Recently, we demonstrated that the lineage-restricted BROMO and WD40 domain containing epigenetic reader BRWD1 opens Ig? and enables assembly of RAG proteins at J?. We now report that BRWD1 has a much broader role in late B cell development. At distances of up to 5 mb from sites of binding, BRWD1 reshaped chromatin landscape by closing enhancers of genes expressed early in B cell development and opening those of genes expressed in late stages. In the absence of BRWD1, over 7000 genes were aberrantly expressed. While many of these genes were normally expressed in earlier developmental stages, many were also part of the germinal center (GC) transcription program. BRWD1 is first expressed in small pre-B cells. However, Brwd1 expression was much higher in nave follicular (FO) and highest in day 12 GC B cells. In light zone (LZ) cells, BRWD1 is expressed in Myc- cells, a transcription factor repressed by BRWD1. Finally, analysis of genes differentially expressed in GC dark zone (DZ) and LZ, compared to those differentially regulated by BRWD1 in small pre-B cells, predicted that BRWD1 represses the DZ program and induces the LZ program. These data suggest similarities between genetic programs that separate proliferation and Ig? recombination in development and those that segregate proliferation from selection in GCs. Based on these findings, we hypothesize that BRWD1 is required for maintaining FO cell identity. Furthermore, we hypothesize that, by repressing DZ and inducing LZ genetic programs, BRWD1 is critical for GC-dependent humoral immunity. These hypotheses will be tested in the following Specific Aims:
Aim 1 : Derive a conditional allele of Brwd1.
Aim 2. Determine role of BRWD1 in maintaining follicular B cell identity.
Aim 3. Determine the role of BRWD1 in adaptive immunity.
In germinal centers, B cells rapidly oscillate between states of differentiation and selection that ensure expansion of B cells secreting protective antibodies against infection. In this grant proposal, we will test a novel model of epigenetic regulation controlling differentiation and selection of B cell developmental states.
