Abstract
Chromatin insulators/boundary elements share the ability to insulate a transgene from its chromosomal context by blocking promiscuous enhancer–promoter interactions and heterochromatin spreading. Several insulating factors target different DNA consensus sequences, defining distinct subfamilies of insulators. Whether each of these families and factors might possess unique cellular functions is of particular interest. Here, we combined chromatin immunoprecipitations and computational approaches to break down the binding signature of the Drosophila boundary element–associated factor (BEAF) subfamily. We identify a dual-core BEAF binding signature at 1,720 sites genome-wide, defined by five to six BEAF binding motifs bracketing 200 bp AT-rich nuclease-resistant spacers. Dual-cores are tightly linked to hundreds of genes highly enriched in cell-cycle and chromosome organization/segregation annotations. siRNA depletion of BEAF from cells leads to cell-cycle and chromosome segregation defects. Quantitative RT-PCR analyses in BEAF-depleted cells show that BEAF controls the expression of dual core–associated genes, including key cell-cycle and chromosome segregation regulators. beaf mutants that impair its insulating function by preventing proper interactions of BEAF complexes with the dual-cores produce similar effects in embryos. Chromatin immunoprecipitations show that BEAF regulates transcriptional activity by restricting the deposition of methylated histone H3K9 marks in dual-cores. Our results reveal a novel role for BEAF chromatin dual-cores in regulating a distinct set of genes involved in chromosome organization/segregation and the cell cycle.
Highlights
Chromatin insulators/boundary elements (BEs) [1,2] are defined as sequences able to insulate a transgene from its chromosomal context and to block promiscuous enhancer– promoter interactions or heterochromatin spreading [1,3,4,5]
We identify hundreds of boundary element–associated factor (BEAF) dual-cores that are defined by a particular arrangement of DNA sequence motifs bracketing nucleosome binding sequences, and that mark the genomic BEAF binding sites
Since BEAF acts by restricting the deposition of repressing epigenetic histone marks, which affects the accessibility of chromatin, its depletion affects the expression of cell-cycle genes
Summary
Chromatin insulators/boundary elements (BEs) [1,2] are defined as sequences able to insulate a transgene from its chromosomal context and to block promiscuous enhancer– promoter interactions or heterochromatin spreading [1,3,4,5]. These elements are thought to subdivide the genome into functional chromosome domains, through their ability to cluster DNA loops [1,2] and to control the deposition of histone epigenetic marks [6,7,8] to regulate chromatin accessibility for gene expression [9,10,11,12,13]. Whether each of these factors and subfamily of insulators might possess distinct cellular functions is of particular interest
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