Non-consensus GLI binding sites in Hedgehog target gene regulation

Abstract

BackgroundThe GLI transcription factors, mediators of the hedgehog signal bind with high affinity to the consensus sequence GACCACCCA. The affinity of variant single substitutions in GLI binding sites has been measured systematically, but the affinities of the variant binding sites appears low compared to the frequency of occurrence of variant sites in known GLI target gene promoters.ResultsWe quantified transcriptional activation by GLI using PTCH1 promoter based luciferase reporters containing all single substitutions of the GLI consensus binding site. As expected variants with very low affinity did not activate the reporter. Many lower affinity binding sequences are, however, functional in the presence of moderate GLI concentration. Using two natural non-consensus GLI site promoters we showed that substitution of the variant sequences by consensus leads to comparable activity.ConclusionsVariant GLI binding sites with relatively low affinity can within natural promoters lead to strong transcriptional activation. This may facilitate the identification of additional direct GLI target genes.

Synergistic cross‐talk of hedgehog and interleukin‐6 signaling drives growth of basal cell carcinoma

“…Interestingly and as depicted in Figure 3 c , all three HH‐IL6 target genes harbor putative GLI and STAT3 binding sites in close proximity as predicted by bioinformatics analysis. By contrast, in silico analysis of the PTCH promoter revealed GLI binding sites 30, but failed to identify STAT3 binding sites (Supporting Information, Fig. S4 c ), consistent with PTCH expression being insensitive to IL6 signaling.…”

“…In silico prediction of putative GLI binding sites was done using the D‐Light Software29 (genome sequence GRCh37/hg19) trained with the GLI binding site matrix according to Winklmayr et al30 The ENCyclopedia Of DNA Elements (ENCODE) Project31 was used to check for STAT3 binding regions. Luciferase reporter assays and site directed mutagenesis were carried out as described previously 23.…”

Integration of Hedgehog and BMP signalling by the engrailed2a gene in the zebrafish myotome

“…Consistent with this, synthetic forms of Gli proteins with differing levels of transcriptional activating activities have been shown to recapitulate the induction of different neuronal cell types elicited by varying levels of Shh activity (Stamataki et al, 2005). Of the few bona fide transcriptional Hh targets that have been characterised in detail, all have been found to have sequences similar or identical to the Gli consensus-binding site upstream of their promoters (Agren et al, 2004;Alexandre et al, 1996;Dai et al, 1999;Gustafsson et al, 2002;Laner-Plamberger et al, 2009;Sasaki et al, 1997;Winklmayr et al, 2010). Genomewide chromatin immunoprecipitation (ChIP) screens, however, have identified hundreds of putative Gli/Hh targets, a significant proportion of which are not associated with Gli consensus-binding sites (Vokes et al, 2007;Vokes et al, 2008).…”

Small noncoding differentially methylated copy-number variants, including lncRNA genes, cause a lethal lung developmental disorder

“…Although only one of the distant GLI-binding sites of SDR perfectly matches the consensus ''core'' sequence, the clustering of nine of these sites within this segment may have synergistic effects on GLI2 binding. Moreover, the variant GLI-binding sites with relatively low affinity have been recently shown to strongly induce transcription when present in native promoters (Winklmayr et al 2010). Since we did not detect interactions between SDR and the FOXF1 promoter in lymphoblasts, where FOXF1 is not expressed, we propose that chromatin looping between SDR and FOXF1 allows GLI2 to increase FOXF1 activity specifically in lung endothelium.…”