Antigen stimulation (signal 1) triggers B cell proliferation, and primes B cells to recruit, engage, and respond to T cell help (signal 2). Failure to receive signal 2 within a defined time window results in B cell apoptosis, yet the mechanisms that enforce dependence upon co-stimulation are incompletely understood. Nr4a1-3 encode a small family of orphan nuclear receptors that are rapidly induced by B cell antigen receptor (BCR) stimulation. Here we showed that Nr4a1 and Nr4a3 play partially redundant roles to restrain B cell responses to antigen in the absence of co-stimulation, and do so in part by repressing expression of BATF and consequently MYC. The NR4A family also restrains B cell access to T cell help by repressing expression of the T cell chemokines CCL3 and CCL4, as well as CD86 and ICAM1. Such NR4A-mediated regulation plays a role specifically under conditions of competition for limiting T cell help.

Interferon-induced proteins, including the largely uncharacterized interferon-induced tetratricopeptide repeat (IFIT) protein family, provide defense against pathogens. Differing from expectation for tetratricopeptide repeat (TPR) proteins and from human IFITs 1, 2, and 3, we show that human IFIT5 recognizes cellular RNA instead of protein partners. In vivo and in vitro, IFIT5 bound to endogenous 5’-phosphate-capped RNAs including transfer RNAs (tRNAs). The crystal structure of IFIT5 revealed a convoluted intramolecular packing of eight TPRs as a fold that we name the TPR eddy. Additional, non-TPR structural elements contribute to an RNA binding cleft. Instead of general cytoplasmic distribution, IFIT5 concentrated in actin-rich protrusions from the apical cell surface co-localized with the RNA-binding retinoic acid-inducible gene-I (RIG-I). These findings establish compartmentalized cellular RNA binding activity as a mechanism for IFIT5 function and reveal the TPR eddy as a scaffold for RNA recognition.

Nur77 (Nr4a1) belongs to a small family of orphan nuclear receptors that are rapidly induced by BCR stimulation, yet little is known about its function in B cells. We have previously characterized a reporter of Nr4a1 transcription, Nur77-eGFP, in which GFP expression faithfully detects Ag encounter by B cells in vitro and in vivo. In this study, we report that Nur77 expression correlates with the degree of self-reactivity, counterselection, and anergy among individual B cell clones from two distinct BCR transgenic mouse models but is dispensable for all of these tolerance mechanisms. However, we identify a role for Nur77 in restraining survival of self-reactive B cells in the periphery under conditions of competition for a limited supply of the survival factor BAFF. We find that Nur77 deficiency results in the progressive accumulation of self-reactive B cells in the mature repertoire with age and is sufficient to break B cell tolerance in V H 3H9 H chain transgenic mice. We thus propose that Nur77 is upregulated in self-reactive B cells in response to chronic Ag stimulation and selectively restricts the survival of these cells, gradually pruning self-reactivity from the mature repertoire to impose a novel layer of peripheral B cell tolerance.

Efficient mechanisms of central tolerance, including receptor editing and deletion, prevent highly self-reactive B cell receptors (BCRs) from populating the periphery.Despite this, modest self-reactivity persists in (and may even be actively selected into) the mature B cell repertoire. In this review, we discuss new insights into mechanisms of peripheral B cell tolerance that restrain mature B cells from mounting inappropriate responses to endogenous antigens, and place recent work into historical context. In particular, we discuss new findings that have arisen from application of a novel in vivo reporter of BCR signaling, Nur77-eGFP, expression of which scales with the degree of self-reactivity in both monoclonal and polyclonal B cell repertoires. We discuss new and historical evidence that self-reactivity is not just tolerated, but actively selected into the peripheral repertoire. We review recent progress in understanding how dual expression of the IgM and IgD BCR isotypes on mature naive follicular B cells tunes responsiveness to endogenous antigen recognition, and discuss how this may be integrated with other features of clonal anergy. Finally, we discuss how expression of Nur77 itself couples chronic antigen stimulation with B cell tolerance. K E Y W O R D Sanergy, B cell, IgD, IgM, Nur77/Nr4a1, tolerance Previous work from our laboratory exploiting an in vivo reporter of antigen recognition, Nur77-eGFP BAC Tg, addresses this important point ( Figure 1A). 15 GFP expression in this reporter line is under the control of the regulatory region of Nr4a1/ Nur77. Nr4a1-3 encode a small family of orphan nuclear hormone receptors which were originally cloned as signal-dependent primary response genes (also known as immediate-early genes), and are highly upregulated by a range of mitogens, including antigen receptor stimulation. [16][17][18] Consequently, antigen stimulation rapidly triggers reporter expression in B and T cells in vitro ( Figure 1B), and GFP is also induced by infection or immunization in antigen-specific lymphocytes in

