Translational regulation of TFH cell differentiation and autoimmune pathogenesis.

Little is known regarding T cell translational regulation. We demonstrate that T follicular helper (TFH) cells use a previously unknown mechanism of selective messenger RNA (mRNA) translation for their differentiation, role in B cell maturation, and in autoimmune pathogenesis. We show that TFH cells have much higher levels of translation factor eIF4E than non-TFH CD4+ T cells, which is essential for translation of TFH cell fate-specification mRNAs. Genome-wide translation studies indicate that modest down-regulation of eIF4E activity by a small-molecule inhibitor or short hairpin RN impairs TFH cell development and function. In mice, down-regulation of eIF4E activity specifically reduces TFH cells among T helper subtypes, germinal centers, B cell recruitment, and antibody production. In experimental autoimmune encephalomyelitis, eIF4E activity down-regulation blocks TFH cell participation in disease pathogenesis while promoting rapid remission and spinal cord remyelination. TFH cell development and its role in autoimmune pathogenesis involve selective mRNA translation that is highly druggable.

BATF acts as an essential regulator of IL-25-responsive migratory ILC2 cell fate and function.

A transitory, interleukin-25 (IL-25)-responsive, group 2 innate lymphoid cell (ILC2) subset induced during type 2 inflammation was recently identified as iILC2s. This study focuses on understanding the significance of this population in relation to tissue-resident nILC2s in the lung and intestine. RNA-sequencing and pathway analysis revealed the AP-1 superfamily transcription factor BATF (basic leucine zipper transcription factor, activating transcription factor-like) as a potential modulator of ILC2 cell fate. Infection of BATF-deficient mice with Nippostrongylus brasiliensis showed a selective defect in IL-25-mediated helminth clearance and a corresponding loss of iILC2s in the lung characterized as IL-17RBhigh, KLRG1high, BATFhigh, and Arg1low BATF deficiency selectively impaired iILC2s because it had no impact on tissue-resident nILC2 frequency or function. Pulmonary-associated iILC2s migrated to the lung after infection, where they represented an early source of IL-4 and IL-13. Although the composition of ILC2s in the small intestine was distinct from those in the lung, their frequency and IL-13 expression remained dependent on BATF, which was also required for optimal goblet and tuft cell hyperplasia. Findings support IL-25-responsive ILC2s as early sentinels of mucosal barrier integrity.

Library Preparation for ATAC-Sequencing of Mouse CD4+ T Cells Isolated from the Lung and Lymph Nodes After Helminth Infection.

Although conventional methods such as MNase-seq, DNase-seq, and ChIP-seq have been used effectively to assess chromatin and locus accessibility at the genome level, these techniques generally require large numbers of input cells. As such, much of what we understand in terms of epigenetic regulation and locus accessibility in CD4+ T cell subsets comes from in vitro culture systems, which allow for the production of large numbers of polarized T cells. However, obtaining such numbers directly ex vivo from tissues of individual mice is difficult. Here we describe a method combining cytokine reporter mice and Assay for Transposase-Accessible Chromatin with high-throughput sequencing (ATAC-seq) to identify genome wide locus accessibility in a small number of cytokine-expressing CD4+ T cells. This method takes you from cell isolation to library generation and quality control to query. Because the Il4 and Ifng loci are reciprocally regulated in polarized CD4+ T cell subsets (Th1 vs. Th2), we investigated the ability of this approach to identify transposase integration in both IL-4- and IFN-γ-expressing CD4+ T cells isolated directly from the lung and lymph nodes after helminth infection.

Irgm1 coordinately regulates autoimmunity and host defense at select mucosal surfaces.

The pathogenesis of primary Sjogren's syndrome (SS), an autoimmune disease that targets the mucosa of exocrine tissues, is poorly understood. Although several mouse models have been developed that display features of SS, most of these are within the larger context of a lupus-like presentation. Immunity-related GTPase family M protein 1 (Irgm1) is an interferon-inducible cytoplasmic GTPase that is reported to regulate autophagy and mitochondrial homeostasis. Here, we report that naive Irgm1-/- mice display lymphocytic infiltration of multiple mucosal tissues including the lung in a manner reminiscent of SS, together with IgA class-predominant autoantibodies including anti-Ro and anti-La. This phenotype persists in the germ-free state, but is abolished by deletion of Irgm3. Irgm1-/- mice have increased local production in the lung of TECP15-idiotype IgA, a natural antibody with dual reactivity against host and pneumococcal phosphorylcholine. Associated with this, Irgm1-/- mice display enhanced opsonization and clearance of Streptococcus pneumoniae from the lung and increased survival from pneumococcal pneumonia. Taken together, our results identify Irgm1 as a master regulator of mucosal immunity that dually modulates evolutionarily conserved self- and other-directed immune responses at the interface of host with environment.

CD36 and Platelet-Activating Factor Receptor Promote House Dust Mite Allergy Development.

