Mucosal-associated invariant T (MAIT) cells recognize microbial compounds presented by the MHC-related 1 (MR1) protein. Although riboflavin precursor derivatives from Gram-positive bacteria have been characterized, some level of ligand heterogeneity has been suggested through the analysis of the MAIT cell TCR repertoire in humans and differential reactivity of human MAIT cell clones according to the bacteria. In this study, using Gram-negative bacteria mutated for the riboflavin biosynthetic pathway, we show a strict correlation between the ability to synthesize the 5-amino-ribityl-uracil riboflavin precursor and to activate polyclonal and quasi-monoclonal mouse MAIT cells. To our knowledge, we show for the first time that the semipurified bacterial fraction and the synthetic ligand activate murine MAIT cells in vitro and in vivo. We describe new MR1 ligands that do not activate MAIT cells but compete with bacterial and synthetic compounds activating MAIT cells, providing the capacity to modulate MAIT cell activation. Through competition experiments, we show that the most active synthetic MAIT cell ligand displays the same functional avidity for MR1 as does the microbial compound. Altogether, these results show that most, if not all, MAIT cell ligands found in Escherichia coli are related to the riboflavin biosynthetic pathway and display very limited heterogeneity.

Direct interactions between bacterial and host glycans have been recently reported to be involved in the binding of pathogenic bacteria to host cells. In the case of Shigella, the Gram-negative enteroinvasive bacterium responsible for acute rectocolitis, such interactions contribute to bacterial adherence to epithelial cells. However, the role of glycans in the tropism of Shigella for immune cells whose glycosylation pattern varies depending on their activation state is unknown. We previously reported that Shigella targets activated, but not nonactivated, human CD4+ T lymphocytes. Here, we show that nonactivated CD4+ T lymphocytes can be turned into Shigella-targetable cells upon loading of their plasma membrane with sialylated glycosphingolipids (also termed gangliosides). The Shigella targeting profile of ganglioside-loaded nonactivated T cells is similar to that of activated T cells, with a predominance of injection of effectors from the type III secretion system (T3SS) not resulting in cell invasion. We demonstrate that gangliosides interact with the O-antigen polysaccharide moiety of lipopolysaccharide (LPS), the major bacterial surface antigen, thus promoting Shigella binding to CD4+ T cells. This binding step is critical for the subsequent injection of T3SS effectors, a step which we univocally demonstrate to be dependent on actin polymerization. Altogether, these findings highlight the critical role of glycan-glycan interactions in Shigella pathogenesis.

The enteropathogen, Shigella, is highly virulent and remarkably adjusted to the intestinal environment of its almost exclusive human host. Key for Shigella pathogenicity is the injection of virulence effectors into the host cell via its type three secretion system (T3SS), initiating disease onset and progression by the vast diversity of the secreted T3SS effectors and their respective cellular targets. The multifaceted modulation of host signaling pathways exerted by Shigella T3SS effectors, which include the subversion of host innate immune defenses and the promotion of intracellular bacterial survival and dissemination, have been extensively reviewed in the recent past. This review focuses on the human species specificity of Shigella by discussing some possible evasion mechanisms towards the human, but not non-human or rodent gut innate defense barrier, leading to the lack of a relevant animal infection model. In addition, subversion mechanisms of the adaptive immune response are highlighted summarizing research advances of the recent years. In particular, the new paradigm of Shigella pathogenicity constituted of invasion-independent T3SS effector-mediated targeting of activated, human lymphocytes is discussed. Along with consequences on vaccine development, these findings offer new directions for future research endeavors towards a better understanding of immunity to Shigella infection.

The enteroinvasive bacterium Shigella is a facultative intracellular bacterium known, in vitro, to invade a large diversity of cells through the delivery of virulence effectors into the cell cytoplasm via a type III secretion system (T3SS). Here, we provide evidence that the injection of T3SS effectors does not necessarily result in cell invasion. Indeed, we demonstrate through optimization of a T3SS injection reporter that effector injection without subsequent cell invasion, termed the injection-only mechanism, is the main strategy used by Shigella to target human immune cells. We show that in vitro-activated human peripheral blood B, CD4+ T, and CD8+ T lymphocytes as well as switched memory B cells are mostly targeted by the injection-only mechanism. B and T lymphocytes residing in the human colonic lamina propria, encountered by Shigella upon its crossing of the mucosal barrier, are also mainly targeted by injection-only. These findings reveal that cells refractory to invasion can still be injected, thus extending the panel of host cells manipulated to the benefit of the pathogen. Future analysis of the functional consequences of the injection-only mechanism toward immune cells will contribute to the understanding of the priming of adaptive immunity, which is known to be altered during the course of natural Shigella infection.Shigella | T3SS | bacterial effectors | human lymphocytes | host-pathogen cross talk

