Jinx, an MCMV susceptibility phenotype caused by disruption of Unc13d: a mouse model of type 3 familial hemophagocytic lymphohistiocytosis

Mouse cytomegalovirus (MCMV) susceptibility often results from defects of natural killer (NK) cell function. Here we describe Jinx, an N-ethyl-N-nitrosourea–induced MCMV susceptibility mutation that permits unchecked proliferation of the virus, causing death. In Jinx homozygotes, activated NK cells and cytotoxic T lymphocytes (CTLs) fail to degranulate, although they retain the ability to produce cytokines, and cytokine levels are markedly elevated in the blood of infected mutant mice. Jinx was mapped to mouse chromosome 11 on a total of 246 meioses and confined to a 4.60–million basepair critical region encompassing 122 annotated genes. The phenotype was ascribed to the creation of a novel donor splice site in Unc13d, the mouse orthologue of human MUNC13-4, in which mutations cause type 3 familial hemophagocytic lymphohistiocytosis (FHL3), a fatal disease marked by massive hepatosplenomegaly, anemia, and thrombocytopenia. Jinx mice do not spontaneously develop clinical features of hemophagocytic lymphohistiocytosis (HLH), but do so when infected with lymphocytic choriomeningitis virus, exhibiting hyperactivation of CTLs and antigen-presenting cells, and inadequate restriction of viral proliferation. In contrast, neither Listeria monocytogenes nor MCMV induces the syndrome. In mice, the HLH phenotype is conditional, which suggests the existence of a specific infectious trigger of FHL3 in humans

Commitment to the Regulatory T Cell Lineage Requires CARMA1 in the Thymus but Not in the Periphery

Regulatory T (Treg) cells expressing forkhead box P3 (Foxp3) arise during thymic selection among thymocytes with modestly self-reactive T cell receptors. In vitro studies suggest Foxp3 can also be induced among peripheral CD4+ T cells in a cytokine dependent manner. Treg cells of thymic or peripheral origin may serve different functions in vivo, but both populations are phenotypically indistinguishable in wild-type mice. Here we show that mice with a Carma1 point mutation lack thymic CD4+Foxp3+ Treg cells and demonstrate a cell-intrinsic requirement for CARMA1 in thymic Foxp3 induction. However, peripheral Carma1-deficient Treg cells could be generated and expanded in vitro in response to the cytokines transforming growth factor beta (TGFβ) and interleukin-2 (IL-2). In vivo, a small peripheral Treg pool existed that was enriched at mucosal sites and could expand systemically after infection with mouse cytomegalovirus (MCMV). Our data provide genetic evidence for two distinct mechanisms controlling regulatory T cell lineage commitment. Furthermore, we show that peripheral Treg cells are a dynamic population that may expand to limit immunopathology or promote chronic infection

Degron mediated BRM/SMARCA2 depletion uncovers novel combination partners for treatment of BRG1/SMARCA4-mutant cancers.

Recent studies have highlighted that cancer cells with a loss of the SWI/SNF complex catalytic subunit BRG1 are dependent on the remaining ATPase, BRM, making it an attractive target for cancer therapy. However, an understanding of the extent of target inhibition required to arrest cell growth, necessary to develop an appropriate therapeutic strategy, remains unknown. Here, we utilize tunable depletion of endogenous BRM using the SMASh degron, and interestingly observe that BRG1-mutant lung cancer cells require near complete depletion of BRM to robustly inhibit growth both in vitro and in vivo. Therefore, to identify pathways that synergize with partial BRM depletion and afford a deeper response, we performed a genome-wide CRISPR screen and discovered a combinatorial effect between BRM depletion and the knockout of various genes of the oxidative phosphorylation pathway and the anti-apoptotic gene MCL1. Together these studies provide an important framework to elucidate the requirements of BRM inhibition in the BRG1-mutant state with implications on the feasibility of targeting BRM alone, as well as reveal novel insights into pathways that can be exploited in combination toward deeper anti-tumor responses

A Toll-Like Receptor 2-Responsive Lipid Effector Pathway Protects Mammals against Skin Infections with Gram-Positive Bacteria

flake (flk), an N-ethyl-N-nitrosourea-induced recessive germ line mutation of C57BL/6 mice, impairs the clearance of skin infections by Streptococcus pyogenes and Staphylococcus aureus, gram-positive pathogens that elicit innate immune responses by activating Toll-like receptor 2 (TLR2) (K. Takeda and S. Akira, Cell. Microbiol. 5:143-153, 2003). Positional cloning and sequencing revealed that flk is a novel allele of the stearoyl coenzyme A desaturase 1 gene (Scd1). flake homozygotes show reduced sebum production and are unable to synthesize the monounsaturated fatty acids (MUFA) palmitoleate (C(16:1)) and oleate (C(18:1)), both of which are bactericidal against gram-positive (but not gram-negative) organisms in vitro. However, intradermal MUFA administration to S. aureus-infected mice partially rescues the flake phenotype, which indicates that an additional component of the sebum may be required to improve bacterial clearance. In normal mice, transcription of Scd1—a gene with numerous NF-κB elements in its promoter—is strongly and specifically induced by TLR2 signaling. Similarly, the SCD1 gene is induced by TLR2 signaling in a human sebocyte cell line. These observations reveal the existence of a regulated, lipid-based antimicrobial effector pathway in mammals and suggest new approaches to the treatment or prevention of infections with gram-positive bacteria

