Transcriptional analysis of scar-free wound healing during early stages of tail regeneration in the green anole lizard, Anolis carolinensis

Abstract

While tail regeneration is observed in a number of vertebrate groups, including teleost fish such as the zebrafish, urodeles such as the axolotl, and anurans such as Xenopus tadpoles, mammalian and avian amniote vertebrates have lost this capacity. Among the amniotes, squamate reptiles such as lizards retain the ability to regrow their tails and also display the capacity to autotomize, or self-amputate, these structures as a predator evasion response. The regenerated tail is a biomechanically functional structure consisting of regrown and repatterned tissues including spinal cord, peripheral nerves, cartilage, skeletal muscle, vasculature, and skin. The green anole lizard, Anolis carolinensis, was the first reptile with a sequenced and annotated genome, thus allowing transcriptomic analyses. Furthermore, anoles exhibit a high degree of conservation of both innate and adaptive immune pathways with mammals. In histological analyses of tail regeneration in the green anole, we observed early cellular infiltration of the tail stump followed by a second phase of epithelial formation of the wound surface. These events preceded the period of rapid tail outgrowth, which typically starts at 10 days post autotomy. To identify genes activated during the initial phase of tail regeneration, we carried out whole transcriptome sequencing at 0.5, 1, 2, 3, 4, and 5 days post-autotomy. We identified that 5315 genes were differentially expressed between any of these time points and clustered into two major groups with elevated expression either in a first phase (0.5–1 DPA) or in a later second phase (3–5 DPA), with a marked shift in expression at 2 DPA. Genes with elevated expression in the first phase included those regulating the immune system, T cell receptor signaling, and the p38 MAPK signaling pathway. Genes upregulated in the second phase included those regulating cell proliferation, developmental growth, and Wnt/Hippo signaling pathways. Identifying the immunomodulatory events that set the stage for regenerative cell proliferation and outgrowth in an amniote model may help guide post-injury treatments as part of regenerative medical therapies.