Snai2 and Snai3 transcriptionally regulate cellular fitness and functionality of T cell lineages through distinct gene programs.

T lymphocytes are essential contributors to the adaptive immune system and consist of multiple lineages that serve various effector and regulatory roles. As such, precise control of gene expression is essential to the proper development and function of these cells. Previously, we identified Snai2 and Snai3 as being essential regulators of immune tolerance partly due to the impaired function of CD4(+) regulatory T cells in Snai2/3 conditional double knockout mice.

Antagonistic Interplay between MicroRNA-155 and IL-10 during Lyme Carditis and Arthritis.

MicroRNA-155 has been shown to play a role in immune activation and inflammation, and is suppressed by IL-10, an important anti-inflammatory cytokine. The established involvement of IL-10 in the murine model of Borrelia burgdorferi-induced Lyme arthritis and carditis allowed us to assess the interplay between IL-10 and miR-155 in vivo. As reported previously, Mir155 was highly upregulated in joints from infected severely arthritic B6 Il10-/- mice, but not in mildly arthritic B6 mice.

β-Glucuronidase, a Regulator of Lyme Arthritis Severity, Modulates Lysosomal Trafficking and MMP-9 Secretion in Response to Inflammatory Stimuli.

The lysosomal enzyme β-glucuronidase (Gusb) is a key regulator of Lyme-associated and K/B×N-induced arthritis severity. The luminal enzymes present in lysosomes provide essential catabolic functions for the homeostatic degradation of a variety of macromolecules. In addition to this essential catabolic function, lysosomes play important roles in the inflammatory response following infection.

Age-related onset of obesity corresponds with metabolic dysregulation and altered microglia morphology in mice deficient for Ifitm proteins.

The IfitmDel mouse lacks all five of the Ifitm genes via LoxP deletion. This animal breeds normally with no obvious defect in development. The IfitmDel animals exhibit a steady and significantly enhanced weight gain relative to wild-type controls beginning about three months of age and under normal feeding conditions.

Fatal autoimmunity results from the conditional deletion of Snai2 and Snai3.

Transcriptional regulation of gene expression is a key component of orchestrating proper immune cell development and function. One strategy for maintaining these transcriptional programs has been the evolution of transcription factor families with members possessing overlapping functions. Using the germ line deletion of Snai2 combined with the hematopoietic specific deletion of Snai3, we report that these factors function redundantly to preserve the development of B and T cells.

Borrelia burgdorferi arthritis-associated locus Bbaa1 regulates Lyme arthritis and K/B×N serum transfer arthritis through intrinsic control of type I IFN production.

Localized upregulation of type I IFN was previously implicated in development of Borrelia burgdorferi-induced arthritis in C3H mice, and was remarkable due to its absence in the mildly arthritic C57BL/6 (B6) mice. Independently, forward genetics analysis identified a quantitative trait locus on Chr4, termed B. burgdorferi-associated locus 1 (Bbaa1), that regulates Lyme arthritis severity and includes the 15 type I IFN genes.

Zfp318 regulates IgD expression by abrogating transcription termination within the Ighm/Ighd locus.

The protein Zfp318 is expressed during the transition of naive B cells from an immature to mature state. To evaluate its role in mature B cell functions, a conditional gene deficiency in Zfp318 was created and deleted in bone marrow lineages via Vav-Cre. B cell development was minimally altered in the absence of the protein, although transitional 2 (T2) B cell populations were depressed in the absence of Zfp318.

MicroRNA-146a provides feedback regulation of lyme arthritis but not carditis during infection with Borrelia burgdorferi.

MicroRNAs have been shown to be important regulators of inflammatory and immune responses and are implicated in several immune disorders including systemic lupus erythematosus and rheumatoid arthritis, but their role in Lyme borreliosis remains unknown. We performed a microarray screen for expression of miRNAs in joint tissue from three mouse strains infected with Borrelia burgdorferi. This screen identified upregulation of miR-146a, a key negative regulator of NF-κB signaling, in all three strains, suggesting it plays an important role in the in vivo response to B.

