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Blimp-1 is known to act as a transcriptional repressor by suppressing genes associated with mature and activated germinal center B cells. Busslinger and colleagues show that Blimp-1 can also directly activate gene expression in plasma cells, including those encoding proteins that regulate the production of membrane-bound immunoglobulin versus secreted immunoglobulin. The transcription factor Blimp-1 is necessary for the generation of plasma cells. Here we studied its functions in plasmablast differentiation by identifying regulated Blimp-1 target genes. Blimp-1 promoted the migration and adhesion of plasmablasts. It directly repressed genes encoding several transcription factors and Aicda (which encodes the cytidine deaminase AID) and thus silenced B cell–specific gene expression, antigen presentation and class-switch recombination in plasmablasts. It directly activated genes, which led to increased expression of the plasma cell regulator IRF4 and proteins involved in immunoglobulin secretion. Blimp-1 induced the transcription of immunoglobulin genes by controlling the 3′ enhancers of the loci encoding the immunoglobulin heavy chain ( Igh ) and κ-light chain ( Igk ) and, furthermore, regulated the post-transcriptional expression switch from the membrane-bound form of the immunoglobulin heavy chain to its secreted form by activating Ell2 (which encodes the transcription-elongation factor ELL2). Notably, Blimp-1 recruited chromatin-remodeling and histone-modifying complexes to regulate its target genes. Hence, many essential functions of plasma cells are under the control of Blimp-1.

Induction of the transcription factor Irf8 in the common dendritic cell progenitor (CDP) is required for classical type 1 dendritic cell (cDC1) fate specification, but the mechanisms controlling this induction are unclear. In the present study Irf8 enhancers were identified via chromatin profiling of dendritic cells and CRISPR/Cas9 genome editing was used to assess their roles in Irf8 regulation. An enhancer 32 kilobases (kb) downstream of the Irf8 transcriptional start site (+32-kb Irf8 ) that was active in mature cDC1s was required for the development of this lineage, but not for its specification. Instead, a +41-kb Irf8 enhancer, previously thought to be active only in plasmacytoid dendritic cells, was found to also be transiently accessible in cDC1 progenitors, and deleting this enhancer prevented the induction of Irf8 in CDPs and abolished cDC1 specification. Thus, cryptic activation of the +41-kb Irf8 enhancer in dendritic cell progenitors is responsible for cDC1 fate specification. The transcription factor IRF8 is essential for classical type 1 dendritic cell (cDC1) development. Murphy and colleagues investigate in detail the molecular control of cDC1 fate specification by systematically unpicking the IRF8 enhancer regions.

The establishment of a diverse B cell antigen receptor (BCR) repertoire by V(D)J recombination also generates autoreactive B cells. Anergy is one tolerance mechanism; it renders autoreactive B cells insensitive to stimulation by self-antigen, whereas Toll-like receptor (TLR) signaling can reactivate anergic B cells. Here, we describe a critical role of the transcription factor Ikaros in controlling BCR anergy and TLR signaling. Mice with specific deletion of Ikaros in mature B cells developed systemic autoimmunity. Ikaros regulated many anergy-associated genes, including Zfp318 , which is implicated in the attenuation of BCR responsiveness by promoting immunoglobulin D expression in anergic B cells. TLR signaling was hyperactive in Ikaros-deficient B cells, which failed to upregulate feedback inhibitors of the MyD88–nuclear factor κB signaling pathway. Systemic inflammation was lost on expression of a non-self-reactive BCR or loss of MyD88 in Ikaros-deficient B cells. Thus, Ikaros acts as a guardian preventing autoimmunity by promoting BCR anergy and restraining TLR signaling. Immature B cells expressing self-reactive BCRs induce anergy programs to promote tolerance. Busslinger and colleagues show that the transcription factor Ikaros enforces anergy by inducing transcription of negative-feedback regulators of the BCR and TLR–MyD88 pathways.