Over 89% of asthmatic children in underdeveloped countries demonstrate sensitivity to house dust mites (HDMs). The allergic response to HDMs is partially mediated by epithelial cell-derived cytokines that activate group 2 innate lymphoid cells, induce migration and activation of dendritic cells, and promote effector differentiation of HDM-specific TH2 cells. However, the contribution of innate receptor engagement on epithelial or dendritic cells by HDMs that ultimately mediates said innate and adaptive allergic responses is poorly understood. We and other investigators have demonstrated that HDMs express phosphorylcholine (PC) moieties. The major PC receptors involved in immune responses include CD36 and platelet-activating factor receptor (PAFR). Because CD36 and PAFR are expressed by epithelial cells and dendritic cells, and expression of these receptors is higher in human asthmatics, we determined whether engagement of CD36 or PAFR on epithelial or dendritic cells contributes to HDM allergy development. Testing bone marrow chimeric mice revealed that CD36 engagement on radioresistant cells and PAFR engagement on radioresistant and radiosensitive cells in the lung promote allergic responses to HDMs. Additionally, passive anti-PC IgM Abs administered intratracheally with HDMs decreased allergen uptake by epithelial cells and APCs in the lungs of C57BL/6 mice but not CD36-/- or PAFR-/- mice. These results show that CD36 and PAFR are important mediators of HDM allergy development and that inhibiting HDM engagement with PC receptors in the lung protects against allergic airway disease.

Immunological Outcomes of Antibody Binding to Glycans Shared between Microorganisms and Mammals.

Glycans constitute basic cellular components of living organisms across biological kingdoms, and glycan-binding Abs participate in many cellular interactions during immune defense against pathogenic organisms. Glycan epitopes are expressed as carbohydrate-only entities or as oligomers or polymers on proteins and lipids. Such epitopes on glycoproteins may be formed by posttranslational modifications or neoepitopes resulting from metabolic-catabolic processes and can be altered during inflammation. Pathogenic organisms can display host-like glycans to evade the host immune response. However, Abs to glycans, shared between microorganisms and the host, exist naturally. These Abs are able to not only protect against infectious disease, but also are involved in host housekeeping functions and can suppress allergic disease. Despite the reactivity of these Abs to glycans shared between microorganisms and host, diverse tolerance-inducing mechanisms permit the B cell precursors of these Ab-secreting cells to exist within the normal B cell repertoire.

Pulmonary α-1,3-Glucan-Specific IgA-Secreting B Cells Suppress the Development of Cockroach Allergy.

There is a higher incidence of allergic conditions among children living in industrialized countries than those in developing regions. One explanation for this is reduced neonatal exposure to microbes and the consequent lack of immune stimulation. Sensitivity to cockroach allergen is highly correlated with the development of severe asthma. In this study, we determined that an Ab to microbial α-1,3-glucan binds an Enterobacter species and cockroach allergen. Neonatal, but not adult, mice immunized with this α-1,3-glucan-bearing Enterobacter (MK7) are protected against cockroach allergy. Following exposure to cockroach allergen, α-1,3-glucan-specific IgA-secreting cells are present in the lungs of mice immunized with MK7 as neonates but not in the lungs of those immunized as adults. Mice that are unable to generate anti-α-1,3-glucan IgA Abs were immunized with MK7 as neonates and were no longer protected against cockroach allergy. Thus, neonatal, but not adult, exposure to α-1,3-glucan results in suppressed development of cockroach allergy via pulmonary α-1,3-glucan-specific IgA-secreting cells.

Neonatal exposure to pneumococcal phosphorylcholine modulates the development of house dust mite allergy during adult life.

Currently, ∼20% of the global population suffers from an allergic disorder. Allergies and asthma occur at higher rates in developed and industrialized countries. It is clear that many human atopic diseases are initiated neonatally and herald more severe IgE-mediated disorders, including allergic asthma, which is driven by the priming of Th2 effector T cells. The hygiene hypothesis attempts to link the increased excessively sanitary conditions early in life to a default Th2 response and increasing allergic phenomena. Despite the substantial involvement of IgE Abs in such conditions, little attention has been paid to the effects of early microbial exposure on the B cell repertoire prior to the initiation of these diseases. In this study, we use Ab-binding assays to demonstrate that Streptococcus pneumoniae and house dust mite (HDM) bear similar phosphorylcholine (PC) epitopes. Neonatal C57BL/6 mice immunized with a PC-bearing pneumococcal vaccine expressed increased frequencies of PC-specific B cells in the lungs following sensitizing exposure to HDM as adults. Anti-PC IgM Abs in the lung decreased the interaction of HDM with pulmonary APCs and were affiliated with lowered allergy-associated cell infiltration into the lung, IgE production, development of airway hyperresponsiveness, and Th2 T cell priming. Thus, exposure of neonatal mice to PC-bearing pneumococci significantly reduced the development of HDM-induced allergic disease during adult life. Our findings demonstrate that B cells generated against conserved epitopes expressed by bacteria, encountered early in life, are also protective against the development of allergic disease during adult life.

Natural antibody repertoires: development and functional role in inhibiting allergic airway disease.

In this review we discuss the effects of microbial exposure on the B cell repertoire. Neonatal exposure to conserved bacterial carbohydrates and phospholipids permanently reprograms the natural antibody repertoire directed toward these antigens by clonal expansion, alterations in clonal dominance, and increased serum antibody levels. These epitopes are present not only in bacterial cell walls, but also in common environmental allergens. Neonatal immunization with bacterial polysaccharide vaccines results in attenuated allergic airway responses to fungi-, house dust mite-, and cockroach-associated allergens in mouse models. The similarities between mouse and human natural antibody repertoires suggest that reduced microbial exposure in children may have the opposite effect, providing a potential mechanistic explanation for the hygiene hypothesis. We propose that understanding the effects of childhood infections on the natural antibody repertoire and the mechanisms of antibody-mediated immunoregulation observed in allergy models will lead to the development of prevention/interventional strategies for treatment of allergic asthma.