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The expanding number of rare immunodeficiency syndromes offers an opportunity to understand key genes that support immune defense against infectious diseases. However, analysis of these in patients is complicated by their treatments and comorbid infections, requiring the use of mouse models for detailed investigations. We developed a mouse model of DOCK2 immunodeficiency and herein demonstrate that these mice have delayed clearance of herpes simplex virus type 1 (HSV-1) infections. We also uncovered a critical, cell-intrinsic role of DOCK2 in the priming of antiviral CD8+ T cells and in particular their initial expansion, despite apparently normal early activation of these cells. When this defect was overcome by priming in vitro, DOCK2-deficient CD8+ T cells were surprisingly protective against HSV-1 disease, albeit not as effectively as wild-type cells. These results shed light on a cellular deficiency that is likely to impact antiviral immunity in DOCK2-deficient patients. [ABSTRACT FROM AUTHOR]

Special AT‐binding protein 1 (SATB1) is a chromatin‐binding protein that has been shown to be a key regulator of T‐cell development and CD4+ T‐cell fate decisions and function. The underlying function for SATB1 in peripheral CD8+ T‐cell differentiation processes is largely unknown. To address this, we examined SATB1‐binding patterns in naïve and effector CD8+ T cells demonstrating that SATB1 binds to noncoding regulatory elements linked to T‐cell lineage–specific gene programs, particularly in naïve CD8+ T cells. We then assessed SATB1 function using N‐ethyl‐N‐nitrosourea‐mutant mice that exhibit a point mutation in the SATB1 DNA‐binding domain (termed Satb1m1Anu/m1Anu). Satb1m1Anu/m1Anu mice exhibit diminished SATB1‐binding, naïve, Satb1m1Anu/m1Anu CD8+ T cells exhibiting transcriptional and phenotypic characteristics reminiscent of effector T cells. Upon activation, the transcriptional signatures of Satb1m1Anu/m1Anu and wild‐type effector CD8+ T cells converged. While there were no overt differences, primary respiratory infection of Satb1m1Anu/m1Anu mice with influenza A virus (IAV) resulted in a decreased proportion and number of IAV‐specific CD8+ effector T cells recruited to the infected lung when compared with wild‐type mice. Together, these data suggest that SATB1 has a major role in an appropriate transcriptional state within naïve CD8+ T cells and ensures appropriate CD8+ T‐cell effector gene expression upon activation. [ABSTRACT FROM AUTHOR]

Clostridium species are a group of Gram-positive bacteria that cause diseases in humans, such as food poisoning, botulism, and tetanus. Here, we analyzed 10 different Clostridium species and identified that Clostridium septicum, a pathogen that causes sepsis and gas gangrene, activates the mammalian cytosolic inflammasome complex in mice and humans. Mechanistically, we demonstrate that α-toxin secreted by C. septicum binds to glycosylphosphatidylinositol (GPI)–anchored proteins on the host plasma membrane, oligomerizing and forming a membrane pore that is permissive to efflux of magnesium and potassium ions. Efflux of these cytosolic ions triggers the activation of the innate immune sensor NLRP3, inducing activation of caspase-1 and gasdermin D, secretion of the proinflammatory cytokines interleukin-1β and interleukin-18, pyroptosis, and plasma membrane rupture via ninjurin-1. Furthermore, α-toxin of C. septicum induces rapid inflammasome-mediated lethality in mice and pharmacological inhibition of the NLRP3 inflammasome using MCC950 prevents C. septicum–induced lethality. Overall, our results reveal that cytosolic innate sensing of α-toxin is central to the recognition of C. septicum infection and that therapeutic blockade of the inflammasome pathway may prevent sepsis and death caused by toxin-producing pathogens. Toxin-triggered tribulations: Lurking within the resident human intestinal microbiota are Clostridium bacteria with the capacity to cause rare but serious deep tissue infections like gas gangrene after hematogenous dissemination. Jing et al. analyzed the downstream cellular signaling pathways triggered when Clostridium septicum α-toxin bound to the plasma membrane surface, triggering pore formation followed by potassium and magnesium ion efflux. The α-toxin–induced ion fluxes activated NLRP3 inflammasomes, leading to gasdermin D–dependent pore formation and cell death by pyroptosis that could be blocked by the NLRP3-specific inhibitor MCC950. These findings indicate that pharmacological inhibition of the NLRP3 inflammasome may provide a new option for the treatment of life-threatening C. septicum infections. [ABSTRACT FROM AUTHOR]

IKZF1 (IKAROS) is essential for normal lymphopoiesis in both humans and mice. Previous Ikzf1 mouse models have demonstrated the dual role for IKZF1 in both B and T cell development and have indicated differential requirements of each zinc finger. Furthermore, mutations in IKZF1 are known to cause common variable immunodeficiency in patients characterized by a loss of B cells and reduced Ab production. Through N-ethyl-N-nitrosourea mutagenesis, we have discovered a novel Ikzf1 mutant mouse with a missense mutation (L132P) in zinc finger 1 (ZF1) located in the DNA binding domain. Unlike other previously reported murine Ikzf1 mutations, this L132P point mutation (Ikzf1L132P) conserves overall protein expression and has a B cell-specific phenotype with no effect on T cell development, indicating that ZF1 is not required for T cells. Mice have reduced Ab responses to immunization and show a progressive loss of serum Igs compared with wild-type littermates. IKZF1L132P overexpressed in NIH3T3 or HEK293T cells failed to localize to pericentromeric heterochromatin and bind target DNA sequences. Coexpression of wild-type and mutant IKZF1, however, allows for localization to pericentromeric heterochromatin and binding to DNA indicating a haploinsufficient mechanism of action for IKZF1L132P. Furthermore, Ikzf1+/L132P mice have late onset defective Ig production, similar to what is observed in common variable immunodeficiency patients. RNA sequencing revealed a total loss of Hsf1 expression in follicular B cells, suggesting a possible functional link for the humoral immune response defects observed in Ikzf1L132P/L132P mice. [ABSTRACT FROM AUTHOR]

