Infection perturbs Bach2- and Bach1-dependent erythroid lineage 'choice' to cause anemia.

Elucidation of how the differentiation of hematopoietic stem and progenitor cells (HSPCs) is reconfigured in response to the environment is critical for understanding the biology and disorder of hematopoiesis. Here we found that the transcription factors (TFs) Bach2 and Bach1 promoted erythropoiesis by regulating heme metabolism in committed erythroid cells to sustain erythroblast maturation and by reinforcing erythroid commitment at the erythro-myeloid bifurcation step. Bach TFs repressed expression of the gene encoding the transcription factor C/EBPß, as well as that of its target genes encoding molecules important for myelopoiesis and inflammation; they achieved the latter by binding to their regulatory regions also bound by C/EBPß. Lipopolysaccharide diminished the expression of Bach TFs in progenitor cells and promoted myeloid differentiation. Overexpression of Bach2 in HSPCs promoted erythroid development and inhibited myelopoiesis. Knockdown of BACH1 or BACH2 in human CD34+ HSPCs impaired erythroid differentiation in vitro. Thus, Bach TFs accelerate erythroid commitment by suppressing the myeloid program at steady state. Anemia of inflammation and myelodysplastic syndrome might involve reduced activity of Bach TFs.

BACH2 regulates CD8(+) T cell differentiation by controlling access of AP-1 factors to enhancers.

T cell antigen receptor (TCR) signaling drives distinct responses depending on the differentiation state and context of CD8(+) T cells. We hypothesized that access of signal-dependent transcription factors (TFs) to enhancers is dynamically regulated to shape transcriptional responses to TCR signaling. We found that the TF BACH2 restrains terminal differentiation to enable generation of long-lived memory cells and protective immunity after viral infection. BACH2 was recruited to enhancers, where it limited expression of TCR-driven genes by attenuating the availability of activator protein-1 (AP-1) sites to Jun family signal-dependent TFs. In naive cells, this prevented TCR-driven induction of genes associated with terminal differentiation. Upon effector differentiation, reduced expression of BACH2 and its phosphorylation enabled unrestrained induction of TCR-driven effector programs.

The transcription repressors Bach2 and Bach1 promote B cell development by repressing the myeloid program.

Mature lymphoid cells express the transcription repressor Bach2, which imposes regulation on humoral and cellular immunity. Here we found critical roles for Bach2 in the development of cells of the B lineage, commencing from the common lymphoid progenitor (CLP) stage, with Bach1 as an auxiliary. Overexpression of Bach2 in pre-pro-B cells deficient in the transcription factor EBF1 and single-cell analysis of CLPs revealed that Bach2 and Bach1 repressed the expression of genes important for myeloid cells ('myeloid genes'). Bach2 and Bach1 bound to presumptive regulatory regions of the myeloid genes. Bach2(hi) CLPs showed resistance to myeloid differentiation even when cultured under myeloid conditions. Our results suggest that Bach2 functions with Bach1 and EBF1 to promote B cell development by repressing myeloid genes in CLPs.

TANK Binding Kinase 1 Promotes BACH1 Degradation through Both Phosphorylation-Dependent and -Independent Mechanisms without Relying on Heme and FBXO22.

BTB and CNC homology 1 (BACH1) represses the expression of genes involved in the metabolism of iron, heme and reactive oxygen species. While BACH1 is rapidly degraded when it is bound to heme, it remains unclear how BACH1 degradation is regulated under other conditions. We found that FBXO22, a ubiquitin ligase previously reported to promote BACH1 degradation, polyubiquitinated BACH1 only in the presence of heme in a highly purified reconstitution assay. In parallel to this regulatory mechanism, TANK binding kinase 1 (TBK1), a protein kinase that activates innate immune response and regulates iron metabolism via ferritinophagy, was found to promote BACH1 degradation when overexpressed in 293T cells. While TBK1 phosphorylated BACH1 at multiple serine and threonine residues, BACH1 degradation was observed with not only the wild-type TBK1 but also catalytically impaired TBK1. The BACH1 degradation in response to catalytically impaired TBK1 was not dependent on FBXO22 but involved both autophagy-lysosome and ubiquitin-proteasome pathways judging from its suppression by using inhibitors of lysosome and proteasome. Chemical inhibition of TBK1 in hepatoma Hepa1 cells showed that TBK1 was not required for the heme-induced BACH1 degradation. Its inhibition in Namalwa B lymphoma cells increased endogenous BACH1 protein. These results suggest that TBK1 promotes BACH1 degradation in parallel to the FBXO22- and heme-dependent pathway, placing BACH1 as a downstream effector of TBK1 in iron metabolism or innate immune response.

