Keratinocyte Integrin Crosstalk During Wound Healing.

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PROJECT SUMMARY Epidermal keratinocytes are vital to normal wound healing by restoring the epidermal barrier and secreting paracrine factors that govern diverse processes including wound angiogenesis and myofibroblast function. In pathogenic settings, impaired epidermal function results in chronically insufficient (e.g., diabetic ulcers) or over- exuberant healing (e.g., hypertrophic scars). Our long-term goal is to develop therapeutic paradigms through which integrins can be manipulated to modulate pathogenic keratinocyte function. While it is well established that integrins regulate proliferation, migration and growth factor signaling, their roles in orchestrating wound keratinocyte functions remain enigmatic. Moreover, while normal and wound keratinocytes express integrin 91, in vivo, upon explanation integrin 91 is lost, confounding observations made in previous studies, in vitro. Using genetically defined, virally transduced keratinocytes that express integrins 31 and/or 91 in different combinations, we discovered in the last project period that 91 exerts a cross-suppressive effect on wound cell function and gene expression that is governed by 31, including paracrine signals that promote endothelial cell function and autocrine signals that regulate basement membrane assembly. Using genetically defined mice that we have derived expressing different combinations of 31 and/or 91 in the epidermis, we also found that deletion of 91 from epidermis promoted wound angiogenesis and enhanced laminin 2 processing in the regenerating, epidermal basement membrane after injury. Based on our recently published studies and new foundation data, we now hypothesize that 91 cross-suppresses 31-dependent keratinocyte functions through inhibition of a novel 31-FAK-YAP/TAZ signaling axis. We further hypothesize that this signaling axis controls a gene expression program that promotes keratinocyte wound functions, including paracrine stimulation of endothelial cells and fibroblasts and autocrine regulation of basement membrane assembly. This hypothesis will be tested in three Aims using a combination of co-culture models, qPCR arrays, proteomics, PAC-seq mRNA analysis, cell biology, and defined genetic mouse models. At the end of this project period, we will have built on the foundation developed in the first project period to elucidate the complex signaling network downstream of integrin signaling in keratinocytes that governs paracrine and autocrine signaling in normal wounds. We will also have determined how these integrin signaling pathways are altered in epidermal tumors in which angiogenesis and other wound processes persist. In doing so, we will have developed the basis for novel integrin targeting therapeutics to modulate keratinocyte function and wound outcome.