Gli3 utilizes Hand2 to synergistically regulate tissue-specific transcriptional networks

Despite a common understanding that Gli TFs are utilized to reiterate a Hh morphogen gradient, genetic analyses suggest craniofacial development does not completely fit this paradigm. We demonstrated that rather than being driven by a Hh threshold, robust Gli3 transcriptional activity during skeletal and glossal development required interaction with the bHLH TF Hand2. Not only did genetic and expression data support a co-factorial relationship, but genomic analysis further revealed that Gli3 and Hand2 were enriched at regulatory elements for genes essential for mandibular patterning and development. Interestingly, motif analysis at sites co-occupied by Gli3 and Hand2 uncovered mandibular-specific, low-affinity, divergent Gli binding motifs (dGBMs). Functional validation revealed these dGBMs conveyed synergistic activation of Gli targets essential for mandibular patterning and development. In summary, this work elucidates a novel, sequence-dependent mechanism for Gli transcriptional activity within the craniofacial complex that is independent of a graded Hh signal.