SUMMARY Listeria monocytogenes infection generates T helper-1 (Th1) effector memory cells and CC chemokine receptor 7 (CCR7)+ cells resembling central memory cells. We tracked endogenous L. monocytogenes-specific CD4+ T cells to determine how these memory cells are formed. Two effector cell populations were already present several days after infection. One highly expressed the T-bet transcription factor and produced Th1 memory cells in an interleukin-2 (IL-2) receptor-dependent fashion. The other resided in the T cell areas, expressed CCR7 and CXC chemokine receptor 5 (CXCR5), and like follicular helper cells depended on the Bcl6 transcription factor and inducible costimulator ligand on B cells. The CCR7+ CXCR5+ effector cells produced similar memory cells that generated diverse effector cell populations in a secondary response. Thus, Th1 effector memory and follicular helper-like central memory cells are produced from early effector cell populations that diverge in response to signals from the IL-2 receptor, Bcl6, and B cells.

Early T lineage progenitors (ETPs) in the thymus are thought to develop from common lymphoid progenitors (CLPs) in the bone marrow (BM). We compared thymic ETPs to BM CLPs in mice and found that they differed in several respects. Thymic ETPs were not interleukin 7 (IL-7)-responsive and generated B lineage progeny with delayed kinetics, whereas BM CLPs were IL-7-responsive and rapidly generated B cells. ETPs sustained production of T lineage progeny for longer periods of time than BM CLPs. Analysis of Ikaros-deficient mice that exhibit ongoing thymopoiesis without B lymphopoeisis revealed near-normal frequencies of thymic ETPs, yet undetectable numbers of BM CLPs. We conclude that ETPs can develop via a CLP-independent pathway.

SUMMARY A naïve CD4+ T cell population specific for a microbial peptide:major histocompatibility complex II ligand (p:MHCII) typically consists of about 100 cells, each with a different T cell receptor (TCR). Following infection, this population produces a consistent ratio of effector cells that activate microbicidal functions of macrophages or help B cells make antibodies. We studied the mechanism that underlies this division of labor by tracking the progeny of single naïve T cells. Different naïve cells produced distinct ratios of macrophage and B cell helpers but yielded the characteristic ratio when averaged together. The effector cell pattern produced by a given naïve cell correlated with the TCR-p:MHCII dwell time or the amount of p:MHCII. Thus, the consistent production of effector cell subsets by a polyclonal population of naïve cells results from averaging the diverse behaviors of individual clones, which are instructed in part by the strength of TCR signaling.

The mammalian gastrointestinal tract contains a large and diverse population of commensal bacteria and is also one of the primary sites of exposure to pathogens. How the immune system perceives commensals in the context of mucosal infection is unclear. Here we show that during a gastrointestinal infection, tolerance to commensals is lost and microbiota-specific T cells are activated and differentiate to inflammatory effector cells. Furthermore, these T cells go on to form memory cells that are phenotypically and functionally consistent with pathogen-specific T cells. Our results suggest that during a gastrointestinal infection, the immune response to commensals parallels the immune response against pathogenic microbes and that adaptive responses against commensals are an integral component of mucosal immunity.

Unraveling the complexity of the adaptive immune system requires the study of T cells in vivo. This protocol describes how populations of T cells specific for a given peptide: Major Histocompatibility Complex (pMHC) epitope can be identified in mice and tracked throughout the course of an immune response. The methodology involves the adoptive transfer of T-cell receptor (TCR) transgenic T cells with defined epitope specificity into histocompatible mice and the subsequent detection of these cells through the use of congenic or clonotypic markers. Alternatively, endogenous epitope-specific T cells can be tracked directly through the use of pMHC tetramers. Using magnetic bead-based enrichment and advanced multi-parameter flow cytometry, populations as small as 5 epitope-specific T cells can be detected from the peripheral lymphoid organs of a mouse. The adoptive transfer procedure can be completed within 3 h, while analysis of epitope-specific cells from mice can be completed within 6 h.