Extended loop extrusion across the immunoglobulin heavy-chain ( Igh ) locus facilitates V H -DJ H recombination following downregulation of the cohesin-release factor Wapl by Pax5, resulting in global changes in the chromosomal architecture of pro-B cells. Here, we demonstrate that chromatin looping and V K -J K recombination at the Igk locus were insensitive to Wapl upregulation in pre-B cells. Notably, the Wapl protein was expressed at a 2.2-fold higher level in pre-B cells compared with pro-B cells, which resulted in a distinct chromosomal architecture with normal loop sizes in pre-B cells. High-resolution chromosomal contact analysis of the Igk locus identified multiple internal loops, which likely juxtapose V K and J K elements to facilitate V K -J K recombination. The higher Wapl expression in Igμ-transgenic pre-B cells prevented extended loop extrusion at the Igh locus, leading to recombination of only the 6 most 3’ proximal V H genes and likely to allelic exclusion of all other V H genes in pre-B cells. These results suggest that pro-B and pre-B cells with their distinct chromosomal architectures use different chromatin folding principles for V gene recombination, thereby enabling allelic exclusion at the Igh locus, when the Igk locus is recombined. V gene recombination at the immunoglobulin heavy chain locus ( Igh ) is facilitated by extended loop extrusion. In this study, the authors find that, unlike Igh , the κ light chain locus does not involve extended loop extrusion but instead involves multiple, short-range loops for V gene combination.

B-1a B cells are a distinct subset of mature B cells that provide innate-like protection against pathogens. Busslinger and colleagues identify the transcription factor Bhlhe41 as being essential for B-1a development and self-renewal. Innate-like B-1a cells provide a first line of defense against pathogens, yet little is known about their transcriptional control. Here we identified an essential role for the transcription factor Bhlhe41, with a lesser contribution by Bhlhe40, in controlling B-1a cell differentiation. Bhlhe41 −/− Bhlhe40 −/− B-1a cells were present at much lower abundance than were their wild-type counterparts. Mutant B-1a cells exhibited an abnormal cell-surface phenotype and altered B cell receptor (BCR) repertoire exemplified by loss of the phosphatidylcholine-specific V H 12V κ 4 BCR. Expression of a pre-rearranged V H 12V κ 4 BCR failed to 'rescue' the mutant phenotype and revealed enhanced proliferation accompanied by increased cell death. Bhlhe41 directly repressed the expression of cell-cycle regulators and inhibitors of BCR signaling while enabling pro-survival cytokine signaling. Thus, Bhlhe41 controls the development, BCR repertoire and self-renewal of B-1a cells.

The transcription factor Ikaros is required for lymphopoiesis. Busslinger and colleagues show Ikaros positively regulates genes encoding pre-BCR signal transducers and thereby promotes pro-B to pre-B cell progression and proliferation. The transcription factor Ikaros is an essential regulator of lymphopoiesis. Here we studied its B cell–specific function by conditional inactivation of the gene encoding Ikaros ( Ikzf1 ) in pro-B cells. B cell development was arrested at an aberrant 'pro-B cell' stage characterized by increased cell adhesion and loss of signaling via the pre-B cell signaling complex (pre-BCR). Ikaros activated genes encoding signal transducers of the pre-BCR and repressed genes involved in the downregulation of pre-BCR signaling and upregulation of the integrin signaling pathway. Unexpectedly, derepression of expression of the transcription factor Aiolos did not compensate for the loss of Ikaros in pro-B cells. Ikaros induced or suppressed active chromatin at regulatory elements of activated or repressed target genes. Notably, binding of Ikaros and expression of its target genes were dynamically regulated at distinct stages of early B lymphopoiesis.

