A Comprehensive Map of Insulator Elements for the Drosophila Genome

Abstract

Insulators are DNA sequences that control the interactions among genomic regulatory elements and act as chromatin boundaries. A thorough understanding of their location and function is necessary to address the complexities of metazoan gene regulation. We studied by ChIP–chip the genome-wide binding sites of 6 insulator-associated proteins—dCTCF, CP190, BEAF-32, Su(Hw), Mod(mdg4), and GAF—to obtain the first comprehensive map of insulator elements in Drosophila embryos. We identify over 14,000 putative insulators, including all classically defined insulators. We find two major classes of insulators defined by dCTCF/CP190/BEAF-32 and Su(Hw), respectively. Distributional analyses of insulators revealed that particular sub-classes of insulator elements are excluded between cis-regulatory elements and their target promoters; divide differentially expressed, alternative, and divergent promoters; act as chromatin boundaries; are associated with chromosomal breakpoints among species; and are embedded within active chromatin domains. Together, these results provide a map demarcating the boundaries of gene regulatory units and a framework for understanding insulator function during the development and evolution of Drosophila.

The chromatin insulator CTCF and the emergence of metazoan diversity

“…Conservation of CTCF-binding sites, for example, is not restricted to Hox complexes, either in vertebrates (30) or in Drosophila (SI Appendix, Fig. S9) (65). Thus, other genomic loci might also have benefited from the emergence of CTCF.…”

Dynamic changes in the genomic localization of DNA replication-related element binding factor during the cell cycle

“…[32][33][34][35] Analysis of the genome-wide distribution of these proteins has given some insights into the issue of whether the different Drosophila insulators play distinct or overlapping roles in genome organization and function. [36][37][38] For example, BEAF-32 is found more often close to gene promoters, where it appears to confer independent regulation to closely adjacent, divergently transcribed genes. 39 On the other hand, Su(Hw) tends to be present in intergenic regions next to genes transcribed at low levels.…”

A conserved gene cluster as a putative functional unit in insect innate immunity

“…Through inter-insulator contacts, boundary elements can create chromatin loops and so provide the opportunity for concerted regulation inside this domain [15]. Indeed, ''Class I" insulators -identified by the binding of BEAF-32, CP190, or CTCF in Drosophila embryos -act as chromatin boundaries and separate differentially expressed genes [16]. In humans, chromosomal domains where the insulator protein CTCF is depleted tend to contain clusters of coordinately regulated genes; moreover, pairs of consecutive CTCF-binding sites often delimit clusters of functionally and/or phylogenetically related genes [17].…”

“…In the Nimrod region, a domain delimited by two sequential Class I insulators [16] considerably overlaps with the SpellmanRubin co-expression block and the Mezey-cluster (Fig. 1B).…”

“…Rectangles in the first lane represent the genes in their chromosomal order, colour-coded as in (A). Data presented in other lanes: conserved synthetic blocks across 12 Drosophila species [7]; clusters of co-expression [12]; clusters of genes with coordinated evolution in Drosophila species [13], overlapping merged windows obtained with different window sizes were fused; Class I insulator sites [16], the sector between two consecutive insulators overlapping the Nimrod gene cluster is highlighted; Polycomb protein binding domains [18]; and regions of Eve transcription factor binding-eve BRICK, the largest BRICK overlapping the cluster is shown [19]. chromosomal distribution of the chromatin proteins -frequently modulated by boundary elements -might also indicate similar regulatory mechanisms operating on the corresponding genes.…”