Opposing effects of SWI/SNF and Mi-2/NuRD chromatin remodeling complexes on epigenetic reprogramming by EBF and Pax5

Abstract

Transcriptionally silent genes are maintained in inaccessible chromatin. Accessibility of these genes requires their modification by chromatin remodeling complexes (CRCs), which are recruited to promoters by sequence-specific DNA-binding proteins. Early B-cell factor (EBF), which is crucial for B-cell lineage specification, reprograms mb-1 (Ig-␣) promoters by increasing chromatin accessibility and initiating the loss of DNA methylation. In turn, this facilitates promoter activation by Pax5. Here, we investigated the roles of ATP-dependent CRCs in these mechanisms. Fusion of EBF and Pax5 with the ligand-binding domain of ER␣ allowed for 4-hydroxytamoxifen-dependent, synergistic activation of mb-1 transcription in plasmacytoma cells. Knockdown of the SWI/SNF ATPases Brg1 and Brm inhibited transcriptional activation by EBF:ER and Pax5:ER. In contrast, knock-down of the Mi-2/NuRD complex subunit Mi-2␤ greatly enhanced chromatin accessibility and mb-1 transcription in response to the activators. The reduction of Mi-2␤ also propagated DNA demethylation in response to EBF:ER and Pax5:ER, resulting in fully unmethylated mb-1 promoters. In EBF-or EBF/Pax5-deficient fetal liver cells, both EBF and Pax5 were required for efficient demethylation of mb-1 promoters. Together, our data suggest that Mi-2/NuRD is important for the maintenance of hypermethylated chromatin in B cells. We conclude that SWI/SNF and Mi-2/NuRD function in opposition to enable or limit the reprogramming of genes by EBF and Pax5 during B-cell development.DNA methylation ͉ mb-1 promoter ͉ Cd19 promoter ͉ chromatin accessibility T he development of B cells from progenitor cells in the bone marrow is controlled by a network of transcriptional regulators (reviewed in 1). Early B-cell Factor (EBF; also known as EBF1/O/E-1/COE1) plays an integral role in this network and has been implicated as a major determinant of the B-cell fate (2, 3). In the absence of EBF, B-cell development is arrested at an early progenitor stage (3, 4). EBF is essential for the rearrangement and expression of Ig (Ig) genes in B cells and is required for expression of the B-cell commitment factor Pax5. EBF and Pax5 synergistically activate transcription of B cell-specific genes including mb-1 (Cd79a), which encodes the Ig-␣ subunit of the pre-B and B-cell receptors (5). We have proposed that EBF functions as a 'pioneer' factor by controlling the epigenetic states of its target genes (6). In response to EBF, the accessibility of mb-1 promoter chromatin is increased, while DNA methylation of the promoter is decreased.Specific biochemical interactions necessary for the pioneer functions of EBF have not been identified. However, transitions between active and inactive states of chromatin can be mediated by the recruitment of chromatin-remodeling complexes (CRCs) by transcription factors (7). CRCs serve as 'molecular motors' that mediate changes in the relative positions of nucleosomes. In this regard, CRCs of the mammalian SWI/SNF (related to yeast switch/sucrose non-Fermenter) subfamily ...

Bivalent recognition of nucleosomes by the tandem PHD fingers of the CHD4 ATPase is required for CHD4-mediated repression

“…The display of mIgM on μM.2 murine plasmacytoma cells is proportional to mb-1 transcript abundance (26). In μM.2 cells, mb-1 transcription is activated approximately 80-fold by the Early B cell factor 1 (EBF1) and Paired box protein 5 (Pax5) transcription factors but is restrained by CHD4/NuRD CRCs (27). In this context, depletion of endogenous CHD4 by shRNA increases activation of mb-1 promoters significantly by enhancing chromatin accessibility and demethylation of mb-1 promoter CpGs (27).…”

“…The μM.2 and μM.2 EBF:ER stably transfected cell lines were cultured as described previously (27). Generation of retroviruses and infection of cells were performed as described (26) with the following exceptions: For generation of CHD4 wild type and mutant retroviruses, 67.2 μg retroviral plasmid DNA and 56 μL Lipofectamine™ 2000 (Invitrogen) were used to transfect 60-80% confluent 100-mm dishes of ΦNX cells.…”

Mechanisms Controlling Hematopoiesis

“…In contrast to Dnmt1, Dnmt3a/3b deficiency affected only the long-term reconstitution ability of HSC, but not their differentiation into committed progenitors (Tadokoro et al, 2007). Nevertheless, the molecular mechanisms mediating DNA methylation and demethylation during hematopoietic development have not been deciphered, although chromatin-remodeling factors as well as Polycomb group/Dnmt3a/3b complexes recruited by transcription factors were supposed to be involved (Gao et al, 2009;Kirillov et al, 1996;Vire et al, 2006).…”

“…For example, Ikaros, a lymphoid-specific transcription factor crucial for the commitment of LMPP into CLP can recruit Mi2/NuRD complexes in order to repress genes Koipally et al, 1999;Sridharan & Smale, 2007). Whereas, EBF1 and E2A are involved in the recruitment of the SWI/SNF complex to the upstream enhancer of the CD19 locus as well as to the CD79a promoter region facilitating the transcriptional activation of these B cell-specific genes (Gao et al, 2009;Walter et al, 2008).…”

Interaction of CCR4–NOT with EBF1 regulates gene-specific transcription and mRNA stability in B lymphopoiesis

“…In particular, the binding of EBF1 to chromatin has been shown to correlate with dimethylation of Lys4 of histone H3 (Treiber et al 2010b). In addition, the function of EBF1 has been associated with SWI/SNF-dependent remodeling of chromatin and DNA demethylation of some promoters (Maier et al 2004;Gao et al 2009;Boller et al 2016). However, no physical interaction between EBF1 and SWI/SNF proteins has yet been demonstrated.…”