Bach2 Promotes B Cell Receptor-Induced Proliferation of B Lymphocytes and Represses Cyclin-Dependent Kinase Inhibitors.

BTB and CNC homology 2 (Bach2) is a transcriptional repressor that is required for the formation of the germinal center (GC) and reactions, including class switch recombination and somatic hypermutation of Ig genes in B cells, within the GC. Although BCR-induced proliferation is essential for GC reactions, the function of Bach2 in regulating B cell proliferation has not been elucidated. In this study, we demonstrate that Bach2 is required to sustain high levels of B cell proliferation in response to BCR signaling. Following BCR engagement in vitro, B cells from Bach2-deficient (Bach2-/-) mice showed lower incorporation of BrdU and reduced cell cycle progression compared with wild-type cells. Bach2-/- B cells also underwent increased apoptosis, as evidenced by an elevated frequency of sub-G1 cells and early apoptotic cells. Transcriptome analysis of BCR-engaged B cells from Bach2-/- mice revealed reduced expression of the antiapoptotic gene Bcl2l1 encoding Bcl-xL and elevated expression of cyclin-dependent kinase inhibitor (CKI) family genes, including Cdkn1a, Cdkn2a, and Cdkn2b Reconstitution of Bcl-xL expression partially rescued the proliferation defect of Bach2-/- B cells. Chromatin immunoprecipitation experiments showed that Bach2 bound to the CKI family genes, indicating that these genes are direct repression targets of Bach2. These findings identify Bach2 as a requisite factor for sustaining high levels of BCR-induced proliferation, survival, and cell cycle progression, and it promotes expression of Bcl-xL and repression of CKI genes. BCR-induced proliferation defects may contribute to the impaired GC formation observed in Bach2-/- mice.

BACH2 represses effector programs to stabilize T(reg)-mediated immune homeostasis.

Through their functional diversification, distinct lineages of CD4(+) T cells can act to either drive or constrain immune-mediated pathology. Transcription factors are critical in the generation of cellular diversity, and negative regulators antagonistic to alternate fates often act in conjunction with positive regulators to stabilize lineage commitment. Genetic polymorphisms within a single locus encoding the transcription factor BACH2 are associated with numerous autoimmune and allergic diseases including asthma, Crohn's disease, coeliac disease, vitiligo, multiple sclerosis and type 1 diabetes. Although these associations point to a shared mechanism underlying susceptibility to diverse immune-mediated diseases, a function for BACH2 in the maintenance of immune homeostasis has not been established. Here, by studying mice in which the Bach2 gene is disrupted, we define BACH2 as a broad regulator of immune activation that stabilizes immunoregulatory capacity while repressing the differentiation programs of multiple effector lineages in CD4(+) T cells. BACH2 was required for efficient formation of regulatory (Treg) cells and consequently for suppression of lethal inflammation in a manner that was Treg-cell-dependent. Assessment of the genome-wide function of BACH2, however, revealed that it represses genes associated with effector cell differentiation. Consequently, its absence during Treg polarization resulted in inappropriate diversion to effector lineages. In addition, BACH2 constrained full effector differentiation within TH1, TH2 and TH17 cell lineages. These findings identify BACH2 as a key regulator of CD4(+) T-cell differentiation that prevents inflammatory disease by controlling the balance between tolerance and immunity.

Inflammatory responses induce an identity crisis of alveolar macrophages, leading to pulmonary alveolar proteinosis.

