DOCK2 Sets the Threshold for Entry into the Virtual Memory CD8+ T Cell Compartment by Negatively Regulating Tonic TCR Triggering.

The control of cytoskeletal dynamics by dedicator of cytokinesis 2 (DOCK2), a hematopoietic cell-specific actin effector protein, has been implicated in TCR signaling and T cell migration. Biallelic mutations in Dock2 have been identified in patients with a recessive form of combined immunodeficiency with defects in T, B, and NK cell activation. Surprisingly, we show in this study that certain immune functions of CD8+ T cells are enhanced in the absence of DOCK2. Dock2-deficient mice have a pronounced expansion of their memory T cell compartment. Bone marrow chimera and adoptive transfer studies indicate that these memory T cells develop in a cell-intrinsic manner following thymic egress. Transcriptional profiling, TCR repertoire analyses, and cell surface marker expression indicate that Dock2-deficient naive CD8+ T cells directly convert into virtual memory cells without clonal effector T cell expansion. This direct conversion to memory is associated with a selective increase in TCR sensitivity to self-peptide MHC in vivo and an enhanced response to weak agonist peptides ex vivo. In contrast, the response to strong agonist peptides remains unaltered in Dock2-deficient T cells. Collectively, these findings suggest that the regulation of the actin dynamics by DOCK2 enhances the threshold for entry into the virtual memory compartment by negatively regulating tonic TCR triggering in response to weak agonists.

Autoimmunity in women: an eXamination of eXisting models.

Women comprise over 80% of the affected individuals for many autoimmune conditions. Although sex-specific differences in sex hormones are thought to contribute to the female predisposition to autoimmunity, emerging evidence also suggests an intriguing role of both physiological and dysregulated X-chromosome inactivation. Furthermore, recent studies have demonstrated that many immune genes encoded on the X chromosome are expressed biallelically, and the contribution of these sex-specific differences in immune gene dosage to autoimmunity remains to be fully explored. This review highlights recent developments in this field and discusses questions that remain unanswered.

Identification of Pseudomonas aeruginosa phenazines that kill Caenorhabditis elegans.

Pathogenic microbes employ a variety of methods to overcome host defenses, including the production and dispersal of molecules that are toxic to their hosts. Pseudomonas aeruginosa, a Gram-negative bacterium, is a pathogen of a diverse variety of hosts including mammals and the nematode Caenorhabditis elegans. In this study, we identify three small molecules in the phenazine class that are produced by P. aeruginosa strain PA14 that are toxic to C. elegans. We demonstrate that 1-hydroxyphenazine, phenazine-1-carboxylic acid, and pyocyanin are capable of killing nematodes in a matter of hours. 1-hydroxyphenazine is toxic over a wide pH range, whereas the toxicities of phenazine-1-carboxylic acid and pyocyanin are pH-dependent at non-overlapping pH ranges. We found that acidification of the growth medium by PA14 activates the toxicity of phenazine-1-carboxylic acid, which is the primary toxic agent towards C. elegans in our assay. Pyocyanin is not toxic under acidic conditions and 1-hydroxyphenazine is produced at concentrations too low to kill C. elegans. These results suggest a role for phenazine-1-carboxylic acid in mammalian pathogenesis because PA14 mutants deficient in phenazine production have been shown to be defective in pathogenesis in mice. More generally, these data demonstrate how diversity within a class of metabolites could affect bacterial toxicity in different environmental niches.

B1a and B2 cells are characterized by distinct CpG modification states at DNMT3A-maintained enhancers.

The B1 and B2 lineages of B cells contribute to protection from pathogens in distinct ways. The role of the DNA CpG methylome in specifying these two B-cell fates is still unclear. Here we profile the CpG modifications and transcriptomes of peritoneal B1a and follicular B2 cells, as well as their respective proB cell precursors in the fetal liver and adult bone marrow from wild-type and CD19-Cre Dnmt3a floxed mice lacking DNMT3A in the B lineage. We show that an underlying foundational CpG methylome is stably established during B lineage commitment and is overlaid with a DNMT3A-maintained dynamic methylome that is sculpted in distinct ways in B1a and B2 cells. This dynamic DNMT3A-maintained methylome is composed of novel enhancers that are closely linked to lineage-specific genes. While DNMT3A maintains the methylation state of these enhancers in both B1a and B2 cells, the dynamic methylome undergoes a prominent programmed demethylation event during B1a but not B2 cell development. We propose that the methylation pattern of DNMT3A-maintained enhancers is determined by the coincident recruitment of DNMT3A and TET enzymes, which regulate the developmental expression of B1a and B2 lineage-specific genes.

Both live and dead Enterococci activate Caenorhabditis elegans host defense via immune and stress pathways.

