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The primary immunoglobulin repertoire develops via opposing forces of expanding diversification balanced by contracting selection mechanisms. The resulting shape is essential for host health and immune fitness. While the molecular mechanisms of Ig diversification have largely been defined, selection forces shaping emerging Ig repertoires are poorly understood. During lifetime, human and mouse early B cell development occurs at distinct locations — beginning in fetal liver before transferring to bone marrow and spleen by the end of gestation. During an early life window of time, the murine gut lamina propria harbors developing immature B cells in proximity to intestinal contents such as commensal microbes and dietary antigens. Location and timing of early B cell development may thus endow neighboring antigens with primary repertoire-shaping capabilities.

Animals and many of their chronic microbial inhabitants form relationships of symbiotic mutualism, which occurs when coexisting life-forms derive mutual benefit from stable associations. While microorganisms receive a secure habitat and constant food source from vertebrate hosts, they are required for optimal immune system development and occupy niches otherwise abused by pathogens. Microbes have also been shown to provide vertebrate hosts with metabolic capabilities that enhance energy and nutrient uptake from the diet. The immune system plays a central role in the establishment and maintenance of host-microbe homeostasis, and B lineage cells play a key role in this regulation. Here, I reviewed the structure and function of the microbiota and the known mechanisms of how nonpathogenic microbes influence B cell biology and immunoglobulin repertoire development early in life. I also discuss what is known about how B lineage cells contribute to the process of shaping the composition of commensal/mutualistic microbe membership.

Immunoglobulin (Ig) E is the most tightly regulated of all Ig heavy chain (IgH) isotypes and plays a key role in atopic disease. The gene encoding for IgH in mature B cells consists of a variable region exon-assembled from component gene segments via V(D)J recombination during early B cell development-upstream of a set of IgH constant region CH exons. Upon activation by antigen in peripheral lymphoid organs, B cells can undergo IgH class switch recombination (CSR), a process in which the initially expressed IgH μ constant region exons (Cμ) are deleted and replaced by one of several sets of downstream CH exons (e.g., Cγ, Cε, and Cα). Activation of the IL-4 receptor on B cells, together with other signals, can lead to the replacement of Cμ with Cε resulting in CSR to IgE through a series of molecular events involving irreversible remodeling of the IgH locus. Here, we discuss the molecular mechanisms of CSR and the unique features surrounding the generation of IgE-producing B cells.

Immunoglobulin (Ig) E is the most tightly regulated of all Ig heavy chain (IgH) isotypes and plays a key role in atopic disease. The gene encoding for IgH in mature B cells consists of a variable region exon—assembled from component gene segments via V(D)J recombination during early B cell development—upstream of a set of IgH constant region CH exons. Upon activation by antigen in peripheral lymphoid organs, B cells can undergo IgH class switch recombination (CSR), a process in which the initially expressed IgH μ constant region exons (Cμ) are deleted and replaced by one of several sets of downstream CH exons (e.g., Cγ, Ce, and Cα). Activation of the IL-4 receptor on B cells, together with other signals, can lead to the replacement of Cμ with Ce resulting in CSR to IgE through a series of molecular events involving irreversible remodeling of the IgH locus. Here, we discuss the molecular mechanisms of CSR and the unique features surrounding the generation of IgE-producing B cells.

Animals and many of the microorganisms that inhabit them form relationships of symbiotic mutualism, which occurs when coexisting life forms derive mutual benefit from stable associations. On one hand, microorganisms receive a secure habitat and constant food source from vertebrate hosts. On the other hand, mutualistic microbes benefit their hosts by providing signals that lead to optimal host immune system development and by occupying niches otherwise abused by pathogens. Microbes have also been shown to provide vertebrate hosts with metabolic capabilities that enhance energy and nutrient uptake from the diet. The immune system plays a central role in the establishment and maintenance of host: microbe homeostasis, and B-lineage cells play a key role in this regulation. The first segment of this chapter describes the current knowledge regarding the structure and function of the commensal microbiota and the known mechanisms of how nonpathogenic microbes influence B cell development early in life. The rest of the chapter discusses what is known about how B-lineage cells contribute to shaping the composition of commensal microbe membership.