Pulmonary alveolar proteinosis (PAP) is a severe respiratory disease characterized by dyspnea caused by accumulation of surfactant protein. Dysfunction of alveolar macrophages (AMs), which regulate the homeostasis of surfactant protein, leads to the development of PAP; for example, in mice lacking BTB and CNC homology 2 (Bach2). However, how Bach2 helps prevent PAP is unknown, and the cell-specific effects of Bach2 are undefined. Using mice lacking Bach2 in specific cell types, we found that the PAP phenotype of Bach2-deficient mice is due to Bach2 deficiency in more than two types of immune cells. Depletion of hyperactivated T cells in Bach2-deficient mice restored normal function of AMs and ameliorated PAP. We also found that, in Bach2-deficient mice, hyperactivated T cells induced gene expression patterns that are specific to other tissue-resident macrophages and dendritic cells. Moreover, Bach2-deficient AMs exhibited a reduction in cell cycle progression. IFN-γ released from T cells induced Bach2 expression in AMs, in which Bach2 then bound to regulatory regions of inflammation-associated genes in myeloid cells. Of note, in AMs, Bach2 restricted aberrant responses to excessive T cell-induced inflammation, whereas, in T cells, Bach2 puts a brake on T cell activation. Moreover, Bach2 stimulated the expression of multiple histone genes in AMs, suggesting a role of Bach2 in proper histone expression. We conclude that Bach2 is critical for the maintenance of AM identity and self-renewal in inflammatory environments. Treatments targeting T cells may offer new therapeutic strategies for managing secondary PAP.

Orchestration of plasma cell differentiation by Bach2 and its gene regulatory network.

Bach2 is a basic region-leucine zipper (bZip) transcription factor that forms heterodimers with small Maf oncoproteins and binds to target genes, thus repressing their expression. Bach2 is required for class switch recombination (CSR) and somatic hypermutation (SHM) of immunoglobulin genes in activated B cells. Bach2 represses the expression of Prdm1 encoding Blimp-1 repressor and thereby inhibits terminal differentiation of B cells to plasma cells. This causes a delay in the induction of Prdm1, thereby securing a time window for the expression of Aicda encoding activation-induced cytidine deaminase (AID) required for both CSR and SHM. Based on the characteristics of a gene regulatory network (GRN) involving Bach2 and Prdm1 and its dynamics, a 'delay-driven diversity' model was introduced to explain the responses of activated B cells. Bach2 is also required for the proper differentiation and function of peripheral T cells. In the absence of Bach2, CD4(+) T cells show increased differentiation to effector cells producing higher levels of Th2-related cytokines, such as IL-4 and IL-10, and a reduction in the generation of regulatory T cells. Bach2 represses many genes in T cells, including Prdm1, suggesting that the Bach2-Prdm1 pathway is also important in maintaining the homeostasis of T cells. Furthermore, Bach2 is essential for the function of alveolar macrophages. Therefore, Bach2 orchestrates both acquired and innate immunity at multiple points. Its connection with disease is also reviewed in this report.

The Transcription Factor Bach2 Is Phosphorylated at Multiple Sites in Murine B Cells but a Single Site Prevents Its Nuclear Localization.

The transcription factor Bach2 regulates the immune system at multiple points, including class switch recombination (CSR) in activated B cells and the function of T cells in part by restricting their terminal differentiation. However, the regulation of Bach2 expression and its activity in the immune cells are still unclear. Here, we demonstrated that Bach2 mRNA expression decreased in Pten-deficient primary B cells. Bach2 was phosphorylated in primary B cells, which was increased upon the activation of the B cell receptor by an anti-immunoglobulin M (IgM) antibody or CD40 ligand. Using specific inhibitors of kinases, the phosphorylation of Bach2 in activated B cells was shown to depend on the phosphatidylinositol 3-kinase (PI3K)-Akt-mammalian target of rapamycin (mTOR) pathway. The complex of mTOR and Raptor phosphorylated Bach2 in vitro. We identified multiple new phosphorylation sites of Bach2 by mass spectrometry analysis of epitope-tagged Bach2 expressed in the mature B cell line BAL17. Among the sites identified, serine 535 (Ser-535) was critical for the regulation of Bach2 because a single mutation of Ser-535 abolished cytoplasmic accumulation of Bach2, promoting its nuclear accumulation in pre-B cells, whereas Ser-509 played an auxiliary role. Bach2 repressor activity was enhanced by the Ser-535 mutation in B cells. These results suggest that the PI3K-Akt-mTOR pathway inhibits Bach2 by both repressing its expression and inducing its phosphorylation in B cells.

