Characterization of canine filaggrin: gene structure and protein expression in dog skin

Abstract

Filaggrin (FLG) is a key protein for skin barrier formation and hydration of the stratum corneum. In humans, a strong association between FLG gene mutations and atopic dermatitis has been reported. Although similar pathogenesis and clinical manifestation have been argued in canine atopic dermatitis, our understanding of canine FLG is limited. The aim of this study was to determine the structure of the canine FLG gene and to raise anti-dog FLG antibodies, which will be useful to detect FLG protein in dog skin. The structure of the canine FLG gene was determined by analysing the publicly available canine genome DNA sequence. Polyclonal anti-dog FLG antibodies were raised based on the canine FLG sequence analysis and used for defining the FLG expression pattern in dog skin by western blotting and immunohistochemistry. Genomic DNA sequence analysis revealed that canine FLG contained four units of repeated sequences corresponding to FLG monomer protein. Western blots probed with anti-dog FLG monomer detected two bands at 59 and 54 kDa, which were estimated sizes. The results of immunohistochemistry showed that canine FLG was expressed in the stratum granulosum of the epidermis as a granular staining pattern in the cytoplasmic region. This study revealed the unique gene structure of canine FLG that results in production of FLG monomers larger than those of humans or mice. The anti-dog FLG antibodies raised in this study identified FLG in dog skin. These antibodies will enable us to screen FLG-deficient dogs with canine atopic dermatitis or ichthyosis.