In type 1 diabetes, autoimmune β-cell destruction may be favored by neoantigens harboring posttranslational modifications (PTMs) such as citrullination. We studied the recognition of native and citrullinated glucose-regulated protein (GRP)78 peptides by CD8+ T cells. Citrullination modulated T-cell recognition and, to a lesser extent, HLA-A2 binding. GRP78-reactive CD8+ T cells circulated at similar frequencies in healthy donors and donors with type 1 diabetes and preferentially recognized either native or citrullinated versions, without cross-reactivity. Rather, the preference for native GRP78 epitopes was associated with CD8+ T cells cross-reactive with bacterial mimotopes. In the pancreas, a dominant GRP78 peptide was instead preferentially recognized when citrullinated. To further clarify these recognition patterns, we considered the possibility of citrullination in the thymus. Citrullinating peptidylarginine deiminase (Padi) enzymes were expressed in murine and human medullary epithelial cells (mTECs), with citrullinated proteins detected in murine mTECs. However, Padi2 and Padi4 expression was diminished in mature mTECs from NOD mice versus C57BL/6 mice. We conclude that, on one hand, the CD8+ T cell preference for native GRP78 peptides may be shaped by cross-reactivity with bacterial mimotopes. On the other hand, PTMs may not invariably favor loss of tolerance because thymic citrullination, although impaired in NOD mice, may drive deletion of citrulline-reactive T cells.

In type 1 diabetes, autoimmune β-cell destruction may be favored by neo-antigens harboring post-translational modifications such as citrullination. We studied the recognition of native and citrullinated glucose-regulated protein (GRP)78 peptides by CD8⁺ T cells. Citrullination modulated T-cell recognition and, to a lesser extent, HLA-A2 binding. GRP78-reactive CD8⁺ T cells circulated at similar frequencies in type 1 diabetic and healthy donors and preferentially recognized either native or citrullinated versions, without cross-reactivity. Rather, the preference for native GRP78 epitopes was associated with CD8⁺ T cells cross-reactive with bacterial mimotopes. In the pancreas, a dominant GRP78 peptide was instead preferentially recognized when citrullinated. To further clarify these recognition patterns, we considered the possibility of citrullination in the thymus. Citrullinating peptidyl-arginine deiminase (Padi) enzymes were expressed in murine and human medullary epithelial cells (mTECs), with citrullinated proteins detected in murine mTECs. However, Padi2 and Padi4 expression was diminished in mature mTECs from NOD mice versus C57BL/6 mice. We conclude that, on one hand, the CD8⁺ T-cell preference for native GRP78 peptides may be shaped by cross-reactivity with bacterial mimotopes. On the other hand, post-translational modifications may not invariably favor loss of tolerance because thymic citrullination, although impaired in NOD mice, may drive deletion of citrulline-reactive T cells.

Strategies employed by pathogenic enteric bacteria, such as Shigella, to subvert the host adaptive immunity are not well defined. Impairment of T lymphocyte chemotaxis by blockage of polarised edge formation has been reported upon Shigella infection. However, the functional impact of Shigella on T lymphocytes remains to be determined. Here, we show that Shigella modulates CD4+ T cell F‐actin dynamics and increases cell cortical stiffness. The scanning ability of T lymphocytes when encountering antigen‐presenting cells (APC) is subsequently impaired resulting in decreased cell–cell contacts (or conjugates) between the two cell types, as compared with non‐infected T cells. In addition, the few conjugates established between the invaded T cells and APCs display no polarised delivery and accumulation of the T cell receptor to the contact zone characterising canonical immunological synapses. This is most likely due to the targeting of intracellular vesicular trafficking by the bacterial type III secretion system (T3SS) effectors IpaJ and VirA. The collective impact of these cellular reshapings by Shigella eventually results in T cell activation dampening. Altogether, these results highlight the combined action of T3SS effectors leading to T cell defects upon Shigella infection.

Purpose of reviewRecent work using immunopeptidomics and deconvolution of the antigenic reactivity of islet-infiltrating CD8þ T cells has expanded our knowledge about the autoimmune target epitopes of type 1 diabetes. The stem-like properties of autoimmune CD8þ T cells have also been described. We here propose a possible link between these findings. Recent findingsWeak major histocompatibility complex (MHC)-binding epitopes list among the major targets of human islet-infiltrating CD8þ T cells, likely resulting in low peptide-MHC presentation that delivers weak T-cell receptor (TCR) signals, especially in the face of low-affinity autoimmune TCRs. These weak TCR signals may favor the maintenance of the partially differentiated stem-like phenotype recently described for islet-reactive CD8þ T cells in the blood and pancreatic lymph nodes. These weak TCR signals may also be physiological, reflecting the need for self-peptide-MHC contacts to maintain homeostatic T-cell survival and proliferation. These features may underlie the universal state of benign autoimmunity that we recently described, which is characterized by islet-reactive, na € ive-like CD8þ T cells circulating in all individuals.