CD8 + T cells (TCD8) confer protective immunity against many infectious diseases, suggesting that microbial TCD8 determinants are promising vaccine targets. Nevertheless, current T cell antigen identification approaches do not discern which epitopes drive protective immunity during active infection -information that is critical for the rational design of TCD8-targeted vaccines. We employed a proteomics-based approach for large-scale discovery of naturally processed determinants derived from a complex pathogen, vaccinia virus (VACV), that are presented by the most frequent representatives of four major HLA class I supertypes. Immunologic characterization revealed that many previously unidentified VACV determinants were recognized by smallpox-vaccinated human peripheral blood cells in a variegated manner. Many such determinants were recognized by HLA class I-transgenic mouse immune TCD8 too and elicited protective TCD8 immunity against lethal intranasal VACV infection. Notably, efficient processing and stable presentation of immune determinants as well as the availability of naive TCD8 precursors were sufficient to drive a multifunctional, protective TCD8 response. Our approach uses fundamental insights into T cell epitope processing and presentation to define targets of protective TCD8 immunity within human pathogens that have complex proteomes, suggesting that this approach has general applicability in vaccine sciences.

The nature and anatomic location of the protective memory CD8+ T cell subset induced by intranasal vaccination remain poorly understood. We developed a vaccination model to assess the anatomic location of protective memory CD8+ T cells and their role in lower airway infections. Memory CD8+ T cells elicited by local intranasal, but not systemic vaccination with an engineered non-replicative CD8+ T cell-targeted antigen confer enhanced protection to a lethal respiratory viral challenge. This protection depends on a distinct CXCR3LO resident memory CD8+ T cell (Trm) population that preferentially localizes to the pulmonary interstitium. Interstitial Trm —by being positioned close to the mucosa where infection occurs—act before inflammation can recruit circulating memory CD8+ T cells into the lung tissue. This results in a local protective immune response as early as one day post-infection. Hence, vaccine strategies that induce lung interstitial Trm may confer better protection against respiratory pathogens

T lineage commitment occurs in a discrete, stage-specific manner during thymic ontogeny. Intrathymic precursor transfer experiments and the identification of CD4 ؉ 8 ؉ double-positive (DP), V␣14J␣18 natural T (iNKT) cells suggest that commitment to this lineage might occur at the DP stage. Nevertheless, this matter remains contentious because others failed to detect V␣14J␣18-positive iNKT cells that are CD4 ؉ 8 ؉ . In resolution to this issue, we demonstrate that retinoic acid receptor-related orphan receptor ␥ (ROR␥) 0/0 thymi, which accumulate immature single-positive (ISP) thymocytes that precede the DP stage, do not rearrange V␣14-to-J␣18 gene segments, suggesting that this event occurs at a post-ISP stage. Mixed radiation bone marrow chimeras revealed that ROR␥ functions in an iNKT cell lineage-specific manner. Further, introgression of a Bcl-x L transgene into ROR␥ 0/0 mice, which promotes survival and permits secondary rearrangements of distal V␣ and J␣ gene segments at the DP stage, rescues V␣14-to-J␣18 recombination. Similarly, introgression of a rearranged V␣14J␣18 transgene into ROR␥ 0/0 mice results in functional iNKT cells. Thus, our data support the ''T cell receptor-instructive (mainstream precursor) model'' of iNKT cell lineage specification where V␣14-to-J␣18 rearrangement, positive selection, and iNKT cell lineage commitment occur at or after the DP stage of ontogeny.ROR␥ ͉ V␣14-to-J␣18 rearrangement ͉ lineage specification ͉ T cell ontogeny C ommitment and differentiation to T lineage lymphocytes occur in the thymus gland from a pluripotent lymphocyte progenitor. The T lineage consists predominantly of ␣␤ T cells and a minor population of ␥␦ cells. Amongst the ␣␤ T cells are the conventional CD4 ϩ 8 Ϫ and CD4 Ϫ 8 ϩ lymphocytes as well as several minor subsets, including CD4 ϩ 25 ϩ T reg and NK1.1 ϩ natural T cells. Lineage decision (i.e., the decision to become ␥␦ or ␣␤) occurs early, whereas commitment to CD4 ϩ 8 Ϫ and CD4 Ϫ 8 ϩ lineages, which are specified by interactions with antigen-presenting molecules, occurs late in thymic ontogeny. These cell-fate specifications during thymic ontogeny of T lymphocytes depend on specific environmental cues, which signal heritable activation and repression of genes.Natural T cells comprise a heterogeneous subset amongst which V␣14J␣18 T (iNKT) lymphocytes predominate. They are CD1d-restricted, innate-like T lymphocytes that express a restricted T cell receptor (TCR) repertoire, which consists of the invariant V␣14J␣18 ␣-chain that predominantly pairs with the V␤8.2 ␤-chain (1). iNKT cell function, albeit elusive, appears to be immunoregulatory in nature (2, 3). Its ontogeny proceeds through the very same early stages (CD3 Ϫ 4 Ϫ 8 Ϫ triple-negative, TN1-4) as do the developing conventional T lymphocytes (4-9). Although it is contentious at which stage commitment to iNKT cell lineage occurs (discussed below), positive selection of this T cell subset relies on the interaction of precursor cells with CD1d1 expressed by CD4 ϩ 8 ϩ double-positive (DP) thymocy...