Epigenetic Regulation of the Blimp-1 Gene (Prdm1) in B Cells Involves Bach2 and Histone Deacetylase 3.

B lymphocyte-induced maturation protein 1 (Blimp-1) encoded by Prdm1 is a master regulator of plasma cell differentiation. The transcription factor Bach2 represses Blimp-1 expression in B cells to stall terminal differentiation, by which it supports reactions such as class switch recombination of the antibody genes. We found that histones H3 and H4 around the Prdm1 intron 5 Maf recognition element were acetylated at higher levels in X63/0 plasma cells expressing Blimp-1 than in BAL17 mature B cells lacking its expression. Conversely, methylation of H3-K9 was lower in X63/0 cells than BAL17 cells. Purification of the Bach2 complex in BAL17 cells revealed its interaction with histone deacetylase 3 (HDAC3), nuclear co-repressors NCoR1 and NCoR2, transducin ß-like 1X-linked (Tbl1x), and RAP1-interacting factor homolog (Rif1). Chromatin immunoprecipitation confirmed the binding of HDAC3 and Rif1 to the Prdm1 locus. Reduction of HDAC3 or NCoR1 expression by RNA interference in B cells resulted in an increased Prdm1 mRNA expression. Bach2 is suggested to cooperate with HDAC3-containing co-repressor complexes in B cells to regulate the stage-specific expression of Prdm1 by writing epigenetic modifications at the Prdm1 locus.

Temporal effects of Notch signaling and potential cooperation with multiple downstream effectors on adenohypophysis cell specification in zebrafish.

The adenohypophysis (AH) consists of six distinct types of hormone-secreting cells. In zebrafish, although proper differentiation of all AH cell types has been shown to require Notch signaling within a period of 14-16 h postfertilization (hpf), the mechanisms underlying this process remain to be elucidated. Herein, we observed using the Notch inhibitor dibenzazepine (DBZ) that Notch signaling also contributed to AH cell specification beyond 16 hpf. Specification of distinct cell types was perturbed by DBZ treatment for different time frames, suggesting that AH cells are specified by Notch-dependent and cell-type-specific mechanisms. We also found that two hes-family genes, her4.1 and hey1, were expressed in the developing AH under the influence of Notch signaling. her4.1 knockdown reduced expression of proopiomelanocortin a (pomca), growth hormone (gh), and prolactin, whereas hey1 was responsible only for gh expression. Simultaneous loss of both Her4.1 and Hey1 produced milder phenotypes than that of DBZ-treated embryos. Moreover, DBZ treatment from 18 hpf led to a significant down-regulation of both gh and pomca genes only when combined with injection of a subthreshold level of her4.1-morpholino. These observations suggest that multiple downstream effectors, including Her4.1 and Hey1, mediate Notch signaling during AH cell specification.

Genetic complementation analysis showed distinct contributions of the N-terminal tail of H2A.Z to epigenetic regulations.

H2A.Z is one of the most evolutionally conserved histone variants. In vertebrates, this histone variant has two isoforms, H2A.Z.1 and H2A.Z.2, each of which is coded by an individual gene. H2A.Z is involved in multiple epigenetic regulations, and in humans, it also has relevance to carcinogenesis. In this study, we used the H2A.Z DKO cells, in which both H2A.Z isoform genes could be inducibly knocked out, for the functional analysis of H2A.Z by a genetic complementation assay, as the first example of its kind in vertebrates. Ectopically expressed wild-type H2A.Z and two N-terminal mutants, a nonacetylable H2A.Z mutant and a chimera in which the N-terminal tail of H2A.Z.1 was replaced with that of the canonical H2A, complemented the mitotic defects of H2A.Z DKO cells similarly, suggesting that both acetylation and distinctive sequence of the N-terminal tail of H2A.Z are not required for mitotic progression. In contrast, each one of these three forms of H2A.Z complemented the transcriptional defects of H2A.Z DKO cells differently. These results suggest that the N-terminal tail of vertebrate H2A.Z makes distinctively different contributions to these epigenetic events. Our results also imply that this genetic complementation system is a novel and useful tool for the functional analysis of H2A.Z.

