Epidermal cell differentiation inhibitor ADP-ribosylates small GTP-binding proteins and induces hyperplasia of epidermis

Abstract

Epidermal cell differentiation inhibitor (EDIN) is a recently discovered protein which inhibits terminal differentiation of cultured keratinocytes (Sugai, M., Enomoto, T., Hashimoto, K., Matsumoto, K., Matsuo, Y., Ohgai, H., Hong, Y.-M., Inoue, S., Yoshikawa, K., and Suginaka, H. (1990) Biochem. Biophys. Res. Commun. 173, 92-98). The amino acid sequenced deduced from the EDIN gene has revealed that EDIN shares high amino acid sequence homology with the exoenzyme C3 of Clostridium botulinum (Inoue, S., Sugai, M., Murooka, Y., Paik, S.-Y., Hong, Y.-M., Ohgai, H., and Suginaka, H. (1991) Biochem. Biophys. Res. Commun. 174, 459-464), which has been shown to ADP-ribosylate the rho/rac proteins (members of the small GTP-binding protein family). We show here that EDIN ADP-ribosylates rhoB p21 in time- and dose-dependent manners in a cell-free system. Kinetic studies of the ADP-ribosylation and peptide mapping of the reaction products of rhoB p21 by EDIN and C3 suggest that the mode of action of the ADP-ribosylation by EDIN is quite similar to that by C3 and that the ADP-ribosylation site of rhoB p21 by EDIN is presumably the same as that by C3. Proteins in epidermal membranes and keratinocyte homogenate with Mr values of about 22,000 are ADP-ribosylated by EDIN or C3. Treatment of cultured human keratinocytes by EDIN or C3 results in an inhibition of terminal differentiation and a stimulation of growth of the cells. Moreover, EDIN and C3 injected into adult mouse skin induce hyperplasia of epidermis. These results suggest that EDIN and C3 affect growth and differentiation of keratinocytes by ADP-ribosylation of protein(s) with a Mr of about 22,000, which may be the rho/rac proteins or related proteins.