PROJECT SUMMARY/ABSTRACT Understanding how to induce skin regeneration instead of scarring will have broad implications clinically and cosmetically because; they inhibit mobility, are painful, and are a source of psychological trauma. In the United States, treatment for burn victims alone amounts to $7.5 billion annually, the total market for scar treatment is estimated to be around $12 billion. A lack of understanding of the role of fibroblast heterogeneity inhibits progress in developing novel treatments for clinical conditions associated with scarring, such as, scleroderma, diabetic ulcers, psoriasis and fibrotic keloid scarring. Consequently, understanding how to regenerate skin has the potential to impact anyone who undergoes surgery, but also individuals with clinical conditions associated with scarring. Adult skin wounds heal with scars, but embryonic skin can regenerate without scarring. However, not knowing how to safely transfer the regeneration abilities of embryonic skin to adult skin remains a gap in knowledge. Thus, the long term goal of our research is to establish methods to induce fibroblasts in adult skin to have the same regenerative potential as their embryonic skin counterparts, without altering development and homeostasis. The objective of this application is to induce adult fibroblasts with embryonic transcription factors to reprogram skin to support regeneration and reverse aging. Embryonic and Neonatal skin have a transient Papillary Fibroblasts (PFs) that are lost during skin maturation and aging. Neonatal PFs reside beneath the epidermis, support skin regeneration, and express the canonical Wnt transcription factor, Lef1. A hallmark of skin aging is the degradation of PFs and the loss of Lef1 expression. Our central hypothesis is that Lef1 is the master regulator of Neonatal PF function, which induces scar-less skin regeneration and will restore PF identity in aged skin. Guided by our exciting preliminary data we will use novel transgenic mouse models and next generation sequencing technologies to investigate this hypothesis by the following specific aims: In aim 1 we will test the hypothesis that Lef1 is the master regulator of Neonatal PF function by tissue specific induction in adult skin. Here we will use wounding experiments and chamber grafting assays to test if adult scarring fibroblasts can be transformed to be regenerative. In aim 2, we will define the downstream pathways that Lef1 regulates to specify Neonatal PF functions. To do this we will perform ChIPSeq and Single-Cell-RNA-Seq with our regenerative transgenic model systems. Finally, in aim 3 we will determine if Lef1 can safely restore Neonatal PF identity and function in aged skin. We will analyze “old” transgenic mice with reprogrammed skin and test their ability to regenerate skin and inhibit “aging”. This proposal is innovative because of our novel approach of manipulating embryonic transcription factors in fibroblasts in vivo. The proposed research i...