The innate immune response of the nematode Caenorhabditis elegans has been extensively studied and a variety of Toll-independent immune response pathways have been identified. Surprisingly little, however, is known about how pathogens activate the C. elegans immune response. Enterococcus faecalis and Enterococcus faecium are closely related enterococcal species that exhibit significantly different levels of virulence in C. elegans infection models. Previous work has shown that activation of the C. elegans immune response by Pseudomonas aeruginosa involves P. aeruginosa-mediated host damage. Through ultrastructural imaging, we report that infection with either E. faecalis or E. faecium causes the worm intestine to become distended with proliferating bacteria in the absence of extensive morphological changes and apparent physical damage. Genetic analysis, whole-genome transcriptional profiling, and multiplexed gene expression analysis demonstrate that both enterococcal species, whether live or dead, induce a rapid and similar transcriptional defense response dependent upon previously described immune signaling pathways. The host response to E. faecium shows a stricter dependence upon stress response signaling pathways than the response to E. faecalis. Unexpectedly, we find that E. faecium is a C. elegans pathogen and that an active wild-type host defense response is required to keep an E. faecium infection at bay. These results provide new insights into the mechanisms underlying the C. elegans immune response to pathogen infection.

The expansion in lymphoid organs of IL-4+ BATF+ T follicular helper cells is linked to IgG4 class switching in vivo.

Distinct T follicular helper (TFH) subsets that influence specific class-switching events are assumed to exist, but the accumulation of isotype-specific TFH subsets in secondary lymphoid organs (SLOs) and tertiary lymphoid organs has not been hitherto demonstrated. IL-4-expressing TFH cells are surprisingly sparse in human SLOs. In contrast, in IgG4-related disease (IgG4-RD), a disorder characterized by polarized Ig class switching, most TFH cells in tertiary and SLOs make IL-4. Human IL-4+ TFH cells do not express GATA-3 but express nuclear BATF, and the transcriptomes of IL-4-secreting TFH cells differ from both PD1hi TFH cells that do not secrete IL-4 and IL-4-secreting non-TFH cells. Unlike IgG4-RD, IL-4+ TFH cells are rarely found in tertiary lymphoid organs in Sjögren's syndrome, a disorder in which IgG4 is not elevated. The proportion of CD4+IL-4+BATF+ T cells and CD4+IL-4+CXCR5+ T cells in IgG4-RD tissues correlates tightly with tissue IgG4 plasma cell numbers and plasma IgG4 levels in patients but not with the total plasma levels of other isotypes. These data describe a disease-related TFH subpopulation in human tertiary lymphoid organs and SLOs that is linked to IgG4 class switching.

Enterococcus infection biology: lessons from invertebrate host models.

The enterococci are commensals of the gastrointestinal tract of many metazoans, from insects to humans. While they normally do not cause disease in the intestine, they can become pathogenic when they infect sites outside of the gut. Recently, the enterococci have become important nosocomial pathogens, with the majority of human enterococcal infections caused by two species, Enterococcus faecalis and Enterococcus faecium. Studies using invertebrate infection models have revealed insights into the biology of enterococcal infections, as well as general principles underlying host innate immune defense. This review highlights recent findings on Enterococcus infection biology from two invertebrate infection models, the greater wax moth Galleria mellonella and the free-living bacteriovorous nematode Caenorhabditis elegans.

The role of Candida albicans SPT20 in filamentation, biofilm formation and pathogenesis.

Candida albicans is a ubiquitous fungus, which can cause very serious and sometimes life-threatening infections in susceptible patients. We used Caenorhabditis elegans as a model host to screen a library of C. albicans mutants for decreased virulence and identified SPT20 as important for virulence. The transcription co-activator SPT20 was identified originally as a suppressor of Ty and solo δ insertion mutations, which can cause transcription defects in Saccharomyces cerevisiae. It is resistant to the toxicity caused by overexpression of GAL4-VP16. We constructed a C. albicans spt20Δ/Δ mutant and found the spt20Δ/Δ strain was significantly less virulent than the wild-type strain SC5314 in C. elegans (p < 0.0001), Galleria mellonella (p < 0.01) and mice (p < 0.001). Morphologically, spt20Δ/Δ mutant cells demonstrated a "snow-flake" shape and clustered together; prolonged culture times resulted in increased size of the cluster. The clustered morphology was associated with defects in nuclei distribution, as the nuclei were not observed in many cellular compartments. In addition, the C. albicans spt20Δ/Δ mutant resulted in defects in hyphae and biofilm formation (compared to the wild-type strain, p < 0.05), and sensitivity to cell wall and osmotic stressors, and to antifungal agents. Thus our study demonstrated a role of C. albicans SPT20 in overall morphology and distribution of nuclear material, which may cause the defects in filamentation and biofilm formation directly when this gene is deleted.