SUMMARY Selectins play a central role in leukocyte trafficking by mediating tethering and rolling on vascular surfaces. Here we have reported that T cell immunoglobulin and mucin domain 1 (TIM-1) is a P-selectin ligand. We have shown that human and murine TIM-1 binds to P-selectin, and that TIM-1 mediates tethering and rolling of T helper-1 (Th1) and Th17, but not Th2 and regulatory T cells on P-selectin. Th1 and Th17 cells lacking the TIM-1 mucin domain showed reduced rolling in thrombin-activated mesenteric venules and inflamed brain microcirculation. Inhibition of TIM-1 had no effect on naive T cell homing, but reduced T cell recruitment in a skin hypersensitivity model and blocked experimental autoimmune encephalomyelitis. Uniquely, the TIM-1 IgV domain was also required for P-selectin binding. Our data demonstrate that TIM-1 is a major P-selectin ligand with a specialized role in T cell trafficking during inflammatory responses and the induction of autoimmune disease.

SummaryCD4+ T cells develop distinct and often contrasting helper, regulatory, or cytotoxic activities. Typically a property of CD8+ T cells, granzyme-mediated cytotoxic T cell (CTL) potential is also exerted by CD4+ T cells. However, the conditions that induce CD4+ CTLs are not entirely understood. Using single-cell transcriptional profiling, we uncover a unique signature of Granzyme B (GzmB)+ CD4+ CTLs, which distinguishes them from other CD4+ T helper (Th) cells, including Th1 cells, and strongly contrasts with the follicular helper T (Tfh) cell signature. The balance between CD4+ CTL and Tfh differentiation heavily depends on the class of infecting virus and is jointly regulated by the Tfh-related transcription factors Bcl6 and Tcf7 (encoding TCF-1) and by the expression of the inhibitory receptors PD-1 and LAG3. This unique profile of CD4+ CTLs offers targets for their study, and its antagonism by the Tfh program separates CD4+ T cells with either helper or killer functions.

CD4+ T cell differentiation is influenced by a plethora of intrinsic and extrinsic factors, providing the immune system with the ability to tailor its response according to specific stimuli. Indeed, different classes of pathogens may induce a distinct balance of CD4+ T cell differentiation programmes. Here, we report an uncommonly strong bias toward follicular helper (Tfh) differentiation of CD4+ T cells reactive with a retroviral envelope glycoprotein model antigen, presented in its natural context during retroviral infection. Conversely, the response to the same antigen, presented in different immunization regimens, elicited a response typically balanced between Tfh and T helper 1 cells. Comprehensive quantitation of variables known to influence Tfh differentiation revealed the closest correlation with the strength of T cell receptor (TCR) signaling, leading to PD-1 expression, but not with surface TCR downregulation, irrespective of TCR clonotypic avidity. In contrast, strong TCR signaling leading to TCR downregulation and induction of LAG3 expression in high TCR avidity clonotypes restrained CD4+ T cell commitment and further differentiation. Finally, stunted Th1 differentiation, correlating with limited IL-2 availability in retroviral infection, provided permissive conditions for Tfh development, suggesting that Tfh differentiation is the default program of envelope-reactive CD4+ T cells.

CD4+ T cells are typically considered as ‘helper’ or ‘regulatory’ populations that support and orchestrate the responses of other lymphocytes. However, they can also develop potent granzyme (Gzm)-mediated cytotoxic activity and CD4+ cytotoxic T cells (CTLs) have been amply documented both in humans and in mice, particularly in the context of human chronic infection and cancer. Despite the established description of CD4+ CTLs, as well as of the critical cytotoxic activity they exert against MHC class II-expressing targets, their developmental and memory maintenance requirements remain elusive. This is at least in part owing to the lack of a murine experimental system where CD4+ CTLs are stably induced. Here, we show that viral and bacterial vectors encoding the same epitope induce distinct CD4+ CTL responses in challenged mice, all of which are nevertheless transient in nature and lack recall properties. Consistent with prior reports, CD4+ CTL differentiation is accompanied by loss of TCF-1 expression, a transcription factor considered essential for memory T cell survival. Using genetic ablation of Tcf7, which encodes TCF-1, at the time of CD4+ T cell activation, we further show that, contrary to observations in CD8+ T cells, continued expression of TCF-1 is not required for CD4+ T cell memory survival. Whilst Tcf7-deficient CD4+ T cells persisted normally following retroviral infection, the CD4+ CTL subset still declined, precluding conclusive determination of the requirement for TCF-1 for murine CD4+ CTL survival. Using xenotransplantation of human CD4+ T cells into murine recipients, we demonstrate that human CD4+ CTLs develop and persist in the same experimental conditions where murine CD4+ CTLs fail to persist. These observations uncover a species-specific defect in murine CD4+ CTL persistence with implications for their use as a model system.

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