Groucho corepressor functions as a cofactor for the Knirps short-range transcriptional repressor

Abstract

Despite the pervasive roles for repressors in transcriptional control, the range of action of these proteins on cis regulatory elements remains poorly understood. Knirps has essential roles in patterning the Drosophila embryo by means of short-range repression, an activity that is essential for proper regulation of complex transcriptional control elements. Short-range repressors function in a local fashion to interfere with the activity of activators or basal promoters within Ϸ100 bp. In contrast, long-range repressors such as Hairy act over distances >1 kb. The functional distinction between these two classes of repressors has been suggested to stem from the differential recruitment of the CtBP corepressor to short-range repressors and Groucho to long-range repressors. Contrary to this differential recruitment model, we report that Groucho is a functional part of the Knirps short-range repression complex. The corepressor interaction is mediated via an eh-1 like motif present in the N terminus and a conserved region present in the central portion of Knirps. We also show that this interaction is important for the CtBP-independent repression activity of Knirps and is required for regulation of even-skipped. Our study uncovers a previously uncharacterized interaction between proteins previously thought to function in distinct repression pathways, and indicates that the Groucho corepressor can be differentially harnessed to execute short-and long-range repression. S hort-range transcriptional repression has a central role in development, and perhaps nowhere have the molecular workings of eukaryotic developmental gene networks been more extensively analyzed than in the Drosophila blastoderm embryo. Here, both transcriptional activators and repressors transduce temporal and spatial information into characteristic patterns of gene expression essential for development. Repressors have key parts in this process, evidenced by the central position in the hierarchy of genes such as hairy, giant, knirps (kni), and Kruppel, all of which function as dedicated repressors. Transcriptional repressors have been characterized based on their range of action; short-range repressors such as Knirps work over distances of Ͻ100 bp to quench activators or basal promoters (1). In contrast, long-range repressors such as Hairy function over distances of 1 kb to silence their target genes in a process suggested to involve extensive spreading of a recruited corepressor, Groucho (2, 3).Seminal work in this area by the Levine laboratory has prompted the suggestion that the functional differences in the range of action of the two classes of repressors reflect the recruiting of distinct corepressors (4). Short-range repressors such as Knirps, Kruppel, Giant, and Snail associate with the C-terminal binding protein (CtBP) corepressor, whereas Groucho is implicated in mediating long-range repression by Hairy (4, 5). Both evolutionarily conserved corepressors have been linked to chromatin-modifying enzymes, and each associates with sequence-specific D...

Long- and Short-Range Transcriptional Repressors Induce Distinct Chromatin States on Repressed Genes

“…However, the mechanisms by which these proteins function have been poorly understood. With the recent demonstration that transcriptional factors considered to be short- and long-range repressors utilize shared cofactors, namely CtBP and Groucho, there has been a question of whether long-range repression is actually functionally distinct from short-range repression [6]. Our study provides evidence that the chromatin states associated with long- and short-range repressors are distinct in several ways.…”

The Snail Repressor Inhibits Release, Not Elongation, of Paused Pol II in the Drosophila Embryo

“…Snail and Knirps might employ distinctive modes of transcriptional repression. Snail recruits the “short range” corepressor, CtBP [29] , while Knirps recruits either CtBP or the “long-range” corepressor, Groucho [30] . When bound to certain cis-regulatory elements within the eve locus, Knirps recruits Groucho, which might propagate a repressive chromatin structure.…”

The Tbx20 homolog Midline represses wingless in conjunction with Groucho during the maintenance of segment polarity

“…Mouse Tbx18 and Tbx15 have been shown to bind Gro and have Gro dependent transcriptional repressor functions (Farin et al, 2007). It has also been noted for other eh1 containing proteins that eh1 F residue substitutions or deletion of the entire eh1 domain reduces but does not eliminate Gro binding (Andrioli et al, 2004;Farin et al, 2007;Goldstein et al, 2005;Hittinger and Carroll, 2008;Payankaulam and Arnosti, 2009;Smith and Jaynes, 1996;Tolkunova et al, 1998). Several lines of evidence suggest that the defect in mid F41E mutant is specific to Gro binding function and does not cause a general disruption of Mid function.…”

“…We also tested GST-Mid NT Á F41E , a construct with a F-E amino acid substitution in the eh1 domain. Similar mutations in other transcription factors have been demonstrated to reduce Gro binding (Goldstein et al, 2005;Payankaulam and Arnosti, 2009;Smith and Jaynes, 1996;Tolkunova et al, 1998). In pull-down assays, equal amounts of the two proteins were loaded onto GST-beads.…”