DOCK8 deficiency in humans and mice leads to multiple defects in immune cell numbers and function. Patients with this immunodeficiency have a high morbidity and mortality, and are distinguished by chronic cutaneous viral infections, including those caused by herpes simplex virus (HSV). The underlying mechanism of the specific susceptibility to these chronic cutaneous viral infections is currently unknown, largely because the effect of DOCK8 deficiency has not been studied in suitable models. A better understanding of these mechanisms is required to underpin the development of more specific therapies. Here we show that DOCK8-deficient mice have poor control of primary cutaneous herpes simplex lesions and this is associated with increased virus loads. Furthermore, DOCK8-deficient mice showed a lack of CD4+ T-cell infiltration into HSV-infected skin. [ABSTRACT FROM AUTHOR]

Gene variants that disrupt TCR signaling can cause severe immune deficiency, yet less disruptive variants are sometimes associated with immune pathology. Null mutations of the gene encoding the scaffold protein Src homology 2 domain-containing leukocyte protein of 76 kDa (SLP-76), for example, cause an arrest of T cell positive selection, whereas a synthetic membrane-targeted allele allows limited positive selection but is associated with proinflammatory cytokine production and autoantibodies. Whether these and other enigmatic outcomes are due to a biochemical uncoupling of tolerogenic signaling, or simply a quantitative reduction of protein activity, remains to be determined. In this study we describe a splice variant of Lcp2 that reduced the amount of wild-type SLP-76 protein by ~90%, disrupting immunogenic and tolerogenic pathways to different degrees. Mutant mice produced excessive amounts of proinflammatory cytokines, autoantibodies, and IgE, revealing that simple quantitative reductions of SLP-76 were sufficient to trigger immune dysrégulation. This allele reveals a dose-sensitive threshold for SLP-76 in the balance of immunity and immune dysrégulation, a common disturbance of atypical clinical immune deficiencies. [ABSTRACT FROM AUTHOR]

Forward genetics screens with N-ethyl-N-nitrosourea (ENU) provide a powerful way to illuminate gene function and generate mouse models of human disease; however, the identification of causative mutations remains a limiting step. Current strategies depend on conventional mapping, so the propagation of affected mice requires non-lethal screens; accurate tracking of phenotypes through pedigrees is complex and uncertain; out-crossing can introduce unexpected modifiers; and Sanger sequencing of candidate genes is inefficient. Here we show how these problems can be efficiently overcome using whole-genome sequencing (WGS) to detect the ENU mutations and then identify regions that are identical by descent (IBD) in multiple affected mice. In this strategy, we use a modification of the Lander-Green algorithm to isolate causative recessive and dominant mutations, even at low coverage, on a pure strain background. Analysis of the IBD regions also allows us to calculate the ENU mutation rate (1.54 mutations per Mb) and to model future strategies for genetic screens in mice. The introduction of this approach will accelerate the discovery of causal variants, permit broader and more informative lethal screens to be used, reduce animal costs, and herald a new era for ENU mutagenesis. [ABSTRACT FROM AUTHOR]

Divergent hypotheses exist to explain how signaling by the B cell receptor (BCR) is initiated after antigen binding and how it is qualitatively altered in anergic B cells to selectively uncouple from nuclear factor κB and c-Jun N-terminal kinase pathways while continuing to activate extracellular signal-regulated kinase and calcium-nuclear factor of activated T cell pathways. Here we find that BCRs on anergic cells are endocytosed at a very enhanced rate upon binding antigen, resulting in a large steady-state pool of intracellularly sequestered receptors that appear to be continuously cycling between surface and intracellular compartments. This endocytic mechanism is exquisitely sensitive to the lowering of plasma membrane cholesterol by methyl-β-cyclodextrin, and, when blocked in this way, the sequestered BCRs return to the cell surface and RelA nuclear accumulation is stimulated. In contrast, when plasma membrane cholesterol is lowered and GM1 sphingo-lipid markers of membrane rafts are depleted in naive B cells, this does not diminish BCR signaling to calcium or RelA. These results provide a possible explanation for the signaling changes in clonal anergy and indicate that a chief function of membrane cholesterol in B cells is not to initiate BCR signaling, but instead to terminate a subset of signals by rapid receptor internalization. [ABSTRACT FROM AUTHOR]

The article discusses the finding of new resistance regulatory genes by the N-ethyl-N-itrosourea (ENU) mutagenesis. It aims to develop new scheme to come up this need by the use of DNA sequencing and computational techniques. Results shows ENU mutant pedigrees used for animals can be usable for experimental models and validation of disease in human.