Dual Mechanism of Rag Gene Repression by c-Myb during Pre-B Cell Proliferation

Abstract

Developing B lymphocytes undergo clonal expansion following successful immunoglobulin heavy chain gene rearrangement. During this proliferative burst, expression of the genes is transiently repressed to prevent the generation of double-stranded DNA (dsDNA) breaks in cycling large pre-B cells. The genes are then reexpressed in small, resting pre-B cells for immunoglobulin light chain gene rearrangement. We previously identified c-Myb as a repressor of transcription during clonal expansion using Abelson murine leukemia virus-transformed B cells. Nevertheless, the molecular mechanisms by which c-Myb achieved precise spatiotemporal repression of expression remained obscure. Here, we identify two mechanisms by which c-Myb represses transcription. First, c-Myb negatively regulates the expression of the activator Foxo1, an activity dependent on M303 in c-Myb's transactivation domain, and likely the recruitment of corepressors to the locus by c-Myb. Second, c-Myb represses transcription directly by occupying the E enhancer and antagonizing Foxo1 binding to a consensus forkhead site in this -regulatory element that we show is crucial for expression in Abelson pre-B cell lines. This work provides important mechanistic insight into how spatiotemporal expression of the genes is tightly controlled during B lymphocyte development to prevent mistimed dsDNA breaks and their deleterious consequences.

Hierarchical assembly and disassembly of a transcriptionally active RAG locus in CD4+CD8+ thymocytes

“…In B cells, the RAG promoters are controlled by proximal, distal, and Erag enhancers distributed upstream of Rag2 (Monroe et al, 1999; Hsu et al, 2003). Studies of the Erag enhancer have revealed FOXO1 as a positive regulator of RAG expression and Gfi1b, Ebf1, and c-Myb as negative regulators (Amin and Schlissel, 2008; Schulz et al, 2012; Timblin and Schlissel, 2013; Lee et al, 2017; Timblin et al, 2017). RAG expression in T cells is controlled by distinct cis-regulatory elements.…”

c-Myb interferes with inflammatory IL1α-NF-κB pathway in breast cancer cells

“…Depending on the MBS position, it can activate transcription of noncoding RNA molecules, thus leading to post-transcriptional repression [48] . Additional regulatory molecules may be essential to facilitate access of the p300 active site [60] or other interacting partners may be involved [61] . Overall, the effect of p300 recruitment to the gene regulatory elements appears to be context-dependent [ 62 , 63 ].…”

The Interplay Between Chromatin Architecture and Lineage-Specific Transcription Factors and the Regulation of Rag Gene Expression

“…In B cells, the deletion of Erag, which is 23 kb upstream of the Rag2 promoter, resulted in a significant reduction in Rag1/2 expression and a partial block at the transition from pro-B to pre-B cells, but did not affect thymocyte development in mice (58). It has been reported that the Erag enhancer is positively regulated by FOXO1 and negatively regulated by Gfi1b, Ebf1, and c-Myb (59)(60)(61)(62). In contrast, an anti-silencer element (ASE), which is 8 kb in length and located 73 kb upstream of the Rag2 promoter, is shown to be required for Rag1/2 expression in DN3 and DP cells but not in B cells (63).…”