STRATIFIN, A KERATINOCYTE SPECIFIC 14-3-3-PROTEIN, HARBORS A PLECKSTRIN HOMOLOGY (PH) DOMAIN AND ENHANCES PROTEIN-KINASE-C ACTIVITY

The intrinsic signal(s) responsible for the onset of human keratinocyt e terminal differentiation is not yet fully understood. Evidence has b een recently accumulated linking the phospholipase-mediated activation of protein kinase C to the coordinate changes in gene expression occu rring during keratinocyte terminal differentiation. Here we report the purification of a keratinocyte-derived protein enhancing protein kina se C enzymatic activity. The stimulator eluted as a peak with estimate d molecular mass of approximately 70 kDa, while analysis by SDS-PAGE s howed a 30 kDa protein migrating as a distinct doublet, suggesting the formation of a 30 kDa homodimer. The amino acid sequence analysis all owed the unambigous identification of the protein kinase C stimulator as a mixture of the highly homologous sigma (stratifin) and zeta isofo rms of 14-3-3 proteins, which are homodimers of identical 30 kDa subun its. Mono Q anion exchange chromatography and immunoblot analysis furt her confirmed that stratifin enhances protein kinase C activity. Strat ifin was originally sequenced from a human keratinocyte protein databa se, but its function was unknown. The pleckstrin homology domain has b een recently related to protein translocation to the cell membrane as well as to functional interactions of intracellular proteins involved in signal transduction. We show here that stratifin (and 14-3-3 zeta) harbors a pleckstrin homology domain, and the consequent functional im plications will be discussed.