Groucho Oligomerization Is Required for Repression In Vivo

Abstract

Drosophila Groucho (Gro) is a member of a family of metazoan corepressors with widespread roles in development. Previous studies indicated that a conserved domain in Gro, termed the Q domain, was required for repression in cultured cells and mediated homotetramerization. Evidence presented here suggests that the Q domain contains two coiled-coil motifs required for oligomerization and repression in vivo. Mutagenesis of the putative hydrophobic faces of these motifs, but not of the hydrophilic faces, prevents the formation of both tetramers and higher order oligomers. Mutagenesis of the hydrophobic faces of both coiled-coil motifs in the context of a Gal4-Gro fusion protein prevents repression of a Gal4-responsive reporter in S2 cells, while mutagenesis of a single motif weakens repression. The finding that the repression directed by the single mutants depends on endogenous wild-type Gro further supports the idea that oligomerization plays a role in repression. Overexpression in the fly of forms of Gro able to oligomerize, but not of a form of Gro unable to oligomerize, results in developmental defects and ectopic repression of Gro target genes in the wing disk. Although the function of several corepressors is suspected to involve oligomerization, these studies represent one of the first direct links between corepressor oligomerization and repression in vivo.

A role of Pygopus as an anti-repressor in facilitating Wnt-dependent transcription

“…Our finding that normal nuclear Armadillo accumulation is restored in gro pygo double-mutants (Fig. 5) is consistent with the idea that Groucho may hinder the access of Armadillo to dTCF target genes by recruiting histone deacetylases (11,12), which results in a compact chromatin structure (21)(22)(23). Loss of Groucho from dTCF target genes might lead to localized chromatin remodeling and histone acetylation (47), which may thus allow Armadillounassisted (i.e., Pygo-independent) access.…”

“…We tested whether this lesion would affect the binding of mutant Groucho protein (called MB5) to dTCF by conducting pull-down assays between bacterially expressed WT Groucho (GST-Gro) and in vitro translated dTCF fragments. As expected (4,5,7,11), Groucho binds to the N-terminal half of dTCF (dTCF1-350), but not to its C-terminal half (dTCF347-750; Fig. 1B).…”

“…Germ-line clones were generated by standard FRT-based methods as described (11,15,24), and homozygous mutant embryos were identified using GFP balancers (for cuticle preparations) or twi-lacZ balancers (for immunohistochemistry). To generate somatic clones in the wing disc, a well characterized driver line vgBEGal4 UAS-flp was used (41).…”

“…These associate with different families of DNA-bound proteins (e.g., the basic helix-loop-helix factor Hairy) and recruit histone deacetylases to silence chromatin locally (9,10). Groucho binds to TCF factors through the Q domain in its N terminus (4,5,7,11). The Q domain also undergoes homo-tetramerization, which is important (11) although not obligatory for Groucho's repressive activity in vivo (12).…”

Crosstalk between the EGFR and other signalling pathways at the level of the global transcriptional corepressor Groucho/TLE

“…Clearly, homo-oligomerisation is a prerequisite for Gro/TLE repressor activity and, further, Gro/TLEs physically bind both histones and histone deacetylases (specifically, Rpd3/HDAC1) (e.g. Chen et al, 1999;Song et al, 2004). Collectively, these observations have led to the following model: Gro is initially tethered to specific gene regulatory regions via its associations with distinct DNA-bound repressors.…”