Marielle C. Gold

Human Mucosal Associated Invariant T Cells Detect Bacterially Infected Cells

A first indication of the biological role of mucosal associated invariant T (MAIT) cells reveals that this discrete T cell subset is broadly reactive to bacterial infection. In particular MAIT cells recognize Mycobacterium tuberculosis-infected lung airway epithelial cells via the most evolutionarily conserved major histocompatibility molecule.

Assembly and antigen-presenting function of MHC class I molecules in cells lacking the ER chaperone calreticulin

MHC class I molecules expressed in a calreticulin-deficient cell line (K42) assembled with beta 2-microglobulin (beta2-m) normally, but their subsequent loading with optimal peptides was defective. Suboptimally loaded class I molecules were released into the secretory pathway. This occurred despite the ability of newly synthesized class I to interact with the transporter associated with antigen processing (TAP) loading complex. The efficiency of peptide loading was reduced by 50%-80%, and impaired T cell recognition was observed for three out of four antigens tested. The peptide-loading function was specific to calreticulin, since the defect in K42 could be rectified by transfection with calreticulin but not a soluble form of calnexin, which shares its lectin-like activity.

Genome-Wide Analysis Reveals a Highly Diverse CD8 T Cell Response to Murine Cytomegalovirus

Human CMV establishes a lifelong latent infection in the majority of people worldwide. Although most infections are asymptomatic, immunocompetent hosts devote an extraordinary amount of immune resources to virus control. To increase our understanding of CMV immunobiology in an animal model, we used a genomic approach to comprehensively map the C57BL/6 CD8 T cell response to murine CMV (MCMV). Responses to 27 viral proteins were detectable directly ex vivo, the most diverse CD8 T cell response yet described within an individual animal. Twenty-four peptide epitopes were mapped from 18 Ags, which together account for most of the MCMV-specific response. Most Ags were from genes expressed at early times, after viral genes that interfere with Ag presentation are expressed, consistent with the hypothesis that the CD8 T cell response to MCMV is largely driven by cross-presented Ag. Titration of peptide epitopes in a direct ex vivo intracellular cytokine staining assay revealed a wide range of functional avidities, with no obvious correlation between functional avidity and the strength of the response. The immunodominance hierarchy varied only slightly between mice and between experiments. However, H-2b-expressing mice with different genetic backgrounds responded preferentially to different epitopes, indicating that non-MHC-encoded factors contribute to immunodominance in the CD8 T cell response to MCMV.

The murine cytomegalovirus immunomodulatory gene m152 prevents recognition of infected cells by M45-specific CTL but does not alter the immunodominance of the M45-specific CD8 T cell response in vivo.

Although in vitro studies have shown that herpesviruses, including murine CMV (MCMV), encode genes that interfere with the MHC class I pathway, their effects on the CTL response in vivo is unclear. We identified a Db-restricted CTL epitope from MCMV M45 by screening an MCMV genomic library using CTL clones isolated from mice infected with MCMV lacking m152. Because m152 severely inhibits CTL recognition of M45 in vitro, we questioned whether an M45-specific response would be generated in mice infected with wild-type MCMV expressing m152. Mice infected with wild-type MCMV or MCMVΔm152 made similar responses to the M45 Ag. Moreover, we saw no skewing of the proportion of M45-specific CD8 T cells within the total MCMV-specific response after infection with MCMV with m152. Despite the profound effect m152 has on presentation of M45 in vitro, it does not affect the immunodominance of M45 in the CTL response in vivo.

MR1-restricted MAIT cells display ligand discrimination and pathogen selectivity through distinct T cell receptor usage

MAIT cells can discriminate between pathogen-derived ligands in a clonotype-dependent manner, and the TCR repertoire is distinct within individuals, indicating that the MAIT cell repertoire is shaped by prior microbial exposure.

Human thymic MR1-restricted MAIT cells are innate pathogen- reactive effectors that adapt following thymic egress

Human mucosal-associated invariant T (MAIT) cells express the semi-invariant T-cell receptor (TCR) Vα7.2 and are restricted by the major histocompatibility complex-Ib molecule MR1. While MAIT cells share similarities with other innate T cells, the extent to which MAIT cells are innate and their capacity to adapt is unknown. We evaluated the function of Vα7.2+ T cells from the thymus, cord blood, and peripheral blood. Although antigen-inexperienced MAIT cells displayed a naïve phenotype, these had intrinsic effector capacity in response to Mycobacterium tuberculosis (Mtb)-infected cells. Vα7.2+ effector thymocytes contained signal joint TCR gene excision circles (sjTRECs) suggesting limited replication and thymic origin. In evaluating the capacity of Mtb-reactive MAIT cells to adapt, we found that those from the peripheral blood demonstrated a memory phenotype and had undergone substantial expansion, suggesting that they responded to antigenic stimulation. MAIT cells, an evolutionarily conserved T-cell subset that detects a variety of intracellular infections, share features of innate and adaptive immunity.