Summary CD4+ T cells differentiate into T helper cell subsets in feed-forward manners with synergistic signals from the T cell receptor (TCR), cytokines, and lineage-specific transcription factors. Naïve CD4+ T cells avoid spontaneous engagement of feed-forward mechanisms but retain a prepared state. T cells lacking the adapter molecule LAT demonstrate impaired TCR-induced signals yet cause a spontaneous lymphoproliferative T helper 2 (TH2) cell syndrome in mice. Thus, LAT constitutes an unexplained maintenance cue. Here we demonstrate that tonic signals through LAT constitutively export the repressor HDAC7 from the nucleus of CD4+ T cells. Without such tonic signals, HDAC7 target genes Nur77 and Irf4 are repressed. We reveal that Nur77 suppresses CD4+ T cell proliferation and uncover a suppressive role for Irf4 in TH2 polarization; halving Irf4 gene-dosage leads to increases in GATA3+ and IL4+ cells. Our studies reveal that naïve CD4+ T cells are dynamically tuned by tonic LAT-HDAC7 signals.

Background: Peptidoglycan hydrolases help elongate, divide, and recycle bacterial cell walls. Results: The structure and activity of Rv3717, a peptidoglycan hydrolase of Mycobacterium tuberculosis, are described. Conclusion: Our data suggest a peptidoglycan-recycling role for Rv3717. Significance: Functional specialization of homologous peptidoglycan hydrolases can be achieved by altering their substrate range via structural variation.

It has long been appreciated that highly autoreactive BCRs are actively removed from the developing B cell repertoire by Agdependent receptor editing and deletion. However, there is persistent debate about whether mild autoreactivity is simply tolerated or positively selected into the mature B cell repertoire as well as at what stage, to what extent, under what conditions, and into which compartments this occurs. In this study, we describe two minor, trackable populations of B cells in B1-8i Ig transgenic mice that express the VH186.2 H chain and recognize a common foreign Ag (the hapten 4-hydroxy-3-nitrophenylacetyl) but differ in L chain expression. We use the Nur77-eGFP reporter of BCR signaling to define their reactivity toward endogenous Ags. The less autoreactive of these two populations is strongly counterselected during the development of mature B1a, follicular, and marginal zone B cells. By genetically manipulating the strength of BCR signal transduction via the titration of surface CD45 expression, we demonstrate that this B cell population is not negatively selected but instead displays characteristics of impaired positive selection. We demonstrate that mild self-reactivity improves the developmental fitness of B cell clones in the context of a diverse population of B cells, and positive selection by endogenous Ags shapes the mature B cell repertoire.

B-1a cells are a unique population of innate-like B cells with a highly restricted and self-reactive BCR repertoire. Preimmune “natural” IgM produced by B-1a–derived plasma cells is essential for homeostatic clearance of cellular debris and forms a primary layer of protection against infection. In this study, we take advantage of a fluorescent reporter of BCR signaling to show that expression of the orphan nuclear hormone receptor Nur77 is upregulated under steady-state conditions in self-reactive B-1a cells in response to chronic Ag stimulation. Nur77-deficient mice exhibit elevated natural serum IgM (but not IgG) and marked expansion of IgM plasma cells of B-1a origin. Moreover, we show that Nur77 restrains the turnover of B-1a cells and the accumulation of immature IgM plasma cells. Thus, we identify a new critical negative-regulatory pathway that serves to maintain B-1a cells in a quiescent state in the face of chronic endogenous Ag stimulation.