Iron-heme-Bach1 axis is involved in erythroblast adaptation to iron deficiency.

Iron plays the central role in oxygen transport by erythrocytes as a constituent of heme and hemoglobin. The importance of iron and heme is also to be found in their regulatory roles during erythroblast maturation. The transcription factor Bach1 may be involved in their regulatory roles since it is deactivated by direct binding of heme. To address whether Bach1 is involved in the responses of erythroblasts to iron status, low iron conditions that induced severe iron deficiency in mice were established. Under iron deficiency, extensive gene expression changes and mitophagy disorder were induced during maturation of erythroblasts. Bach1-/- mice showed more severe iron deficiency anemia in the developmental phase of mice and a retarded recovery once iron was replenished when compared with wild-type mice. In the absence of Bach1, the expression of globin genes and Hmox1 (encoding heme oxygenase-1) was de-repressed in erythroblasts under iron deficiency, suggesting that Bach1 represses these genes in erythroblasts under iron deficiency to balance the levels of heme and globin. Moreover, an increase in genome-wide DNA methylation was observed in erythroblasts of Bach1-/- mice under iron deficiency. These findings reveal the principle role of iron as a regulator of gene expression in erythroblast maturation and suggest that the iron-heme-Bach1 axis is important for a proper adaptation of erythroblast to iron deficiency to avoid toxic aggregates of non-heme globin.

Histone H3 Lysine 27 Trimethylation Leads to Loss of Mesendodermal Competence During Gastrulation in Zebrafish Ectodermal Cells.

Previous studies in Xenopus have shown that forced expression of Nodal signaling can change ectodermal cells to a mesodermal fate by the early gastrula stage, suggesting mesodermal competence in early ectoderm cells. This mesodermal competence in ectodermal cells has been shown to be regulated at the level of nucleocytoplasmic localization of Smad2 in Xenopus. However, the regulation of mesodermal competence through epigenetic mechanisms has not been fully elucidated. Here, we used a constitutively active form of zebrafish Smad2 (Smad2ca) to overcome the inhibition of Nodal signaling via the nuclear exclusion of Smad2. While heat-shock-dependent expression of Smad2ca at 5 h post fertilization (hpf) induced ectopic expression of mesendodermal genes in zebrafish ectodermal cells, responsiveness to Smad2ca was lost by 7 hpf. Chromatin immunoprecipitation-quantitative PCR analyses revealed that in ectodermal cells, levels of H3K27me3, but not H3K9me3, at both transcriptional start site (TSS) and 3'-flanking regions of mesendodermal genes at 9 hpf were markedly higher than those at 5 hpf. In contrast to mesendodermal genes, the levels of H3K27me3 at the TSS, but not 3'-flanking regions, of ectodermal genes remained low in ectodermal cells even at 9 hpf. We also found that chemical inhibition of H3K27me3 modification was able to recover the mesendodermal competence in ectodermal cells at 7 hpf, but not at 10 hpf. Taken together, our results suggest that the mesendodermal competence in zebrafish ectodermal cells is restricted by multiple mechanisms, including upregulation of H3K27me3 levels at the TSS of mesendodermal genes during early gastrulation.

Nipbl and mediator cooperatively regulate gene expression to control limb development.

