Study of Long-Distance Functional Interactions between Su(Hw) Insulators That Can Regulate Enhancer-Promoter Communication in Drosophila melanogaster

Abstract

The Su(Hw) insulator found in the gypsy retrotransposon is the most potent enhancer blocker in Drosophila melanogaster. However, two such insulators in tandem do not prevent enhancer-promoter communication, apparently because of their pairing interaction that results in mutual neutralization. Furthering our studies of the role of insulators in the control of gene expression, here we present a functional analysis of a large set of transgenic constructs with various arrangements of regulatory elements, including two or three insulators. We demonstrate that their interplay can have quite different outcomes depending on the order of and distance between elements. Thus, insulators can interact with each other over considerable distances, across interposed enhancers or promoters and coding sequences, whereby enhancer blocking may be attenuated, cancelled, or restored. Some inferences concerning the possible modes of insulator action are made from collating the new data and the relevant literature, with tentative schemes illustrating the regulatory situations in particular model constructs.

New Properties of Drosophila Fab-7 Insulator

“…The 1.2-kb Fab-7 insulator and its 0.86-kb subfragment block the yellow and white enhancers with similar efficiency: To test the enhancer-blocking activity of the Fab-7 insulators and functional interaction between them, we employed the test system with the yellow and white genes that have been extensively used in insulator studies (Geyer and Corces 1992;Kellum and Schedl 1992;Roseman et al 1993;Muravyova et al 2001;Gruzdeva et al 2005;Savitskaya et al 2006). The yellow gene (designated Y) is required for dark pigmentation of larval and adult cuticle and its derivatives.…”

Insulators form gene loops by interacting with promoters in Drosophila

“…The 3 kb SalI-BamHI fragment containing the yellow gene regulatory region (yr) was cloned into pGEM7 digested with XhoI and BamHI (yr-pGEM7). The 5 kb BamHI-BglII fragment containing the coding region (yc) was inserted in forward orientation into the C⌬ plasmid (Savitskaya et al, 2006) digested with BamHI (C⌬-yc). The XbaI-BamHI fragment containing yr was then cloned from the yr-pGEM7 vector into C⌬-yc digested with XbaI and BamHI (C⌬-y).…”

“…Experiments with transgenic lines suggest that the interaction between insulators can result in the formation of chromatin loops that either block or facilitate long-distance enhancer-promoter communication depending on the nature of the interacting insulators as well as on the distances between all the elements involved (enhancers, insulators and promoters) and their relative 'strength' (Muravyova et al, 2001;Cai and Shen, 2001;Conte et al, 2002;Gruzdeva et al, 2005;Savitskaya et al, 2006;Kyrchanova et al, 2008a). Alternatively, insulator action can be explained by the ability of insulators to form direct contacts with either an enhancer (the decoy model) or a promoter, thereby inactivating them.…”

GATA simple sequence repeats function as enhancer blocker boundaries

“…Much of this non-coding DNA consists of a variety of repetitive DNA including transposable elements (TEs) and simple sequence repeats (SSRs). Certain repetitive sequences have been implicated in chromatin-mediated transcriptional regulation, chromosome organization, imprinting, chromatin domain boundary function and complex nuclear features such as heterochromatin, telomeres and nucleolar organization [1][2][3][4][5][6][7][8] . SSRs, on the other hand, have not been directly implicated in such regulatory roles although this class of DNA has also accumulated in complex genomes, particularly in vertebrates 9,10 .…”