Deciphering transcription factor spatiotemporal and dosage effects in face development

PROJECT SUMMARY The face is at the center of our physical identity. Common genetic variants can lead to changes in gene expression dosage impacting normal-range facial differences among human populations. Additionally, deleterious genetic variants can negatively impact gene expression and lead to a craniofacial disorder which collectively account for approximately one-third of birth defects. Genetic studies have revealed that several sequence-specific transcription factors (TFs) are associated with both normal-range facial variation and craniofacial disorders with hallmarks of haploinsufficiency and distinct facial regional changes. For example, facial development is sensitive to changes in SOX9, a key facial TF, over a broad range with smaller changes in TF levels associated with more subtle but increased specificity in phenotypes. Besides affecting normal-range facial variation, 30-50% SOX9 gene dosage reduction in facial progenitor cells leads to a set of craniofacial features termed Pierre Robin sequence (PRS) characterized by underdevelopment of the lower jaw (micrognathia). These findings indicate the importance of precise regulation of TF expression and dosage during facial development. However, it remains unknown how TF changes directly link to gene regulation and ultimately translate into facial morphological consequences in vivo. The proposed study will use SOX9 as an example to start to address this knowledge gap with the newly generated Sox9 dTAG mouse model utilizing the dTAG degron system allowing stage-specific, reversible, and dosage-dependent protein perturbation. Using the Sox9 dTAG mice, this proposal will determine the stage-specific (Aim1) and dosage (Aim2) effects of SOX9 perturbation during facial development. Aim1 will aim to determine the consequences of transient complete SOX9 depletion in the developing face from molecular to morphological levels using single cell sequencing and quantitative morphometric analysis. Genome-wide SOX9 binding region dynamics will also be identified at key facial development timepoints. Aim 2 will aim to determine how in vivo SOX9 dosage titration at key facial development windows leads to acute gene expression changes and final morphology outcomes. Successful completion of these aims will advance the fundamental knowledge about how our faces are shaped and these findings will also shed light on future preventative therapies of the associated craniofacial disorders. My long-term career goal is to lead my independent research program investigating the transcriptional and epigenetic mechanisms of facial structure development and regeneration. To reach this goal, the proposed research and training plan will combine my scientific background in mouse developmental genetics and “web- lab” techniques and postdoctoral training in genomic and “dry-lab” techniques. I will be mentored by Dr. Joanna Wysocka in a rigorous scientific environment (Stanford Chemical and Systems Biology) with additional support from my collaborator Dr. Nathanael Gray. I believe this fellowship award will facilitate the successful execution of the proposed research and training plan for my transition into an independent craniofacial research scientist.