Haploinsufficiency for Nipbl, a cohesin loading protein, causes Cornelia de Lange Syndrome (CdLS), the most common "cohesinopathy". It has been proposed that the effects of Nipbl-haploinsufficiency result from disruption of long-range communication between DNA elements. Here we use zebrafish and mouse models of CdLS to examine how transcriptional changes caused by Nipbl deficiency give rise to limb defects, a common condition in individuals with CdLS. In the zebrafish pectoral fin (forelimb), knockdown of Nipbl expression led to size reductions and patterning defects that were preceded by dysregulated expression of key early limb development genes, including fgfs, shha, hand2 and multiple hox genes. In limb buds of Nipbl-haploinsufficient mice, transcriptome analysis revealed many similar gene expression changes, as well as altered expression of additional classes of genes that play roles in limb development. In both species, the pattern of dysregulation of hox-gene expression depended on genomic location within the Hox clusters. In view of studies suggesting that Nipbl colocalizes with the mediator complex, which facilitates enhancer-promoter communication, we also examined zebrafish deficient for the Med12 Mediator subunit, and found they resembled Nipbl-deficient fish in both morphology and gene expression. Moreover, combined partial reduction of both Nipbl and Med12 had a strongly synergistic effect, consistent with both molecules acting in a common pathway. In addition, three-dimensional fluorescent in situ hybridization revealed that Nipbl and Med12 are required to bring regions containing long-range enhancers into close proximity with the zebrafish hoxda cluster. These data demonstrate a crucial role for Nipbl in limb development, and support the view that its actions on multiple gene pathways result from its influence, together with Mediator, on regulation of long-range chromosomal interactions.

Using mouse and zebrafish models to understand the etiology of developmental defects in Cornelia de Lange Syndrome.

Cornelia de Lange Syndrome (CdLS) is a multisystem birth defects disorder that affects every tissue and organ system in the body. Understanding the factors that contribute to the origins, prevalence, and severity of these developmental defects provides the most direct approach for developing screens and potential treatments for individuals with CdLS. Since the majority of cases of CdLS are caused by haploinsufficiency for NIPBL (Nipped-B-like, which encodes a cohesin-associated protein), we have developed mouse and zebrafish models of CdLS by using molecular genetic tools to create Nipbl-deficient mice and zebrafish (Nipbl(+/-) mice, zebrafish nipbl morphants). Studies of these vertebrate animal models have yielded novel insights into the developmental etiology and genes/gene pathways that contribute to CdLS-associated birth defects, particularly defects of the gut, heart, craniofacial structures, nervous system, and limbs. Studies of these mouse and zebrafish CdLS models have helped clarify how deficiency for NIPBL, a protein that associates with cohesin and other transcriptional regulators in the nucleus, affects processes important to the emergence of the structural and physiological birth defects observed in CdLS: NIPBL exerts chromosome position-specific effects on gene expression; it influences long-range interactions between different regulatory elements of genes; and it regulates combinatorial and synergistic actions of genes in developing tissues. Our current understanding is that CdLS should be considered as not only a cohesinopathy, but also a "transcriptomopathy," that is, a disease whose underlying etiology is the global dysregulation of gene expression throughout the organism. © 2016 Wiley Periodicals, Inc.

Bach2 maintains T cells in a naive state by suppressing effector memory-related genes.

The transcriptional repressor BTB and CNC homology 2 (Bach2) is thought to be mainly expressed in B cells with specific functions such as class switch recombination and somatic hypermutation, but its function in T cells is not known. We found equal Bach2 expression in T cells and analyzed its function using Bach2-deficient (-/-) mice. Although T-cell development was normal, numbers of peripheral naive T cells were decreased, which rapidly produced Th2 cytokines after TCR stimulation. Bach2(-/-) naive T cells highly expressed genes related to effector-memory T cells such as CCR4, ST-2 and Blimp-1. Enhanced expression of these genes induced Bach2(-/-) naive T cells to migrate toward CCR4-ligand and respond to IL33. Forced expression of Bach2 restored the expression of these genes. Using Chromatin Immunoprecipitation (ChIP)-seq analysis, we identified S100 calcium binding protein a, Heme oxigenase 1, and prolyl hydroxylase 3 as Bach2 direct target genes, which are highly expressed in effector-memory T cells. These findings indicate that Bach2 suppresses effector memory-related genes to maintain the naive T-cell state and regulates generation of effector-memory T cells.

