Abstract
After injury to the animal epidermis, a variety of genes are transcriptionally activated in nearby cells to regenerate the missing cells and facilitate barrier repair. The range and types of diffusible wound signals that are produced by damaged epidermis and function to activate repair genes during epidermal regeneration remains a subject of very active study in many animals. In Drosophila embryos, we have discovered that serine protease function is locally activated around wound sites, and is also required for localized activation of epidermal repair genes. The serine protease trypsin is sufficient to induce a striking global epidermal wound response without inflicting cell death or compromising the integrity of the epithelial barrier. We developed a trypsin wounding treatment as an amplification tool to more fully understand the changes in the Drosophila transcriptome that occur after epidermal injury. By comparing our array results with similar results on mammalian skin wounding we can see which evolutionarily conserved pathways are activated after epidermal wounding in very diverse animals. Our innovative serine protease-mediated wounding protocol allowed us to identify 8 additional genes that are activated in epidermal cells in the immediate vicinity of puncture wounds, and the functions of many of these genes suggest novel genetic pathways that may control epidermal wound repair. Additionally, our data augments the evidence that clean puncture wounding can mount a powerful innate immune transcriptional response, with different innate immune genes being activated in an interesting variety of ways. These include puncture-induced activation only in epidermal cells in the immediate vicinity of wounds, or in all epidermal cells, or specifically in the fat body, or in multiple tissues.
Highlights
Drosophila’s epithelial barriers provide an organismal shield from physical damage and microbial infection
As a positive control for BSA-Green proteolysis, wild-type embryos that were puncture wounded with BSA-Green pre-incubated with trypsin, showed fluorescent signal throughout the entire embryonic body cavity (Figure 1D). These results reveal that localized endogenous proteolytic activity occurs around clean puncture wound sites
Our results indicate that a protease function is activated around embryonic puncture wound sites, and that serine protease activity is required to activate wound-induced transcription around wound sites
Summary
Drosophila’s epithelial barriers provide an organismal shield from physical damage and microbial infection. In Drosophila, the epidermal barrier consists of a single cell layer that secretes an impermeable, multilayered cuticle at the apical surface. The epidermis consists of several layers, the outermost being the stratum corneum, which is composed of dead squamous epithelial cells encased in a cornified cellular envelope, analogous to the Drosophila cuticle [2]. Drosophila and mammalian skin are structurally different, some of the genes that control the formation and repair of epidermal barriers are evolutionarily conserved between Drosophila and mammals, making Drosophila an advantageous model organism for studying the process of epidermal wound healing [3,4,5]. The grainy head (grh) gene encodes a conserved transcriptional regulator of epidermal barrier regeneration in both
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