Inflammation and autoimmunity caused by a SHP1 mutation depend on IL-1, MyD88, and a microbial trigger

A recessive phenotype called spin (spontaneous inflammation) was induced by N-ethyl-N-nitrosourea (ENU) mutagenesis in C57BL/6J mice. Homozygotes display chronic inflammatory lesions affecting the feet, salivary glands and lungs, and antichromatin antibodies. They are immunocompetent and show enhanced resistance to infection by Listeria monocytogenes. TLR-induced TNF and IL-1 production are normal in macrophages derived from spin mice. The autoinflammatory phenotype of spin mice is fully suppressed by compound homozygosity for Myd88poc, Irak4otiose, and Il1r1-null mutations, but not Ticam1Lps2, Stat1m1Btlr, or Tnf-null mutations. Both autoimmune and autoinflammatory phenotypes are suppressed when spin homozygotes are derived into a germ-free environment. The spin phenotype was ascribed to a viable hypomorphic allele of Ptpn6, which encodes the tyrosine phosphatase SHP1, mutated in mice with the classical motheaten alleles me and me-v. Inflammation and autoimmunity caused by SHP1 deficiency are thus conditional. The SHP1-deficient phenotype is driven by microbes, which activate TLR signaling pathways to elicit IL-1 production. IL-1 signaling via MyD88 elicits inflammatory disease

Loss of T cell and B cell quiescence precedes the onset of microbial flora-dependent wasting disease and intestinal inflammation in Gimap5-deficient mice

Homeostatic control of the immune system involves mechanisms that ensure the self-tolerance, survival and quiescence of hematopoietic-derived cells. In this study, we demonstrate that the GTPase of immunity associated protein (Gimap)5 regulates these processes in lymphocytes and hematopoietic progenitor cells. As a consequence of a recessive N-ethyl-N-nitrosourea–induced germline mutation in the P-loop of Gimap5, lymphopenia, hepatic extramedullary hematopoiesis, weight loss, and intestinal inflammation occur in homozygous mutant mice. Irradiated fetal liver chimeric mice reconstituted with Gimap5-deficient cells lose weight and become lymphopenic, demonstrating a hematopoietic cell-intrinsic function for Gimap5. Although Gimap5-deficient CD4(+) T cells and B cells appear to undergo normal development, they fail to proliferate upon Ag-receptor stimulation although NF-κB, MAP kinase and Akt activation occur normally. In addition, in Gimap5-deficient mice, CD4(+) T cells adopt a CD44(high) CD62L(low) CD69(low) phenotype and show reduced IL-7rα expression, and T-dependent and T-independent B cell responses are abrogated. Thus, Gimap5-deficiency affects a noncanonical signaling pathway required for Ag-receptor–induced proliferation and lymphocyte quiescence. Antibiotic-treatment or the adoptive transfer of Rag-sufficient splenocytes ameliorates intestinal inflammation and weight loss, suggesting that immune responses triggered by microbial flora causes the morbidity in Gimap5-deficient mice. These data establish Gimap5 as a key regulator of hematopoietic integrity and lymphocyte homeostasis

Loss of T cell and B cell quiescence precedes the onset of microbial flora-dependent wasting disease and intestinal inflammation in Gimap5-deficient mice

Homeostatic control of the immune system involves mechanisms that ensure the self-tolerance, survival and quiescence of hematopoietic-derived cells. In this study, we demonstrate that the GTPase of immunity associated protein (Gimap)5 regulates these processes in lymphocytes and hematopoietic progenitor cells. As a consequence of a recessive N-ethyl-N-nitrosourea-induced germline mutation in the P-loop of Gimap5, lymphopenia, hepatic extramedullary hematopoiesis, weight loss, and intestinal inflammation occur in homozygous mutant mice. Irradiated fetal liver chimeric mice reconstituted with Gimap5-deficient cells lose weight and become lymphopenic, demonstrating a hematopoietic cell-intrinsic function for Gimap5. Although Gimap5-deficient CD4(+) T cells and B cells appear to undergo normal development, they fail to proliferate upon Ag-receptor stimulation although NF-kappaB, MAP kinase and Akt activation occur normally. In addition, in Gimap5-deficient mice, CD4(+) T cells adopt a CD44(high)CD62L(low)CD69(low) phenotype and show reduced IL-7ralpha expression, and T-dependent and T-independent B cell responses are abrogated. Thus, Gimap5-deficiency affects a noncanonical signaling pathway required for Ag-receptor-induced proliferation and lymphocyte quiescence. Antibiotic-treatment or the adoptive transfer of Rag-sufficient splenocytes ameliorates intestinal inflammation and weight loss, suggesting that immune responses triggered by microbial flora causes the morbidity in Gimap5-deficient mice. These data establish Gimap5 as a key regulator of hematopoietic integrity and lymphocyte homeostasis