Animals and many of their chronic microbial inhabitants form relationships of symbiotic mutualism, which occurs when coexisting life-forms derive mutual benefit from stable associations. While microorganisms receive a secure habitat and constant food source from vertebrate hosts, they are required for optimal immune system development and occupy niches otherwise abused by pathogens. Microbes have also been shown to provide vertebrate hosts with metabolic capabilities that enhance energy and nutrient uptake from the diet. The immune system plays a central role in the establishment and maintenance of host-microbe homeostasis, and B lineage cells play a key role in this regulation. Here, I reviewed the structure and function of the microbiota and the known mechanisms of how nonpathogenic microbes influence B cell biology and immunoglobulin repertoire development early in life. I also discuss what is known about how B lineage cells contribute to the process of shaping the composition of commensal/mutualistic microbe membership.

Tumor necrosis factor receptor 1 (TNFR1)-associated death domain protein (TRADD) is essential in recruiting signaling molecules to the TNFR1 receptor complex. Interferon-γ (IFN-γ) is a potent activator of macrophages and uses signal transducer and activator of transcription 1-α (STAT1-α) for signal transduction. Here we demonstrate that IFN-γ induces the formation of a nuclear-localized TRADD–STAT1-α complex. IFN-γ-mediated STAT1-α phosphorylation was prolonged in cells with reduced TRADD expression. Moreover, we noted an increase in IFN-γ-mediated STAT1-α DNA-binding activity, nuclear presence and transcriptional potential in the TRADD knockdown cells. These data indicate that TRADD may be involved in IFN-γ signaling by forming a complex with STAT1-α within the nucleus and regulating IFN-γ-mediated STAT1-α activation.

Inflammatory events in the central nervous system (CNS) contribute to the disease process in a variety of neuroinflammatory diseases such as multiple sclerosis (MS), Alzheimer's Disease (AD), and cerebral ischemia, and activated macrophages/microglia are central to this response. Immunological activation of these cells leads to the production of a wide array of cytokines, chemokines, matrix metalloproteinases and neurotoxins, and ultimately to glial/neuronal injury and death. The CD40 molecule has an important role in promoting inflammatory responses by macrophages/microglia, since interaction with its cognate ligand, CD154, leads to secretion of cytokines and neurotoxins. Aberrant CD40 expression by macrophages/microglia, induced by cytokines such as IFN-gamma and TNF-alpha, contributes to neuroimmunologic cascades in the CNS. Strategies to suppress CD40 expression may attenuate inflammation and neuronal damage within the CNS, which will ultimately be of benefit in neuroinflammatory diseases. The mediators that regulate expression of CD40 in macrophages/microglia (both induction and inhibition) function at the level of gene transcription. In this review, we present an overview of the molecular basis of CD40 expression in macrophages/microglia. The signal transduction pathways and transcription factors employed by IFN-gamma and TNF-alpha to induce CD40 expression are described, as are the cis-elements in the CD40 promoter that are critical for CD40 transcription. Information is provided on the mechanism(s) underlying suppression of CD40 in macrophages/microglia by immunomodulatory agents such as IL-4, TGF-beta, neuropeptides, neurotrophins, and statins. A comprehensive assessment of CD40 production and function in macrophages/microglia will establish the foundation for future therapeutic manipulation of this critical immunoregulatory protein.

CD40 is a type I membrane-bound molecule belonging to the TNFR superfamily that is expressed on various immune cells including macrophages and microglia. The aberrant expression of CD40 is involved in the initiation and maintenance of various human diseases including multiple sclerosis, arthritis, atherosclerosis, and Alzheimer's disease. Inhibition of CD40 signaling has been shown to provide a significant beneficial effect in a number of animal models of human diseases including the aforementioned examples. We have previously shown that IFN-gamma induces CD40 expression in macrophages and microglia. IFN-gamma leads to STAT-1alpha activation directly and up-regulation of NF-kappaB activity due to the secretion and subsequent autocrine signaling of TNF-alpha. However, TNF-alpha alone is not capable of inducing CD40 expression in these cells. Suppressor of cytokine signaling 1 protein (SOCS-1) is a cytokine-inducible Src homology 2-containing protein that regulates cytokine receptor signaling by inhibiting STAT-1alpha activation via a specific interaction with activated Janus kinase 2. Given the important role of CD40 in inflammatory events in the CNS as well as other organ systems, it is imperative to understand the molecular mechanisms contributing to both CD40 induction and repression. We show that ectopic expression of SOCS-1 abrogates IFN-gamma-induced CD40 protein expression, mRNA levels, and promoter activity. Additionally, IFN-gamma-induced TNF-alpha secretion, as well as STAT-1alpha and NF-kappaB activation, are inhibited in the presence of SOCS-1. We conclude that SOCS-1 inhibits cytokine-induced CD40 expression by blocking IFN-gamma-mediated STAT-1alpha activation, which also then results in suppression of IFN-gamma-induced TNF-alpha secretion and subsequent NF-kappaB activation.

