Insulator speckles associated with long-distance chromatin contacts

Abstract

Nuclear foci of chromatin binding factors are, in many cases, discussed as sites of long-range chromatin interaction in the three-dimensional nuclear space. Insulator binding proteins have been shown to aggregate into insulator bodies, which are large structures not involved in insulation; however, the more diffusely distributed insulator speckles have not been analysed in this respect. Furthermore, insulator binding proteins have been shown to drive binding sites for Polycomb group proteins into Polycomb bodies. Here we find that insulator speckles, marked by the insulator binding protein dCTCF, and Polycomb bodies show differential association with the insulator protein CP190. They differ in number and three-dimensional location with only 26% of the Polycomb bodies overlapping with CP190. By using fluorescence in situ hybridization (FISH) probes to identify long-range interaction (kissing) of the Hox gene clusters Antennapedia complex (ANT-C) and Bithorax complex (BX-C), we found the frequency of interaction to be very low. However, these rare kissing events were associated with insulator speckles at a significantly shorter distance and an increased speckle number. This suggests that insulator speckles are associated with long-distance interaction.

Multiple interactions are involved in a highly specific association of the Mod(mdg4)-67.2 isoform with the Su(Hw) sites inDrosophila

“…Previously, we and others have shown that CP190, Mod(mdg4)-67.2 and Su(Hw) are co-localized in the nuclear speckles, named also the insulator bodies [ 51 , 70 , 77 – 80 ]. According to the current model [ 70 , 78 ], the insulator bodies help to form protein complexes that subsequently bind to the regulatory elements such as insulators and promoters.…”

Role of Su(Hw) zinc finger 10 and interaction with CP190 and Mod(mdg4) proteins in recruiting the Su(Hw) complex to chromatin sites in Drosophila

“…The genome-wide studies have shown that binding sites for the Su(Hw), Mod(mdg4)-67.2, and CP190 proteins could be grouped into two main classes which are characterized by the binding of either Su(Hw) alone (SBS-O) or of all the three proteins (SBS-CM) [ 35 , 56 , 57 ]. In the interphase nucleus the Su(Hw), CP190, and Mod(mdg4)-67.2 proteins colocalize in speckles, named insulator bodies [ 43 , 58 – 62 ]. It was speculated [ 60 ] that protein complexes are formed in the insulator bodies before binding to the chromatin.…”

Drosophila CP190- and dCTCF-mediated enhancer blocking is augmented by SUMOylation

“…Nuclear foci of insulator proteins have been identified in different quality and quantity, depending on stress conditions of the analyzed cells as well as on microscopic resolution. Recently, we have refined the analysis of insulator speckles [ 43 ] and found more than 100 such foci, when analyzed by structured illumination microscopy (SIM) or 20 or less foci, when detected by confocal laser scanning microscopy (CLSM). Furthermore, we determined that insulator speckles are associated with long-distance chromatin contacts.…”