Exp Dermatol 2001: 10: 193–203 Copyright C Munksgaard 2001 Printed in Denmark ¡ All rights reserved EXPERIMENTAL DERMATOLOGY ISSN 0906-6705 Proprotein convertase expression and localization in epidermis: evidence for multiple roles and substrates Pearton DJ, Nirunsuksiri W, Rehemtulla A, Lewis SP, Presland RB, Dale David J. Pearton 1 , BA. Proprotein convertase expression and localization in epidermis: evi- Wilas Nirunsuksiri 1,3 , dence for multiple roles and substrates. Alnawaz Rehemtulla 2 , Exp Dermatol 2001: 10: 193–203. C Munksgaard, 2001 S. Patrick Lewis 1 , Richard B. Presland 1 and Abstract: Specific proteolysis plays an important role in the terminal dif- Beverly A. Dale 1 ferentiation of keratinocytes in the epidermis and several types of proteases 1 Departments of Oral Biology, Periodontics, have been implicated in this process. The proprotein convertases (PCs) Biochemistry, Medicine/Dermatology, University are a family of Ca 2π -dependent serine proteases involved in processing and of Washington, Seattle, WA 98195; 2 Department activation of several types of substrates. In this study we examined the of Radiation Oncology, University of Michigan, expression and some potential substrates of PCs in epidermis. Four PCs Ann Arbor, MI; 3 Present address Dow are expressed in epidermis: furin, PACE4, PC5/6 and PC7/8. Furin is de- AgroSciences LLC, Indianapolis, IN tected in two forms, either with or without the transmembrane domain, suggesting occurrence of post-translational cleavage to produce a soluble enzyme. In addition the furin active site has differential accessibility in the granular layer of the epidermis relative to the basal layer, whereas antibodies to the transmembrane domain stain both layers. These find- ings suggest that furin has access to different types of substrates in granu- Key words: epidermal differentiation – lar cells as opposed to basal cells. PC7/8, in contrast, is detected throughout keratinocytes – furin – PACE4 – PC5/PC6 – the epidermis with antibodies to both the transmembrane and active site PC7/PC8 – profilaggrin – Notch-1 and no soluble form observed. A peptide PC inhibitor (dec-RVKR-CMK) Dr Beverly A. Dale, Dept of Oral Biology, inhibits cleavage of Notch-1, a receptor important in cell fate determi- University of Washington, Box 357132, Seattle, nation that is found throughout the epidermis. Profilaggrin, found in WA 98195 7132 the granular layer, is specifically cleaved by furin and PACE4 in vitro at Tel.: 206 543 4393 Fax: 206 685 3162 a site between the amino terminus and the first filaggrin repeat. This e-mail: bdale/u.washington.edu work suggests that the PCs play multiple roles during epidermal differen- tiation. Accepted for publication 12 January 2001 Introduction The sequential differentiation of keratinocytes in the epidermis is a complex, highly regulated series of events that results in the formation of the anu- cleate cornified cells of the stratum corneum from the relatively undifferentiated cells of the basal layer. This differentiation process is calcium de- pendent and requires extensive and tightly con- trolled changes in both protein expression and en- zymatic activities resulting in characteristic morphological changes of the cells. Proteolytic processing is an important mechanism for many of these events, including signaling, proprotein acti- vation, structural remodeling and desquamation. A number of different proteases have been iden- 193 tified that play a role in epidermal differentiation. These include calpain I (1), cathepsins C (2, 3), B, H and L (4), profilaggrin endopeptidase (PEP1) (5), the stratum corneum chymotryptic-like pro- tease (SCCP) and the stratum corneum thiol pro- tease (SCTP) (6). The importance of proteases in the epidermis is highlighted by the finding that null mutations of cathepsin C result in palmoplantar keratoderma disorders, specifically Haim–Munk and Papillon–Lefevre syndromes (2, 7, 8). A protease family that has been implicated in processing and differentiation in a number of tissues is the Proprotein Convertase (PC) family. This is a family of Ca 2π -dependent serine pro- teases related to bacterial subtilisin (for reviews see 9–12). To date 7 family members have been iden-