Immunoglobulin heavy chain (IgH) isotypes (e.g. IgM, IgG and IgE) are generated as secreted/soluble antibodies (sIg) or as membrane-bound (mIg) B cell receptors (BCR) through alternative RNA splicing. IgH isotype dictates soluble antibody function, but how mIg isotype influences B cell behavior is unclear. We examined IgH isotype-specific BCR function by engineering polyclonal Ighγ1/γ1 and Ighɛ/ɛ mice, which initially produce IgG1 or IgE from their respective native genomic configurations. Transcripts producing IgM, IgG1, and IgE are produced in an alternative splice form bias hierarchy, where mIgμ>mIgγ1>mIgɛ—influencing respective BCR densities and fitness for populating the periphery—a pattern replicated in activated and memory B cells from wild-type mice. Restrained B cell development from Ighγ1/γ1 and Ighɛ/ɛ mice was proportional to sIg/mIg ratios, and was rescued by enforced expression of the respective mIgs. In addition, enhanced BCR signaling by way of PTEN deletion in IgE B cells generated functional IgE memory responses from IgE memory B cells themselves, which are normally absent under normal conditions, indicating that weak signal strength normally restrains IgE responses. Thus, IgH isotype-specific mIg dosage regulates B cell fate by influencing quantitative baseline BCR signal strength.

B lymphocyte immunoglobulin heavy chain (IgH) class switch recombination (CSR) is a process wherein initially expressed IgM switches to other IgH isotypes, such as IgA, IgE and IgG. Measurement of IgH CSR in vitro is a key method for the study of a number of biologic processes ranging from DNA recombination and repair to aspects of molecular and cellular immunology. In vitro CSR assay involves the flow cytometric measurement surface Ig expression on activated B cells. While measurement of IgA and IgG subclasses is straightforward, measurement of IgE by this method is problematic due to soluble IgE binding to FcεRII/CD23 expressed on the surface of activated B cells. Here we describe a unique procedure for accurate measurement of IgE-producing mouse B cells that have undergone CSR in culture. The method is based on trypsin-mediated cleavage of IgE-CD23 complexes on cell surfaces, allowing for detection of IgE-producing B lineage cells by cytoplasmic staining. This procedure offers a convenient solution for flow cytometric analysis of CSR to IgE.

The N-terminal variable regions of immunoglobulin (Ig) heavy (IgH) and light (IgL) chains are involved in specific antigen binding and assembled from germline variable (V), diversity (D), and joining (J) gene segments through a somatic recombination reaction known as V(D)J recombination. V(D)J recombination is initiated by RAG endonuclease and completed by non-homologous DNA end-joining. V(D)J recombination is tightly regulated in the contexts of order, lineage, and allelic exclusion. For IgL this regulation additionally involves isotypic exclusion, in which most B cells express Ig molecules containing only one of the two IgL isotypes, κ and λ. Controlled V(D)J recombination is important beyond its role in generating diverse antibody repertoires against diverse antigens, since dysregulation of V(D)J recombination can not only underlie various immune disorders, but lead to oncogenic translocations that are frequently found in human B and T cell leukemias and lymphomas. Many of such translocations involve IgL chain loci. To provide new insights into mechanisms of chromosomal translocations involving IgL chain loci, we sought to investigate mechanisms of normal V(D)J recombination at IgL loci by developing and applying multiple new approaches. In this study, we employed high-throughput genome-wide translocation sequencing (HTGTS) to identify RAG activities at Igκ and Igλ loci in v-Abl transformed progenitor B (pro-B) line. Remarkably, we found such RAG activities were largely confined to unanticipated distinct chromatin topologically associated loop domains within Igκ and Igλ loci, adding a new layer of regulation of gene rearrangements at IgL loci. Furthermore, we developed a highly sensitive and unbiased assay, referred to as HTGTS repertoire sequencing (HTGTS-Rep-seq), to quantify Igκ and Igλ repertoires in mouse bone marrow precursor B (pre-B) and splenic B cell populations. HTGTS-Rep-seq detected diverse Igκ and Igλ VJ rearrangements involving individual functional Jκ and Jλ segments, respectively. Currently we are integrating such enormous Igκ and Igλ repertoire data with long-range chromatin interaction map data revealed by Hi-C and our recently developed high-resolution HTGTS chromosome conformation capture sequencing (HTGTS-3C-seq) methods to further address the mechanisms of gene rearrangement regulation by chromatin loop domains at these loci. In addition, to address the mechanisms of isotypic exclusion that has been debated, we generated germline mouse models harboring endogenously-generated Igκ deletions and a productively assembled IgH variable region exon by generating induced-pluripotent stems cells (iPSCs) from peripheral B cells. We then applied global nuclear run-on sequencing (GRO-seq) to bone marrow pre-B cells purified from iPSC-generated mice in a RAG-deficient background (which abrogates V(D)J recombination) to elucidate transcriptional features at Igκ and Igλ loci and specifically to reveal potential transcriptional alterations at Igλ in the context of Igκ deletion. Compared with Igκ-intact RAG-deficient pre-B cells, Igκ-deleted RAG-deficient pre-B cells showed remarkably increased transcription across the whole Igλ locus, indicating specific Igκ-deletion-triggered transcriptional activation at Igλ. Citation Format: Zhaoqing Ba, Jiazhi Hu, Zhou Du, Sherry G. Lin, Duane R. Wesemann, Frederick W. Alt. Mechanisms that mediate intralocus and interlocus regulation of V(D)J recombination at immunoglobulin light chain loci [abstract]. In: Proceedings of the Second CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; 2016 Sept 25-28; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2016;4(11 Suppl):Abstract nr A033.

