ISOFORMS OF CATHEPSIN-D AND HUMAN EPIDERMAL DIFFERENTIATION

Cathepsin D is an ubiquitously expressed lysosomal aspartic proteinase , with well-determined structural and chemical properties but a less c learly defined biological role. In stratified epithelia, the chronolog y of cathepsin D activation and degradation can be connected with stag es of cellular differentiation. We partially purified cathepsin D from human epidermis and from separated stratum corneum by standard bioche mical procedures, monitored by SDS-PAGE and Western blotting, and veri fied its identity as to molecular mass, pH optimum, N-terminal sequenc ing, reactivity with the specific antibody, inhibition by pepstatin A, and specific enzyme activity. It had hemoglobin-degrading activity ov er the acid range, with maximum at pH 3. It also degraded bovine serum albumin, human keratins, and stratum corneum extracts at pH 4. We dis cerned all three isoforms of human cathepsin D (the 52 kDa proenzyme a nd the active forms at 48 kDa and 33 kDa) in the epidermis; both activ e forms were also seen in the stratum corneum, but the proenzyme was n ot. Gene expression of cathepsin D in epidermal keratinocytes resemble d that of suprabasal structural proteins (involucrin, keratin K10, tra nsglutaminase) in its response to the calcium switch. An antibody to t he 33 kDa isoform immunolocalized to the granular layer and the stratu m corneum (whereas antibodies to the 48 kDa isoform have been reported to stain mainly the upper spinous and granular layers). A plausible h ypothesis to harmonize these results is that cathepsin D is first expr essed as the proenzyme in the upper spinous layer, is activated in the lysosomes in the granular layer to the 48 kDa form, and is degraded t o the 33 kDa form in the transition zone between the granular layer an d the stratum corneum. As the stratum corneum is an acid environment, with an ambient pi-I of approximately 4.5, cathepsin D is available an d suited to contribute to desquamation. (C) Societe francaise de bioch imie et biologie moleculaire / Elsevier, Paris.