Trichohyalin is a major differentiation product of the inner root sheath cells of hair follicle, the medulla of the hair fiber, and is expressed in the epidermis, and other tissues including the hard palate, and filiform ridges of the tongue. It is thought to function in part as a keratin intermediate filament associated protein in these tissues. Trichohyalin is a substrate for transglutaminases (TGases), which crosslink it into polymers, and for the enzyme peptidylarginine deiminase (PAD) which converts protein-bound arginines to citrullines. We have expressed in bacteria domain 8 (representing about 40%) of human trichohyalin, and used it to study these two postsynthetic modification events. PAD converts 60% of the arginines to citrullines, resulting in the complete loss of its a-helical structure. Trichohyalin is used by all three TGases known to be present in the epidermis, but by calculation of kinetic parameters, TGase 3 enzyme uses it most efficiently. The kcat for the TGase 3 enzyme increases by a factor of about 10 following PAD modification. These data suggest that trichohyalin is first modified by PAD before crosslinking. We have explored the regulation of expression of the human trichohyalin gene. Constructs containing 3 kbp of upstream sequences coupled with a b-galactosidase reporter can confer expression in transgenic mice in the tongue and palate, but not epidermis or hair follicle. The proximal promoter of the gene exists in the first 135 bp above the transcription start site, and consists of an AP1 element which confers keratinocyte specific expression, and contains a calcium responsive element.
