Filaggrin Linker Segment Peptide and Cystatin α Are Parts of a Complex of the Cornified Envelope of Epidermis

We showed that filaggrin linker segment peptide (FLSP) is a glutamine-rich substrate of epidermal transglutaminase (TGase), conjugating enzymatically with a phosphorylated cystatin α (P-cystatin α) which is a lysine-rich substrate of the enzyme. This finding suggested that FLSP would be a component of cornified envelope of the epidermis as well as P-cystatin α. Here, we investigated thein vivolocation of FLSP. An antibody against the peptide conjugated with keyhole limpet hemocyanin (FLSP/KLH) reacted specifically with FLSP on immunoblots. Immunofluorescence histochemistry located specific staining with this antibody on keratohyalin granules and the cell membrane region of the stratum corneum. Specific staining was not detected when the antiserum was first absorbed by FLSP. Preembedding immunoelectron microscopic analysis showed that anti-FLSP/KLH antibody labeled with gold particles reacted with cornified envelope prepared from newborn rat stratum corneum. The high molecular weight protein enzymatically synthesized from phosphorylated cystatin α and FLSP by TGase reacted with both anti-FLSP/KLH antibody and anti-P-cystatin α antibody on Western blotting. These findings suggest that the FLSP–cystatin α conjugate is a component of the cornified envelope of the epidermis in rats.