Induced pluripotent stem cells (iPSCs) can be formed from somatic cells by a defined set of genetic factors; however, aberrant epigenetic silencing of the imprinted Dlk1-Dio3 gene cluster often hinders their developmental potency and ability to contribute to high-grade chimerism in mice. Here, we describe an approach that allows splenic B cells activated to undergo Ig heavy-chain ( IgH ) class-switch recombination (CSR) to be reprogrammed into iPSCs that contribute to high-grade chimerism in mice. Treatment of naïve splenic B cells in culture with anti-CD40 plus IL-4 induces IgH CSR from IgM to IgG1 and IgE. CSR leads to irreversible IgH locus deletions wherein the IgM-producing Cμ exons are permanently excised from the B-cell genome. We find that anti-CD40 plus IL-4–activated B cells produce iPSCs that are uniformly hypermethylated in the imprinted Dlk1-Dio3 gene cluster and fail to produce chimerism in mice. However, treatment of activated B cells with the methyltransferase inhibitor 5-aza-2′-deoxycytidine before and at early stages of reprogramming attenuates hypermethylation of the Dlk1-Dio3 locus in resultant iPSCs and enables them to form high-grade chimerism in mice. These conditions allowed us to produce chimeric mice in which all mature B cells were derived entirely from IgG1-expressing B-cell–derived iPSCs. We conclude that culture conditions of activated B cells before and at early stages of reprogramming influence the developmental potency of resultant iPSCs.

Variable, diversity and joining gene segment (V(D)J) recombination assembles immunoglobulin heavy or light chain (IgH or IgL) variable region exons in developing bone marrow B cells, whereas class switch recombination (CSR) exchanges IgH constant region exons in peripheral B cells. Both processes use directed DNA double-strand breaks (DSBs) repaired by non-homologous end-joining (NHEJ). Errors in either V(D)J recombination or CSR can initiate chromosomal translocations, including oncogenic IgH locus (Igh) to c-myc (also known as Myc) translocations of peripheral B cell lymphomas. Collaboration between these processes has also been proposed to initiate translocations. However, the occurrence of V(D)J recombination in peripheral B cells is controversial. Here we show that activated NHEJ-deficient splenic B cells accumulate V(D)J-recombination-associated breaks at the lambda IgL locus (Igl), as well as CSR-associated Igh breaks, often in the same cell. Moreover, Igl and Igh breaks are frequently joined to form translocations, a phenomenon associated with specific Igh-Igl co-localization. Igh and c-myc also co-localize in these cells; correspondingly, the introduction of frequent c-myc DSBs robustly promotes Igh-c-myc translocations. Our studies show peripheral B cells that attempt secondary V(D)J recombination, and determine a role for mechanistic factors in promoting recurrent translocations in tumours.

