The problem of origination and diversification of integument derivatives in vertebrates is still a challenge. The homeobox (Hox) genes Hoxc13 control integument formation in vertebrate. Hoxc13 show strong expression in the integument development, are highly conserved across vertebrates, and show mutations that are associated with skin and appendages. To test whether the evolution of the integument is associated with positive selection or relaxation of Hoxc13, we obtained these genes in a wide range of vertebrates. In Hoxc13, we found evidence of diversifying selection after speciation during the origin of vertebrates. In addition, we found the glycine-rich regions in Hoxc13 protein in mammals, but not among non-mammalian taxa. Our results strongly implicate that Hoxc13 genes could have played an important role in the evolution of integument structure. J. Exp. Zool. (Mol. Dev. Evol.) 320B: 412–419, 2013. © 2013 Wiley Periodicals, Inc.