Snail transcription factors in hematopoietic cell development: a model of functional redundancy.

Coordinated gene expression is crucial in facilitating proper lymphoid cell development and function. The precise patterns of gene expression during B- and T-cell development are regulated through a complex interplay between a multitude of transcriptional regulators, both activators and repressors. We have recently identified the Snail family of transcription factors as playing significant and overlapping roles in lymphoid cell development, in that deletion of both SNAI2 and SNAI3 was required to fully impact the generation of mature T and B cells.

Murine complement receptor 1 is required for germinal center B cell maintenance but not initiation.

Germinal centers are the anatomic sites for the generation of high affinity immunoglobulin expressing plasma cells and memory B cells. The germinal center B cells that are precursors of these cells circulate between the light zone B cell population that interact with antigen laden follicular dendritic cells (FDC) and the proliferative dark zone B cell population. Antigen retention by follicular dendritic cells is dependent on Fc receptors and complement receptors, and complement receptor 1 (Cr1) is the predominant complement receptor expressed by FDC.

Lysosomal β-glucuronidase regulates Lyme and rheumatoid arthritis severity.

Lyme disease, caused by the spirochete Borrelia burgdorferi, is the most prevalent arthropod-borne illness in the United States and remains a clinical and social challenge. The spectrum of disease severity among infected patients suggests that host genetics contribute to pathogenic outcomes, particularly in patients who develop arthritis. Using a forward genetics approach, we identified the lysosomal enzyme β-glucuronidase (GUSB), a member of a large family of coregulated lysosomal enzymes, as a key regulator of Lyme-associated arthritis severity.

Quantification of complement receptor 2 calcium signaling enhancement using flow cytometry.

The complement receptor 2 (Cr2) gene is exclusively expressed in B cells and follicular dendritic cells (FDC) in mice and in humans. CR2 is a receptor for the complement component 3 (C3) cleavage fragments C3d(g) and iC3b. On B cells CR2 acts as the B cell co-receptor in which ligand binding of CR2 effectively lowers the threshold for B cell activation.

Detection of complement receptors 1 and 2 on mouse splenic B cells using flow cytometry.

The complement receptor 2 (Cr2) gene is exclusively expressed in B cells and follicular dendritic cells (FDC) in mice and in humans. However, mice also express an alternative splice variant, CR1, of the Cr2 gene. CR2 and CR1 are receptors for the complement component 3 (C3) cleavage fragments C3d(g) and iC3b.

Generation of a novel Cr2 gene allele by homologous recombination that abrogates production of Cr2 but is sufficient for expression of Cr1.

The enhancing effects of the complement system for humoral immunity have primarily focused upon the recognition of complement-bound foreign antigens by a co-receptor complex of the antigen-specific B cell receptor (BCR) and complement receptor 2 (Cr2). In vivo experiments using Cr2 gene deficient mice (which lack the expression of both the Cr1 and Cr2 proteins) do demonstrate depressed humoral responses to immunization but cannot be used to define specific contributions of the singular Cr1 or Cr2 proteins on B cell functions. To study the effect of a Cr2 deficiency in a Cr1 sufficient environment we created a mouse line in which the alternative splice site required for the expression of the Cr2 isoform was removed.

Deletion of Snai2 and Snai3 results in impaired physical development compounded by lymphocyte deficiency.

The Snail family of transcriptional regulators consists of three highly conserved members. These proteins regulate (repress) transcription via the recruitment of histone deacetylases to target gene promoters that possess the appropriate E-box binding sequences. Murine Snai1 is required for mouse development while Snai2 deficient animals survive with some anomalies.

Optimal germinal center B cell activation and T-dependent antibody responses require expression of the mouse complement receptor Cr1.