Malignant gliomas are the most common primary brain tumors in adults, and the most malignant form, glioblastoma multiforme (GBM), is usually rapidly fatal. Most GBMs do not have p53 mutations, although the p53 tumor suppressor pathway appears to be inactivated. GBMs grow in a hypoxic and inflammatory microenvironment, and increased levels of the free radicals nitric oxide (NO) and superoxide () occur in these malignancies in vivo. Peroxynitrite (ONOO(-)) is a highly reactive molecule produced by excess NO and that can posttranslationally modify and inactivate proteins, especially zinc finger transcription factors such as p53. We demonstrated previously that GBMs have evidence of tyrosine nitration, the "footprint" of peroxynitrite-mediated protein modification in vivo, and that peroxynitrite could inhibit the specific DNA binding ability of wild-type p53 protein in glioma cells in vitro. Here we show that both authentic peroxynitrite and SIN-1 (3-morpholinosydnonimine hydrochloride), a molecule that decomposes into NO and to form peroxynitrite, can inhibit wild-type p53 function in malignant glioma cells. Concentrations of peroxynitrite associated with a tumor inflammatory environment caused dysregulation of wild-type p53 transcriptional activity and downstream p21(WAF1) expression.

Tumor necrosis factor receptor 1 (TNFR1)-associated death domain protein (TRADD) is essential in recruiting signaling molecules to the TNFR1 receptor complex. Interferon-gamma (IFN-gamma) is a potent activator of macrophages and uses signal transducer and activator of transcription 1-alpha (STAT1-alpha) for signal transduction. Here we demonstrate that IFN-gamma induces the formation of a nuclear-localized TRADD-STAT1-alpha complex. IFN-gamma-mediated STAT1-alpha phosphorylation was prolonged in cells with reduced TRADD expression. Moreover, we noted an increase in IFN-gamma-mediated STAT1-alpha DNA-binding activity, nuclear presence and transcriptional potential in the TRADD knockdown cells. These data indicate that TRADD may be involved in IFN-gamma signaling by forming a complex with STAT1-alpha within the nucleus and regulating IFN-gamma-mediated STAT1-alpha activation.

Since the discovery of the STAT proteins, there have been a great many reports on the involvement of STAT signals with other signaling pathways. These interactions can be classified into at least four general groups. First, STAT-mediated signals may be the means of the transcriptional upregulation of genes that impact on other pathways. Second, STATs interact with other transcriptional regulators at the promoter level. These interactions occur as STATs bind regions of DNA that are adjacent to or overlap with binding sites for other transcription factors. Moreover, STATs contain binding sites for a number of different co-activators such as the CREB binding protein (CBP) that enables the indirect association with many other transcriptional regulators such as those involved in TGF-β signaling. Third, STAT function can be manipulated by post-translational modification induced by other signaling pathways. An example of such is serine phosphorylation that can be induced by kinases that are activated by other signaling pathways (i.e. TNF-α and IL-1β). And fourth, STATs have been shown to interact with components from other signaling pathways that are not linked to DNA binding or transcription, such as the TNF Receptor 1 (TNFR1) the TNF-Associated Death Domain protein (TRADD). Demonstration of these interactions has provided clues to the mechanisms involved in how cells can integrate multiple signals and respond in a specific manner to a given set of signals from the environment. This chapter will review the interactions of various STAT molecules with signaling components from the TNF, IL-1β, TGF-β and hormone receptor signaling pathways. Specifically, the focus will be on STAT interactions with NF-кB, TNFR1, TRADD, SMADs and glucocorticoid receptor. Since this is not a comprehensive outline of all of the interaction partners of the STATs, emphasis will be placed on the general modes of interaction and the relation to functional modification.

Progenitor (pro-) and precursor (pre-) B cells express the recombination-activating gene (RAG1/RAG2) endonuclease, which initiates the V(D)J recombination reaction that assembles Immunoglobulin heavy (IgH) and light (IgL) chain variable region exons from germline gene segments. Expression of productively assembled IgH μ chains and IgL (Igκ or Igλ) chains generates IgM molecules that form the B-cell antigen binding receptor (BCR) with a diverse primary repertoire of binding specificities. As the newly assembled and expressed IgM interacts with local antigens, RAG expression can continue, allowing continued Igκ V(D)J recombination that can replace the previously assembled VκJκ exon with one that generates a new specificity. This "receptor editing" process provides an antigen-mediated regulatory checkpoint that helps shape the pre-immune BCR repertoire. As the principal location of early B-cell development, the bone marrow antigenic environment is the main force affecting receptor editing. However, the extent to which self-antigens outside of the bone marrow — as well as environmental antigens — may shape the primary Ig repertoire is not known. Here we show that early B-cell development takes place in peripheral sites, including the small intestinal mucosa, in young mice. Cells in these sites co-express RAG with cytoplasmic Igκ and Igμ, indicating active receptor editing; and, correspondingly, they have significant differences in Vκ usage compared to RAG2-expressing bone marrow B-lineage cells and other RAG2-expressing B-lineage cells from other peripheral sites. In addition, the luminal microbial environment appears to also affect Igλ/Igκ ratios. We conclude that early B-cell development can occur in the periphery, including the intestinal mucosa, where commensal microbes may influence BCR diversification and gut pre-immune Ig repertoires. Disclosures No relevant conflicts of interest to declare.

