Low-affinity transcription factor binding sites shape morphogen responses and enhancer evolution.
In the era of functional genomics, the role of transcription factor (TF)-DNA binding affinity is of increasing interest: for example, it has recently been proposed that low-affinity genomic binding events, though frequent, are functionally irrelevant. Here, we investigate the role of binding site affinity in the transcriptional interpretation of Hedgehog (Hh) morphogen gradients. We noted that enhancers of several Hh-responsive Drosophila genes have low predicted affinity for Ci, the Gli family TF that transduces Hh signalling in the fly. Contrary to our initial hypothesis, improving the affinity of Ci/Gli sites in enhancers of dpp, wingless and stripe, by transplanting optimal sites from the patched gene, did not result in ectopic responses to Hh signalling. Instead, we found that these enhancers require low-affinity binding sites for normal activation in regions of relatively low signalling. When Ci/Gli sites in these enhancers were altered to improve their binding affinity, we observed patterning defects in the transcriptional response that are consistent with a switch from Ci-mediated activation to Ci-mediated repression. Synthetic transgenic reporters containing isolated Ci/Gli sites confirmed this finding in imaginal discs. We propose that the requirement for gene activation by Ci in the regions of low-to-moderate Hh signalling results in evolutionary pressure favouring weak binding sites in enhancers of certain Hh target genes.
['Ramos|Andrea I|AI|', 'Barolo|Scott|S|']
D000818:Animals D001483:Base Sequence D001665:Binding Sites D004268:DNA-Binding Proteins D004330:Drosophila D029721:Drosophila Proteins D019143:Evolution, Molecular D018507:Gene Expression Regulation, Developmental D053823:Hedgehog Proteins D060227:Imaginal Discs D007150:Immunohistochemistry D008957:Models, Genetic D008969:Molecular Sequence Data D011485:Protein Binding D012641:Selection, Genetic D016415:Sequence Alignment D014157:Transcription Factors D014921:Wings, Animal
enhancers binding affinity morphogen gradient Ci/Gli gene regulation hedgehog signalling