Follicular dendritic cells (FDCs) and complement receptor (Cr)1 and complement receptor (Cr)2 are important for the generation of humoral immunity. Cr1/2 expression on B cells and FDCs was shown to provide a secondary signal for B cell activation, to facilitate transport of Ag in immune follicles, and to enhance retention of immune complexes by FDCs. We show in this study that murine B cells predominantly express the Cr2 product from the Cr2 gene, whereas FDCs almost exclusively express the Cr1 isoform generated from the Cr2 gene.

Bone marrow-induced Mef2c deficiency delays B-cell development and alters the expression of key B-cell regulatory proteins.

The Mef2 family transcriptional regulator Mef2c (myocyte enhancer factor 2c) is highly expressed in maturing bone marrow and peripheral mature B-cells. To evaluate the role of this transcription factor in B-cell development, we generated a B-cell-specific conditional deletion of Mef2c using the Mb-1-Cre transgene that is expressed during the early stages of immunoglobulin rearrangement. Young mice possessing this defect demonstrated a significant impairment in B-cell numbers in bone marrow and spleen.

Overexpression of Snai3 suppresses lymphoid- and enhances myeloid-cell differentiation.

The altered expression of transcription factors in hematopoietic stem cells and their subsequent lineages can alter the development of lymphoid and myeloid lineages. The role of the transcriptional repressor Snai3 protein in the derivation of cells of the hemato-poietic system was investigated. Snai3 is expressed in terminal T-cell and myeloid lineages, therefore, we chose to determine if expressing Snai3 in the early stages of hematopoietic development would influence cell-lineage determination.

Interferon-inducible transmembrane proteins of the innate immune response act as membrane organizers by influencing clathrin and v-ATPase localization and function.

The innate response interferon-inducible transmembrane (Ifitm) proteins have been characterized as influencing proliferation, signaling complexes and restricting virus infections. Treatment of cells lacking these proteins (IfitmDel) with IFN-β resulted in the loss of clathrin from membrane compartments and the inhibition of clathrin-mediated phagocytosis, suggesting a molecular interaction between clathrin and Ifitm proteins. The pH of endosomes of IfitmDel cells, with or without IFN activation, was neutralized, suggesting the function of the vacular ATPase proton pumps in such cells was compromised.

High-throughput genotyping of advanced congenic lines by high resolution melting analysis for identification of Bbaa2, a QTL controlling Lyme arthritis.

Congenic mapping is a powerful strategy to identify genomic loci regulating quantitative traits. Generating congenic lines is an iterative process of refinement that is both time and resource intensive. Here we report an alternative to traditional microsatellite marker analysis or costly high-density oligonucleotide single nucleotide polymorphism (SNP) arrays for congenic genotyping.

Localized production of IL-10 suppresses early inflammatory cell infiltration and subsequent development of IFN-γ-mediated Lyme arthritis.

IL-10 is a nonredundant inflammatory modulator that suppresses arthritis development in Borrelia burgdorferi-infected mice. Infected C57BL/6 (B6) IL-10(-/-) mice were previously found to have a prolonged IFN-inducible response in joint tissue. Infection of B6 IL-10 reporter mice identified macrophages and CD4(+) T cells as the primary sources of IL-10 in the infected joint tissue, suggesting that early local production of IL-10 dampened the proarthritic IFN response.

The Lyme disease spirochete Borrelia burgdorferi utilizes multiple ligands, including RNA, for interferon regulatory factor 3-dependent induction of type I interferon-responsive genes.

We recently discovered a critical role for type I interferon (IFN) in the development of murine Lyme arthritis. Borrelia burgdorferi-mediated induction of IFN-responsive genes by bone marrow-derived macrophages (BMDMs) was dependent upon a functional type I IFN receptor but independent of Toll-like receptor 2 (TLR2), TLR4, TLR9, and the adapter molecule MyD88. We now demonstrate that induction of the IFN transcriptional profile in B.