Delineating key molecular programs that induce hair-follicle-inductive dermal fibroblasts using a novel single-cell genomic approach

Summary: Abstract Body: Hair follicles (HFs) form by signals communicated between epithelial cells and specialized dermal cells,dermal condensate (DC) cells. Although it has been long recognized that DC cells can induce new HF growth, how they initially form has remained elusive. We recently showed that Wnt and SHH corporate to induce DCs through a rapid sequence of cell cycle exit followed by molecular DC differentiation. This sequence spatially corresponds to different levels of Wnt and SHH activity in the dermis. If and how different levels of Wnt and SHH are responsible for the distinct steps of DC genesis is unknown, largely because they occur spatiotemporally tightly coupled. Here, we genetically repatterned levels of Wnt and SHH and found that this disrupts the sequence of cell cycle exit and molecular DC differentiation such that they occur independently of each other. This resulted in disorganized DCs and revealed that the two typically coupled processes can be separately triggered by changing Wnt and SHH levels. We applied a novel computational method, GeneTrajectory, to dissect out these two distinct processes and their regulation. Unexpectedly, we show that SHH activity augments Wnt signaling and that high Wnt signaling is sufficient to elicit cell cycle exit irrespective of DC gene expression. Further, we found that this Wnt level utilizes GLI3, a SHH transcription factor, to coordinate cell cycle exit with SHH activity. Next, by recreating covarying Wnt and SHH levels over the entire dermis, we show that the mere coupling of Wnt and SHH gradients is sufficient to initiate a proper sequence of cell cycle exit and molecular differentiation. Finally, we established an in vitro system to show that Wnt and SHH can reciprocally regulate each other and induce DC processes, suggesting a feedback mechanism to ensure coupled levels. This study provides insight into how Wnt and SHH gradients coordinate biological processes in DC formation, which will help guide efforts to recreate DCs to generate organotypic HF models and to promote adult HF regeneration. Yiqun Jiang¹, Ruiqi Li¹, Sarah Van¹, Sarah Platt¹, Hainan Lam¹, Yuval Kluger¹, Peggy Myung¹ 1. Yale University, New Haven, CT, United States. Cell Communication Networks and Stromal Biology