Tissue-Specific Regulation of Chromatin Insulator Function

Leah H Matzat,Nellie Moshkovich,Elissa P Lei,Ryan K Dale,Gregory P Copenhaver

doi:10.1371/journal.pgen.1003069

Abstract

Chromatin insulators organize the genome into distinct transcriptional domains and contribute to cell type–specific chromatin organization. However, factors regulating tissue-specific insulator function have not yet been discovered. Here we identify the RNA recognition motif-containing protein Shep as a direct interactor of two individual components of the gypsy insulator complex in Drosophila. Mutation of shep improves gypsy-dependent enhancer blocking, indicating a role as a negative regulator of insulator activity. Unlike ubiquitously expressed core gypsy insulator proteins, Shep is highly expressed in the central nervous system (CNS) with lower expression in other tissues. We developed a novel, quantitative tissue-specific barrier assay to demonstrate that Shep functions as a negative regulator of insulator activity in the CNS but not in muscle tissue. Additionally, mutation of shep alters insulator complex nuclear localization in the CNS but has no effect in other tissues. Consistent with negative regulatory activity, ChIP–seq analysis of Shep in a CNS-derived cell line indicates substantial genome-wide colocalization with a single gypsy insulator component but limited overlap with intact insulator complexes. Taken together, these data reveal a novel, tissue-specific mode of regulation of a chromatin insulator.

Highlights

Chromatin insulators are DNA-protein complexes that influence eukaryotic gene expression by organizing the genome into distinct transcriptional domains
The shep gene was named based on its identification in a gravitaxis screen [33] and is predicted computationally and suggested by EST data to produce four different protein isoforms with distinct N-terminal domains that share a mostly common C-terminal region bearing two highly conserved, tandemly arranged RNA recognition motifs (RRMs; Figure 1A)
We found that insulator phenotypes and synthetic lethality remained the same or insulator function was slightly increased compared to homozygous P-elements, except when shepBG00836 and shepd05714 are transheterozygous with Df(3L)Exel6104 (Table 1)

Summary

Introduction

Chromatin insulators are DNA-protein complexes that influence eukaryotic gene expression by organizing the genome into distinct transcriptional domains. Chromatin insulators are thought to exert effects on gene expression by constraining the topology of chromatin and facilitating the formation of intra- and inter-chromosomal looping (for review, see [2]). These higher order interactions can vary between cell types, thereby facilitating tissue-specific transcriptional output. In addition to Su(Hw), the core gypsy insulator complex contains Centrosomal protein 190 (CP190), which harbors a zinc finger domain, and the nonDNA-binding protein, Modifier of mdg4 2.2 (Mod(mdg4)2.2). Both CP190 and Mod(mdg4)2.2 contain broad complex, tramtrack, bric-a-brac (BTB) dimerization domains that have been suggested to mediate insulator-insulator interactions and facilitate the formation of long range insulator-mediated loops along the chromatin fiber [4,8]

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Results

Conclusion