Co-dependents: MR1-restricted MAIT cells and their antimicrobial function

Mucosal-associated invariant T (MAIT) cells are a unique T cell subset in mammals. They are present at high frequencies at mucosal tissue sites and have an intrinsic capacity to respond to microbial infections. The semi-invariant antigen recognition receptor of MAIT cells detects the non-polymorphic antigen-presenting molecule major histocompatibility complex class I-related protein 1 (MR1), which can bind microorganism-derived riboflavin metabolites. The striking evolutionary conservation in both the MR1 molecule and the MAIT T cell receptor suggests that strong selective pressures maintain this T cell pattern recognition system which detects microbial infection.

Murine Cytomegalovirus Interference with Antigen Presentation Has Little Effect on the Size or the Effector Memory Phenotype of the CD8 T Cell Response

As with most herpesviruses, CMVs encode viral genes that inhibit Ag presentation to CD8 T cells (VIPRs). VIPR function has been assumed to be essential for CMV to establish its characteristic lifetime infection of its host. We compared infection of C57BL/6 mice with wild-type murine CMV (MCMV) and a virus lacking each of MCMV’s three known VIPRs: m4, m6, and m152. During acute infection, there was very little difference between the two viruses with respect to the kinetics of viral replication and clearance, or in the size and kinetics of the virus-specific CD8 T cell response. During chronic infection, a large, effector memory, virus-specific CD8 T cell population (CD8lowCD62L−CD11c+NKG2A+) was maintained in both infections; the size and phenotype of the CD8 T cell response to both viruses was remarkably similar. The characteristic effector memory phenotype of the CD8 T cells suggested that both wild-type and Δm4+m6+m152 virus continued to present Ag to CD8 T cells during the chronic phase of infection. During the chronic phase of infection, MCMV cannot be isolated from immunocompetent mice. However, upon immunosuppression, both Δm4+m6+m152 and wild-type virus could be reactivated from mice infected for 6 wk. Thus, restoring the ability of CD8 T cells to detect MCMV had little apparent effect on the course of MCMV infection and on the CD8 T cell response to it. These results challenge the notion that VIPR function is necessary for CMV persistence in the host.

Human Lung Epithelial Cells Contain Mycobacterium tuberculosis in a Late Endosomal Vacuole and Are Efficiently Recognized by CD8+ T Cells

Mycobacterium tuberculosis (Mtb) is transmitted via inhalation of aerosolized particles. While alveolar macrophages are thought to play a central role in the acquisition and control of this infection, Mtb also has ample opportunity to interact with the airway epithelium. In this regard, we have recently shown that the upper airways are enriched with a population of non-classical, MR1-restricted, Mtb-reactive CD8+ T cells (MAIT cells). Additionally, we have demonstrated that Mtb-infected epithelial cells lining the upper airways are capable of stimulating IFNγ production by MAIT cells. In this study, we demonstrate that airway epithelial cells efficiently stimulate IFNγ release by MAIT cells as well as HLA-B45 and HLA-E restricted T cell clones. Characterization of the intracellular localization of Mtb in epithelial cells indicates that the vacuole occupied by Mtb in epithelial cells is distinct from DC in that it acquires Rab7 molecules and does not retain markers of early endosomes such as Rab5. The Mtb vacuole is also heterogeneous as there is a varying degree of association with Lamp1 and HLA-I. Although the Mtb vacuole shares markers associated with the late endosome, it does not acidify, and the bacteria are able to replicate within the cell. This work demonstrates that Mtb infected lung epithelial cells are surprisingly efficient at stimulating IFNγ release by CD8+ T cells.

The Role of Mucosal Associated Invariant T Cells in Antimicrobial Immunity.

Mucosal associated invariant T (MAIT) cells are an innate-like T cell subset prevalent in humans and distributed throughout the blood and mucosal sites. Human MAIT cells are defined by the expression of the semi-invariant TCRα chain TRAV1-2/TRAJ12/20/33 and are restricted by the non-polymorphic major histocompatibility complex (MHC) class I-like molecule, MHC-related protein 1, MR1. MAIT cells are activated by small organic molecules, derived from the riboflavin biosynthesis pathway of bacteria and fungi, presented by MR1. Traditionally, MAIT cells were thought to recognize a limited number of antigens due to usage of an invariant TCRα chain and restriction by a non-polymorphic MHC molecule. However, recent studies demonstrate that the TCR repertoire of MAIT cells is more heterogeneous, suggesting there is a more diverse array of MR1 antigens that MAIT cells can recognize. In response to infected cells, MAIT cells produce the pro-inflammatory cytokines, IFN-γ and TNF, and are cytolytic. Studies performed in MR1-deficient mice suggest that MAIT cells can provide anti-bacterial control within the first few days post-infection, as well as contribute to enhanced adaptive immunity in murine models of respiratory infections. In humans, the role of MAIT cells is unclear; however, evidence points to interplay between MAIT cells and microbial infections, including Mycobacterium tuberculosis. Given that MAIT cells are pro-inflammatory, serve in early control of bacterial infections, and appear enriched at tissue sites where microbes interface and gain access to the body, we postulate that they play an important role in antimicrobial immune responses. In this review, we discuss the most recent studies on the function and phenotype of MAIT cells, including their TCR diversity and antigenic repertoire, with a focus on the contribution of human MAIT cells in the immune response to microbial infection.