Heme binds to an intrinsically disordered region of Bach2 and alters its conformation.

The transcriptional repressor Bach2 regulates humoral and cellular immunity, including antibody class switching. It possesses a basic leucine zipper domain that mediates DNA binding. Heme inhibits the DNA-binding activity of Bach2 in vitro and induces the degradation of Bach2 in B cells. However, the structural basis of the heme-Bach2 interaction has not been identified. Spectroscopic analyses revealed that Bach2(331-520) is the heme-binding domain, as it includes three Cys-Pro motifs known to be important for heme binding. Heme-titration experiments demonstrated the presence of 5- and 6-coordinated heme-binding modes. Circular dichroism measurements indicated that Bach2(331-520) exists mostly in a random-coil conformation. However, dynamic light scattering analyses showed that, upon heme binding to Bach2(331-520), this region becomes denatured at a lower temperature, as compared with unbound Bach2(331-520). In addition, small-angle X-ray scattering and chemical modification analyses revealed that heme binding induces conformational alterations within the unstructured region. A GAL4-based luciferase assay in 293T cells showed that heme alters the protein interactions mediated by Bach2(331-520). These observations suggested that the unstructured region of Bach2 is important for heme binding, and consequently for its functional regulation.

Association between BACH2 expression and clinical prognosis in diffuse large B-cell lymphoma.

BACH2, a B cell-specific transcriptional repressor, plays a significant role in B cell maturation. Despite a number of previous studies, the clinicopathological significance of BACH2 expression in diffuse large B cell lymphoma (DLBCL) remains to be established. The present study was performed to validate the significance of BACH2 expression as a predictor of prognosis in DLBCL. A total of 94 DLBCL cases were included in the present study. All were diagnosed between 2008 and 2011, and thorough clinical and pathological investigations were possible, including immunohistochemical analysis of BACH2. Eighteen cases were selected by positive MYC gene alteration (MYC+ group) according to cytogenetic study. The remaining 76 cases were subclassified into germinal center B cell phenotype (GCB group, 38 cases) or non-GCB phenotype (non-GCB group, 38 cases). There were no significant differences between the two groups with regard to clinical characteristics and outcomes. In the GCB group, 21 cases were judged to have high BACH2 expression, with 19 cases in the non-GCB group. In cases with high BACH2 expression in GCB and non-GCB groups, the 3-year overall survival (OS) rate was significantly shorter than that with low expression (71.7% vs 91.3%, P = 0.0256). In the MYC+ group, 15 cases had high BACH2 expression levels. Although overall the MYC+ group showed short survival time (3-year OS 35.0%), 3 out of 4 cases with low BACH2 expression are alive without disease relapse at the time of publication of this paper. In conclusion, BACH2 expression level is a promising predictor of prognosis for DLBCL.

Bach2 represses plasma cell gene regulatory network in B cells to promote antibody class switch.

Two transcription factors, Pax5 and Blimp-1, form a gene regulatory network (GRN) with a double-negative loop, which defines either B-cell (Pax5 high) or plasma cell (Blimp-1 high) status as a binary switch. However, it is unclear how this B-cell GRN registers class switch DNA recombination (CSR), an event that takes place before the terminal differentiation to plasma cells. In the absence of Bach2 encoding a transcription factor required for CSR, mouse splenic B cells more frequently and rapidly expressed Blimp-1 and differentiated to IgM plasma cells as compared with wild-type cells. Genetic loss of Blimp-1 in Bach2(-/-) B cells was sufficient to restore CSR. These data with mathematical modelling of the GRN indicate that Bach2 achieves a time delay in Blimp-1 induction, which inhibits plasma cell differentiation and promotes CSR (Delay-Driven Diversity model for CSR). Reduction in mature B-cell numbers in Bach2(-/-) mice was not rescued by Blimp-1 ablation, indicating that Bach2 regulates B-cell differentiation and function through Blimp-1-dependent and -independent GRNs.