Mammalian tRNA 3' processing endoribonuclease (3' tRNase) can remove a 3' trailer from various pre-tRNAs without 5' leader nucleotides. To examine how 5[prime] leader sequences affect 3' processing efficiency, we performed in vitro 3' processing reactions with purified pig 3' tRNase and pre-tRNAArgs containing a 13-nt 3' trailer and a 5[prime] leader of various lengths. The 3' processing was slightly stimulated by 5[prime] leaders containing up to 7 nt, whereas leaders of 9 nt or longer severely inhibited the reaction. Structure probing indicated that the 5' leader sequences had little effect on pre-tRNA folding. Similar results were obtained using pre-tRNA(Val)s containing a 5' leader of various lengths. We also investigated whether 3'tRNase can remove 3' trailers that are stably base-paired with 5' leaders to form an extended acceptor stem. Even such small 5' leaders as 3 and 6 nt, when base-paired with a 3' trailer, severely hindered removal of the 3' trailer by 3' tRNase.

The classical nonhomologous end-joining (C-NHEJ) DNA double-strand break (DSB) repair pathway employs the Ku70/80 complex (Ku) for DSB recognition and the XRCC4/DNA ligase 4 (Lig4) complex for ligation. During IgH class switch recombination (CSR) in B lymphocytes, switch (S) region DSBs are joined by C-NHEJ to form junctions either with short microhomologies (MHs; “MH-mediated” joins) or no homologies (“direct” joins). In the absence of XRCC4 or Lig4, substantial CSR occurs via “alternative” end-joining (A-EJ) that generates largely MH-mediated joins. Because upstream C-NHEJ components remain in XRCC4- or Lig4-deficient B cells, residual CSR might be catalyzed by C-NHEJ using a different ligase. To address this, we have assayed for CSR in B cells deficient for Ku70, Ku80, or both Ku70 and Lig4. Ku70- or Ku80-deficient B cells have reduced, but still substantial, CSR. Strikingly, B cells deficient for both Ku plus Lig4 undergo CSR similarly to Ku-deficient B cells, firmly demonstrating that an A-EJ pathway distinct from C-NHEJ can catalyze CSR end-joining. Ku-deficient or Ku- plus Lig4-deficient B cells are also biased toward MH-mediated CSR joins; but, in contrast to XRCC4- or Lig4-deficient B cells, generate substantial numbers of direct CSR joins. Our findings suggest that more than one form of A-EJ can function in CSR.

Microglia and macrophages are important antigen-presenting cells (APCs) in the central nervous system (CNS). By virtue of their ability to express class II MHC antigens and costimulatory molecules such as CD40 and B7, microglia/macrophages promote Th1 cell activation and subsequent immune/inflammatory responses within the CNS. We have previously demonstrated that IFN-γ is the most potent inducer of CD40 expression on microglia. Our more recent studies have focused on the molecular basis of IFN-γ induced CD40 expression, and mechanisms by which this gene can be inhibited. The suppressive effects of IL-4 on CD40 expression will be discussed, as will the involvement of SH2 containing proteins called SOCS (for Suppressors Of Cytokine Signalling). Expression of CD40 by activated microglia/macrophages may contribute to the complex neuroimmunologic cascades that result in inflammation, demyelination and neuronal dysfunction. As such, understanding the mechanisms of inhibition of this molecule will be beneficial in diseases such as multiple sclerosis, HIV-1 associated dementia and Alzheimer's disease.

Nature Immunology 5, 199–207 (2004). On page 204, column 2, line 3, the number should read “(1,700%).” Nature Immunology regrets the error.

Nature Immunology 5, 199–207 (2004). Figure 3b should have a line separating the top and bottom halves.