Endoplasmic reticulum (ER)-associated aminopeptidase (ERAP)1 has been implicated in the final proteolytic processing of peptides presented by major histocompatibility complex (MHC) class I molecules. To evaluate the in vivo role of ERAP1, we have generated ERAP1-deficient mice. Cell surface expression of the class Ia molecules H-2Kb and H-2Db and of the class Ib molecule Qa-2 was significantly reduced in these animals. Although cells from mutant animals exhibited reduced capacity to present several self- and foreign antigens to Kb-, Db-, or Qa-1b–restricted CD8+ cytotoxic T cells, presentation of some antigens was unaffected or significantly enhanced. Consistent with these findings, mice generated defective CD8+ T cell responses against class I–presented antigens. These findings reveal an important in vivo role of ER-associated peptidase activity in tailoring peptides for presentation by MHC class Ia and class Ib molecules.

Type I natural killer T (NKT) cells are innate-like T lymphocytes that recognize glycolipid antigens presented by the MHC class I-like protein CD1d. Agonistic activation of NKT cells leads to rapid pro-inflammatory and immune modulatory cytokine and chemokine responses. This property of NKT cells, in conjunction with their interactions with antigen-presenting cells, controls downstream innate and adaptive immune responses against cancers and infectious diseases, as well as in several inflammatory disorders. NKT cell properties are acquired during development in the thymus and by interactions with the host microbial consortium in the gut, the nature of which can be influenced by NKT cells. This latter property, together with the role of the host microbiota in cancer therapy, necessitates a new perspective. Hence, this review provides an initial approach to understanding NKT cells from an ecological evolutionary developmental biology (eco-evo-devo) perspective.

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Invariant natural killer T (iNKT) cell-derived cytokines have important functions in inflammation, host defense, and immunoregulation. Yet, when and how iNKT cells undergo effector differentiation, which endows them with the capacity to rapidly secrete cytokines upon activation, remains unknown. We discovered that granulocyte-macrophage colony-stimulating factor (Csf-2)-deficient mice developed iNKT cells that failed to respond to the model antigen alpha-galactosylceramide because of an intrinsic defect in the fusion of secretory vesicles with the plasma membrane. Exogenous Csf-2 corrected the functional defect only when supplied during the development of thymic, but not mature, splenic Csf-2-deficient iNKT cells. Thus, we ascribe a unique function to Csf-2, which regulates iNKT cell effector differentiation during development by a mechanism that renders them competent for cytokine secretion.

Endoplasmic reticulum (ER)-associated aminopeptidase (ERAP)1 has been implicated in the fi nal proteolytic processing of peptides presented by major histocompatibility complex (MHC) class I molecules. To evaluate the in vivo role of ERAP1, we have generated ERAP1-defi cient mice. Cell surface expression of the class Ia molecules H-2K b and H-2D b and of the class Ib molecule Qa-2 was signifi cantly reduced in these animals. Although cells from mutant animals exhibited reduced capacity to present several self-and foreign antigens to K b-, D b-, or Qa-1 b-restricted CD8 + cytotoxic T cells, presentation of some antigens was unaffected or signifi cantly enhanced. Consistent with these fi ndings, mice generated defective CD8 + T cell responses against class I-presented antigens. These fi ndings reveal an important in vivo role of ER-associated peptidase activity in tailoring peptides for presentation by MHC class Ia and class Ib molecules.