Transcriptome Analysis of Mycobacteria-Specific CD4+ T Cells Identified by Activation-Induced Expression of CD154

Autor: Steven A. Porcelli, William R. Jacobs, Jiayong Xu, John Chan, Neeraj K. Saini, Michael F. Goldberg, Alison J. Johnson, Tony W. Ng, Christopher T. Johndrow, Shajo Kunnath-Velayudhan
Rok vydání: 2017
Předmět:
Zdroj: The Journal of Immunology. 199:2596-2606
ISSN: 1550-6606
0022-1767
Popis: 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.
Databáze: OpenAIRE
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