Summary Our long term objective is to heal skin wounds with full functional restoration (regenerative wound healing) instead of scarring (reparative wound healing). We aspire to learn what factors control regeneration as supposed to repair during wound healing under the newly established paradigm of ?Wound-Induced Hair Neogenesis? (WIHN). In this model, new hair follicles emerge from the wound center when a large (>1cm) skin wound is made on the mice (e.g., C57BL/6). Our new finding in Spiny mice (Acomys) shows, however, that WIHN starts to form from the periphery of wounds toward the center. Our preliminary data further shows a distinct spatial distribution of mechanical stiffness across the wound field, and that perturbation of mechanotransduction in the wound bed alters the outcome of WIHN. These new findings prompted us to hypothesize that tissue mechanics modulate tissue regeneration and WIHN. WIHN is easily accessible and is a good model to evaluate this hypothesis. While epithelial placode formation can be initiated by different chemical morphogens present during embryonic development (which converge to induce beta-catenin signaling), in WIHN of both C57BL/6 and spiny mouse, the mechanical environment of the wound can modulate, in parallel or independently, the threshold of successful placode formation. This acts to alter the status of epithelia activation, basement membrane remodeling, and dermal condensation. In Aim 1A, we will compare the different cellular and molecular events leading to WIHN, contrasting spiny and C57BL/6 mice. Supported by the bioinformatic analyses, Twist1 is proposed as a master regulator for placode formation in the spiny mouse. Therefore, the role of Twist1 and its downstream Msx2 in WIHN will be examined in Aim1B. In Aim 2A, we will map the stiffness in different parts of the wound field employing atomic force microscopy accompanied by cell shape analyses and FRET-biosensors to indirectly ?visualize? the consequence of cell forces within the wound site. The role of tissue mechanics in WIHN will be further tested by perturbation studies. Aim 2B will investigate and define the molecular circuits which are functioning in the conductive mechanical environment for placode regeneration. Overall, the proposal aims to explore novel epidermal-dermal networks during regenerative wound healing from the perspective of epigenetic, molecular, and mechanical inputs that construct the grand theme of elements necessary for future progression of regenerative medicine. !