Pax5 is a critical regulator of B‐cell commitment. Here, we identified direct Pax5 target genes by streptavidin‐mediated ChIP‐chip analysis of pro‐B cells expressing in vivo biotinylated Pax5. By binding to promoters and enhancers, Pax5 directly regulates the expression of multiple transcription factor, cell surface receptor and signal transducer genes. One of the newly identified enhancers was shown by transgenic analysis to confer Pax5‐dependent B‐cell‐specific activity to the Nedd9 gene controlling B‐cell trafficking. Profiling of histone modifications in Pax5‐deficient and wild‐type pro‐B cells demonstrated that Pax5 induces active chromatin at activated target genes, while eliminating active chromatin at repressed genes in committed pro‐B cells. Pax5 rapidly induces these chromatin and transcription changes by recruiting chromatin‐remodelling, histone‐modifying and basal transcription factor complexes to its target genes. These data provide novel insight into the regulatory network and epigenetic regulation, by which Pax5 controls B‐cell commitment. The mechanism of Pax5‐mediated gene activation and repression during early B‐cell development is unclear. This study identifies Pax5‐binding sites in pro‐B cells and the changes in chromatin modifications induced by the recruitment of chromatin‐modifying and transcription factors.

The gene encoding the receptor for macrophage colony-stimulating factor (CSF-1R) is expressed exclusively in cells of the myeloid lineages as well as trophoblasts. A conserved element in the second intron, Fms-Intronic Regulatory Element (FIRE), is essential for macrophage-specific transcription of the gene. However, the molecular details of how FIRE activity is regulated and how it impacts the Csf1r promoter have not been characterised. Here we show that agents that down-modulate Csf1r mRNA transcription regulated promoter activity altered the occupancy of key FIRE cis-acting elements including RUNX1, AP1, and Sp1 binding sites. We demonstrate that FIRE acts as an anti-sense promoter in macrophages and reversal of FIRE orientation within its native context greatly reduced enhancer activity in macrophages. Mutation of transcription initiation sites within FIRE also reduced transcription. These results demonstrate that FIRE is an orientation-specific transcribed enhancer element.

The murine c‐fms ( Csf1r ) gene encodes the macrophage colony‐stimulating factor receptor, which is essential for macrophage development. It is expressed at a low level in haematopoietic stem cells and is switched off in all non‐macrophage cell types. To examine the role of chromatin structure in this process we studied epigenetic silencing of c‐fms during B‐lymphopoiesis. c‐fms chromatin in stem cells and multipotent progenitors is in the active conformation and bound by transcription factors. A similar result was obtained with specified common myeloid and lymphoid progenitor cells. In developing B cells, c‐fms chromatin is silenced in distinct steps, whereby first the binding of transcription factors and RNA expression is lost, followed by a loss of nuclease accessibility. Interestingly, regions of de novo DNA methylation in B cells overlap with an intronic antisense transcription unit that is differently regulated during lymphopoiesis. However, even at mature B cell stages, c‐fms chromatin is still in a poised conformation and c‐fms expression can be re‐activated by conditional deletion of the transcription factor Pax5.

Nuclear processes, such as V(D)J recombination, are orchestrated by the three-dimensional organization of chromosomes at multiple levels, including compartments 1 and topologically associated domains (TADs) 2 , 3 consisting of chromatin loops 4 . TADs are formed by chromatin-loop extrusion 5 – 7 , which depends on the loop-extrusion function of the ring-shaped cohesin complex 8 – 12 . Conversely, the cohesin-release factor Wapl 13 , 14 restricts loop extension 10 , 15 . The generation of a diverse antibody repertoire, providing humoral immunity to pathogens, requires the participation of all V genes in V(D)J recombination 16 , which depends on contraction of the 2.8-Mb-long immunoglobulin heavy chain ( Igh ) locus by Pax5 17 , 18 . However, how Pax5 controls Igh contraction in pro-B cells remains unknown. Here we demonstrate that locus contraction is caused by loop extrusion across the entire Igh locus. Notably, the expression of Wapl is repressed by Pax5 specifically in pro-B and pre-B cells, facilitating extended loop extrusion by increasing the residence time of cohesin on chromatin. Pax5 mediates the transcriptional repression of Wapl through a single Pax5-binding site by recruiting the polycomb repressive complex 2 to induce bivalent chromatin at the Wapl promoter. Reduced Wapl expression causes global alterations in the chromosome architecture, indicating that the potential to recombine all V genes entails structural changes of the entire genome in pro-B cells. Pax5 regulates contraction of the immunoglobulin heavy chain ( Igh ) locus—an essential step in V(D)J recombination—by promoting chromatin loop extrusion via repression of Wapl expression.