced expression of CD154 has been reported to detect live Ag-specific CD4+ T cells, but this approach remains underexplored and, to our knowledge, has not previously been applied in mycobacteria-infected animals. In this article, we show that CD154 expression identifies adoptively transferred or endogenous Ag-specific CD4+ T cells induced by Mycobacterium bovis bacillus Calmette-Guérin vaccination. We confirmed that Ag-specific cytokine production was positively correlated with CD154 expression by CD4+ T cells from bacillus Calmette-Guérin-vaccinated mice and show that high-quality microarrays can be performed from RNA isolated from CD154+ cells purified by cell sorting. Analysis of microarray data demonstrated that the transcriptome of CD4+ CD154+ cells was distinct from that of CD154- cells and showed major enrichment of transcripts encoding multiple cytokines and pathways of cellular activation. One notable finding was the identification of a previously unrecognized subset of mycobacteria-specific CD4+ T cells that is characterized by the production of IL-3. Our results support the use of CD154 expression as a practical and reliable method to isolate live Ag-specific CD4+ T cells for transcriptomic analysis and potentially for a range of other studies in infected or previously immunized hosts."],"journal":["Journal of immunology (Baltimore, Md. : 1950)"],"pubmed_title":["Transcriptome Analysis of Mycobacteria-Specific CD4+ T Cells Identified by Activation-Induced Expression of CD154."],"pmcid":["PMC5605459"],"funding_grant_id":["T32 GM007491","P30 CA013330","R21 AI092448","R01 AI098925","P01 AI063537"],"pubmed_authors":["Ng TW","Jacobs WR","Chan J","Johnson AJ","Goldberg MF","Porcelli SA","Kunnath-Velayudhan S","Johndrow CT","Saini NK","Xu J"],"additional_accession":[]},"is_claimable":false,"name":"Transcriptome Analysis of Mycobacteria-Specific CD4+ T Cells Identified by Activation-Induced Expression of CD154.","description":"Analysis of Ag-specific CD4+ T cells in mycobacterial infections at the transcriptome level is informative but technically challenging. Although several methods exist for identifying Ag-specific T cells, including intracellular cytokine staining, cell surface cytokine-capture assays, and staining with peptide:MHC class II multimers, all of these have significant technical constraints that limit their usefulness. Measurement of activation-induced expression of CD154 has been reported to detect live Ag-specific CD4+ T cells, but this approach remains underexplored and, to our knowledge, has not previously been applied in mycobacteria-infected animals. In this article, we show that CD154 expression identifies adoptively transferred or endogenous Ag-specific CD4+ T cells induced by Mycobacterium bovis bacillus Calmette-Guérin vaccination. We confirmed that Ag-specific cytokine production was positively correlated with CD154 expression by CD4+ T cells from bacillus Calmette-Guérin-vaccinated mice and show that high-quality microarrays can be performed from RNA isolated from CD154+ cells purified by cell sorting. Analysis of microarray data demonstrated that the transcriptome of CD4+ CD154+ cells was distinct from that of CD154- cells and showed major enrichment of transcripts encoding multiple cytokines and pathways of cellular activation. One notable finding was the identification of a previously unrecognized subset of mycobacteria-specific CD4+ T cells that is characterized by the production of IL-3. Our results support the use of CD154 expression as a practical and reliable method to isolate live Ag-specific CD4+ T cells for transcriptomic analysis and potentially for a range of other studies in infected or previously immunized hosts.","dates":{"release":"2017-01-01T00:00:00Z","publication":"2017 Oct","modification":"2024-11-12T02:00:52.188Z","creation":"2019-03-26T23:57:52Z"},"accession":"S-EPMC5605459","cross_references":{"pubmed":["28821584"],"doi":["10.4049/jimmunol.1700654"]}}