Graphical AbstractHighlights d Mouse and human tumors harbor relatively undifferentiated Tcf1 + PD-1 + CD8 + T cells d These intratumoral cells have expansion, regeneration, and differentiation capacity d They produce differentiated Tcf1 À PD-1 + CD8 + T cells in response to immunotherapy d These stem-like cells are critical for tumor control in response to immunotherapy In BriefSince chronic activation promotes terminal T cell differentiation (exhaustion), it has remained unclear how checkpoint blockade mediates a proliferative response of tumorinfiltrating T cells. Siddiqui et al. identify intratumoral, tumor-reactive Tcf1 + PD-1 + CD8 + T cells that display stem-like properties and that promote tumor control in response to vaccination and checkpoint blockade immunotherapy. SUMMARYCheckpoint blockade mediates a proliferative response of tumor-infiltrating CD8 + T lymphocytes (TILs). The origin of this response has remained elusive because chronic activation promotes terminal differentiation or exhaustion of tumor-specific T cells. Here we identified a subset of tumor-reactive TILs bearing hallmarks of exhausted cells and central memory cells, including expression of the checkpoint protein PD-1 and the transcription factor Tcf1. Tcf1 + PD-1 + TILs mediated the proliferative response to immunotherapy, generating both Tcf1 + PD-1 + and differentiated Tcf1 À PD-1 + cells. Ablation of Tcf1 + PD-1 + TILs restricted responses to immunotherapy. Tcf1 was not required for the generation of Tcf1 + PD-1 + TILs but was essential for the stem-like functions of these cells. Human TCF1 + PD-1 + cells were detected among tumor-reactive CD8 + T cells in the blood of melanoma patients and among TILs of primary melanomas. Thus, immune checkpoint blockade relies not on reversal of T cell exhaustion programs, but on the proliferation of a stem-like TIL subset.

Chronic infections promote the terminal differentiation (or "exhaustion") of T cells and are thought to preclude the formation of memory T cells. In contrast, we discovered a small subpopulation of virus-specific CD8(+) T cells that sustained the T cell response during chronic infections. These cells were defined by, and depended on, the expression of the transcription factor Tcf1. Transcriptome analysis revealed that this population shared key characteristics of central memory cells but lacked an effector signature. Unlike conventional memory cells, Tcf1-expressing T cells displayed hallmarks of an "exhausted" phenotype, including the expression of inhibitory receptors such as PD-1 and Lag-3. This population was crucial for the T cell expansion that occurred in response to inhibitory receptor blockade during chronic infection. These findings identify a memory-like T cell population that sustains T cell responses and is a prime target for therapeutic interventions to improve the immune response in chronic infections.

γδ T cells are an important innate source of interleukin-17 (IL-17). In contrast to T helper 17 (Th17) cell differentiation, which occurs in the periphery, IL-17-producing γδ T cells (γδT17 cells) are probably committed during thymic development. To study when γδT17 cells arise during ontogeny, we used TcrdH2BeGFP reporter mice to monitor T cell receptor (TCR) rearrangement and IL-17 production in the embryonic thymus. We observed that several populations such as innate lymphoid cells and early T cell precursors were able to produce IL-17 prior to (and thus independent of) TCR recombination. γδT17 cells were absent after transplantation of IL-17-sufficient bone marrow into mice lacking both Il17a and Il17f. Also, γδT17 cells were not generated after genetic restoration of defective Rag1 function in adult mice. Together, these data suggested that these cells developed exclusively before birth and subsequently persisted in adult mice as self-renewing, long-lived cells.

cd T cells are a potent source of innate IL-17A and IFN-c, and they acquire the capacity to produce these cytokines within the thymus. However, the precise stages and required signals that guide this differentiation are unclear. Here we show that the CD24 low CD44 high effector cd T cells of the adult thymus are segregated into two lineages by the mutually exclusive expression of CCR6 and NK1.1. Only CCR6 1 cd T cells produced IL-17A, while NK1.1 1 cd T cells were efficient producers of IFN-c but not of IL-17A. Their effector phenotype correlated with loss of CCR9 expression, particularly among the NK1.1 1 cd T cells. Accordingly, both cd T-cell subsets were rare in gut-associated lymphoid tissues, but abundant in peripheral lymphoid tissues. There, they provided IL-17A and IFN-c in response to TCR-specific and TCR-independent stimuli. IL-12 and IL-18 induced IFN-c and IL-23 induced IL-17A production by NK1.1 1 or CCR6 1 cd T cells, respectively. Importantly, we show that CCR6 1 cd T cells are more responsive to TCR stimulation than their NK1.1 1 counterparts. In conclusion, our findings support the hypothesis that CCR6 1 IL-17A-producing cd T cells derive from less TCR-dependent selection events than IFN-c-producing NK1.1 1 cd T cells.Key words: gd T cells . CCR6 . IFN-g . IL-17A . Innate lymphocytes . NK1.1 Introduction IL-17A and IFN-g are generally regarded as pro-inflammatory effector cytokines that can be produced by Th cells but also by innate lymphocytes such as NK cells, NKT cells and gd T cells. While macrophage activation is supposed to be the main role of IFN-g, the induction of granulopoiesis is ascribed as a key biological function of IL-17A [1]. It is currently emerging that gd T cells are a potent source of IL-17A in the early phases of immune responses (reviewed in [2]). gd T cells constitute a large fraction of all IL-17A-producing cells in healthy mice and humans and are able to secrete IL-17A much more rapidly than CD4 1 Th17 cells [3][4][5]. These observations led to the concept that gd T cells are important players in a transitional response between innate and adaptive immune reactions [6]. The production of innate IL-17A by gd T cells appears to be essential in situations where an effective defence against extra-cellular bacteria or fungi relies on the fast mobilization of neutrophils [4,[6][7][8][9]. Moreover, IL-17A-producing gd T cells have been described to play important roles in immunopathologic diseases such as collageninduced arthritis [10], experimental pulmonary fibrosis [11], and in experimental autoimmune encephalitis [12,13].In contrast to CD4 1 Th cells that can develop into Th17 cells after encounter of specific cognate TCR Ag [14][15][16], it is not clear which stimuli induce IL-17A production by gd T cells in vivo. This essentially results from a lack of information about physiological gd TCR ligands. However, the current literature suggests that the decision whether a gd T cell will produce IL-17A is linked to 3488thymic development. Jensen et al. introduced a concept th...