Molecular and Comparative Genomics Analysis of Proteins and Peptides Involved in Epidermal Differentiation of Reptiles and Birds (Sauropsida)

The epidermis protects the body of vertebrates against many biological, chemical, physical and mechanical hazards present in the environment. The adaptation of the epidermis to a permanent life outside of the water was an essential step in the evolution of reptiles, birds, and mammals. The common ancestral origin of these terrestrial vertebrates is reflected in the shared presence of a gene cluster called the Epidermal Differentiation Complex (EDC), which controls the formation of the cornified layers of the epidermis. The aim of this study was to characterize novel factors and processes that contribute to the protective functions of the epidermis in reptiles and birds (Sauropsida). Comparative studies of genome and transcriptome sequences led to the first description of the EDC and its protein products in turtles, crocodilians and snakes. In each of these reptilian clades 50-100 epidermal differentiation genes were characterized at the sequence level. By reverse transcription-polymerase chain reaction experiments, turtle EDC genes were confirmed to be expressed in the skin or specifically in the scutes of the turtle shell. A newly generated antibody was used to demonstrate the expression of an avian EDC protein in feathers. Furthermore, specific features of the epidermal barrier, such as the defense mechanisms against microbes and the molecular cross-linking of corneous beta proteins (beta-keratins), were investigated in vitro. Reptilian skin-specific peptides were tested for their antimicrobial activities on Gram-negative and Gram-positive bacteria. Disulfide bonds of corneous beta proteins were studied by exposure to reduction and alkylation or oxidation followed by western blot analysis. Together, the results of these bioinformatics studies, analyses of gene expression, and mechanistic assays provided important new insights into the evolution and functions of epidermal proteins in sauropsids and the epidermal barrier to the environment in general.