Selective interactions between diverse STEs organize the ANT-C Hox cluster

Mo Li,Zhibo Ma,Carly R Duffy,Derrick C Lane,Sapna K Patel,Haini N Cai,Sharmila Roy

doi:10.1038/s41598-018-33588-4

Abstract

The three-dimensional organization of the eukaryotic genome is important for its structure and function. Recent studies indicate that hierarchies of chromatin loops underlie important aspects of both genomic organization and gene regulation. Looping between insulator or boundary elements interferes with enhancer-promoter communications and limits the spread active or repressive organized chromatin. We have used the SF1 insulator in the Drosophila Antennapedia homeotic gene complex (ANT-C) as a model to study the mechanism and regulation of chromatin looping events. We reported previously that SF1 tethers a transient chromatin loop in the early embryo that insulates the Hox gene Sex comb reduce from the neighbor non-Hox gene fushi tarazu for their independent regulation. To further probe the functional range and connectivity of SF1, we used high-resolution chromosomal conformation capture (3C) to search for SF1 looping partners across ANT-C. We report here the identification of three distal SF1 Tether Elements (STEs) located in the labial, Deformed and Antennapedia Hox gene regions, extending the range of SF1 looping network to the entire complex. These novel STEs are bound by four different combinations of insulator proteins and exhibit distinct behaviors in enhancer block, enhancer-bypass and boundary functions. Significantly, the six STEs we identified so far map to all but one of the major boundaries between repressive and active histone domains, underlining the functional relevance of these long-range chromatin loops in organizing the Hox complex. Importantly, SF1 selectively captured with only 5 STEs out of ~20 sites that display similar insulator binding profiles, indicating that presence of insulator proteins alone is not sufficient to determine looping events. These findings suggest that selective interaction among diverse STE insulators organize the Drosophila Hox genes in the 3D nuclear space.

Highlights

The three-dimensional (3D) organization of the genome is critical for its function including transcription regulation[1,2,3,4,5,6,7,8]
We have previously identified a hub of chromatin loops anchored by the SF1 insulator in the Drosophila Antennapedia homeotic complex (ANT-C, Fig. 1A)[22]
We tested SF1 capture of 39 EcoRI fragments using high-resolution chromosomal conformation capture (3C) in 0–20 hour old embryos (Fig. 1A, see Methods)[20,51,52,53,54]. These elements cover most of the non-transcribed regions in Antennapedia homeotic gene complex (ANT-C) outside the Scr-ftz interval, which was described previously[22]

Summary

Introduction

The three-dimensional (3D) organization of the genome is critical for its function including transcription regulation[1,2,3,4,5,6,7,8]. The Drosophila Homeotic/Hox genes are activated in a tissue-specific pattern by numerous enhancers during early embryonic development Their transcriptional status are maintained by epigenetic mechanisms mediated by the Polycomb Group (PcG) and Trithorax Group (Trx-G) complexes in late development[39,40,41,42,43]. SF1 and STEs collocalize with all but one of the major domain boundaries between repressive chromatin domains around Hox genes and the less repressive domains around non Hox domains, underscoring the functional relevance of these looping events in organizing the ANT-C Hox complex in Drosophila embryos

Methods

Results

Conclusion