De novo DNA Methyltransferases Dnmt3a and Dnmt3b regulate the onset of Igκ light chain rearrangement during early B‐cell development

Abstract

Immunoglobulin genes V(D)J rearrangement during early lymphopoiesis is a critical process involving sequential recombination of the heavy and light chain loci. A number of transcription factors act together with temporally activated recombinases and chromatin accessibility changes to regulate this complex process. Here, we deleted the de novo DNA methyltransferases Dnmt3a and Dnmt3b in early B cells of conditionally targeted mice, and monitored the process of V(D)J recombination. Dnmt3a and Dnmt3b deletion resulted in precocious recombination of the immunoglobulin κ light chain without impairing the differentiation of mature B cells or overall B-cell development. Ex vivo culture of IL-7 restricted early B-cell progenitors lacking Dnmt3a and Dnmt3b showed precocious Vκ-Jκ rearrangements that are limited to the proximal Vκ genes. Furthermore, B-cell progenitors deficient in Dnmt3a and Dnmt3b showed elevated levels of germline transcripts at the proximal Vκ genes, alterations in methylation patterns at Igκ enhancer sites and increased expression of the transcription factor E2A. Our data suggest that Dnmt3a and Dnmt3b are critical to regulate the onset of Igκ light chain rearrangement during early B-cell development.Keywords: B-cell r DNA methylation r Immunoglobulin r V(D)J rearrangement Additional supporting information may be found in the online version of this article at the publisher's web-site IntroductionThe development of lymphocytes (B or T cells) through differentiation of HSCs into several successive progenitor stages involves highly coordinated events orchestrated by many different genes, which often act at specific stages of cell differentiation. These genes encode lineage-specific transcription factors, growth factors, and chemokines and their receptors [1][2][3][4]. The primary function Correspondence: Prof. Patrick Matthias e-mail: patrick.matthias@fmi.ch of B lymphocyte is to produce high level of antigen-specific antibodies. To generate a very large repertoire of antigen receptors (antibodies) early B cells undergo a somatic recombination process known as V(D)J recombination that assembles variable (V), diversity (D), and joining (J) gene segments of the immunoglobulin (Ig) locus to create a large diversity of antibodies [5,6].The mouse Ig heavy (IgH) and light (IgL) chain (κ or λ) loci become activated during B-cell development in a step-wise manner for V(D)J recombination. Eur. J. Immunol. 2015Immunol. . 45: 2343Immunol. -2355 the small pre-BII cell stage, thereby giving rise to functional Ig genes in successive stages of B-cell development [6,7]. The V(D)J recombinase proteins Rag-1 and Rag-2 are responsible for the induction of cleavage and recombination at conserved flanking regions (recombination signal sequences, RSS) of the V, D, and J gene segments within the Ig loci. Accessibility of RSS sites at the Ig locus for Rag-1 and Rag-2 mediated synapsis and cleavage is dependent on chromatin structure and epigenetic marks [7][8][9]. DNA methylation is a covalent modification of the genomic DNA...

Tet2 and Tet3 cooperate with B-lineage transcription factors to regulate DNA modification and chromatin accessibility

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Defining B Cell Chromatin: Lessons from EBF1

“…Apart from histone modifications, DNA methylation also plays a role in Ig locus recombination. Here, knockdown of the DNA methyltransferases Dnmt3a and Dnmt3b or the DNA demethylases Tet2 and Tet3 results in precocious or strongly impaired rearrangement, respectively [56,57,92]. EBF1 contributes to V(D)J recombination mainly indirectly via the activation of Pax5 [93].…”

TET Methylcytosine Oxidases in T Cell and B Cell Development and Function

“…B cells undergo dynamic changes in DNA modification status during their development, with an estimated 30% of all CpGs exhibiting changes at distinct genomic regions depending on developmental stage ( 104 ). Ablation of the gene encoding the maintenance methyltransferase Dnmt1 completely halted early B cell development ( 105 ), but the de novo methyltransferases Dnmt3a and Dnmt3b were dispensable for B cell development in conditional Mb1Cre Dnmt3a/b-deficient mice, although the B cell receptor repertoire was skewed toward increased usage of proximal Vκ genes ( 106 ). Thus, maintenance of global DNA methylation is essential for B cell development, while de novo methylation is important for proper immunoglobulin (Ig) gene rearrangement.…”