Chromatin and epigenetic memory

Eunice Kennedy Shriver National Institute Of Child Health & Human Development

Investigators

Linked publications & trials

Abstract

1. BRD4 directs HSC self renewal: Context dependent significance in peripheral immune cells. As part of efforts to understand the role of BRD4 hematopoietic cells and innate immunity, we examined mice with Brd4 conditional KO. We have shown that Brd4f/f,Vav-Cre mice, lacking Brd4 in the HSCs die in utero due to impaired blood cell production, Analysis of fetal liver cells revealed a striking paucity of LT-HSCs, the source of all immune cells and red blood cells. The MPP, progenitor immediately downstream of HSCs were also scarce. As a result, cells of myeloid and lymphoid lineages were hardly present in the Brd4 KO fetus. These results demonstrate that BRD4 is essential for hematopoiesis in the fetal liver, presumably because BRD4 is required for HSCs self renewal and progenitor differentiation capacity. On the other hand, Bd4f/f LysM-Cre mice, lacking BRD4 in macrophages showed milder effects, in that the Brd4 deletion did not completely inhibit development of macrophages with the Cd11b and F4/80 markers, although Brd4 KO macrophages were deficient in IL-4 induced proliferation. Furthermore Brd4 KO macrophages were only partially defective in LPS induced inflammation. ChIP-seq analysis revealed that BRD4 bound to super-enhancers enriched with H3K27ac. LPS rapidly changed the location of super-enhancers. Interestingly, many of the super-enhancers targeting inflammatory genes were still present in Brd4 KO macrophages. Our results pointed multiple compensatory mechanisms that cover the loss of BRD4 sparing the ability to elicit inflammation, a process necessary for innate immunity. Similar, context dependent roles of Brd4 were observed in our collaboration on thymic T cell development and adipocytes. 2. BRD4 marks cell cycle genes and orchestrates mitotic cell division Cell cycle progression is driven by sequential activation of transcription factors, kinases and other effectors 69. BRD4 promotes growth of many cancers. Inhibitors that interfere binding of BRD4 with acetylated histones or inhibitors that degrade BRD4 collectively called BETi are shown to arrest growth of many types of cancers, particularly effective for leukemia. However, whether BRD4 promotes proliferation of normal cells has remained inconclusive. Also, the mechanisms and pathways through which BRD4 regulate cell growth has been unclear, even for cancers. By testing Brd4 KO mouse embryonic fibroblasts, we found that BRD4 is critically required for cell cycle progression, not only in G1-to-S transition, but S-to- G2 and G2-to-M progression. Transcriptome analysis revealed that genes required for S phase and G2/M progression were markedly downregulated in Brd4KO cells, including multiple histone genes at S phase, and G2/M master regulators, FOXM1 and ATM/ATR. FOXM1 is a transcription factor required for activation of many G2/M genes. ATM/ATR are kinases previously known to be involved in DNA damage recognition/repair. They are recently shown to have a role in G2/M transition to lead mitotic cell division. By ChIP-seq analyses we found that BRD4 constitutively occupies numerous cell cycle genes active at not only G0/G1, but S and G2/M. Our results indicate that BRD sets an epigenome structure, ensuring cell cycle progressions in the subsequent generations. Our imaging analysis found that t Brd4 KO cells undergo a catastrophic failure in mitosis. Nuclear materials in Brd4 KO cells disintegrated without assembling at the mitotic plate, and leading to apoptosis Together, our study demonstrates that BRD4 orchestrates the entire program of cell cycle progression for both normal and cancer cells.