Phosphorylated cystatin alpha is a natural substrate of epidermal transglutaminase for formation of skin cornified envelope.
Both keratohyalin granules (KHG) and cornified envelopes were stained histochemically in an indirect immunofluorescent study by antiphosphorylated cystatin alpha antibody, indicating that phosphorylated cystatin alpha is a component of the cornified envelope proteins. When phosphorylated cystatin alpha (P-cystatin alpha) was incubated with epidermal transglutaminase (TGase) and Ca2+ ions, polymerized protein was produced by formation of epsilon-(gamma-glutamyl)lysine cross-linking peptide bonds between lysine residues of cystatin alpha and glutamine residues of suitable protein(s) in the enzyme preparation. However, phosphorylated and non-phosphorylated cystatins were polymerized to similar extents by the TGase. Immunofluorescent and immunoelectron microscopic observations revealed that P-cystatin alpha could be detected in vivo in the KHG and cornified envelopes. Treatment of sphingosine, a specific inhibitor of protein kinase C, markedly suppressed the incorporation of cystatin alpha into KHG. Thus phosphorylation of cystatin alpha by protein kinase C may play an important role in targeting cystatin alpha into KHG.
Department of Dermatology, Kinki University School of Medicine, Osaka, Japan.
M
M
Takahashi
T
T
Tezuka
N
N
Katunuma
D000818:Animals; D015891:Cystatins / Q000378:metabolism*; D004591:Electrophoresis, Polyacrylamide Gel; D004817:Epidermis / Q000201:enzymology*; D008856:Microscopy, Fluorescence; D016253:Microscopy, Immunoelectron; D010766:Phosphorylation; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D051381:Rats; D012867:Skin / Q000648:ultrastructure; D012879:Skin Physiological Phenomena; D013110:Sphingosine / Q000494:pharmacology; D013379:Substrate Specificity; D011503:Transglutaminases / Q000378:metabolism*
C070572:Csta protein, rat; D015891:Cystatins; D011503:Transglutaminases; D011493:Protein Kinase C; D013110:Sphingosine