INVOLVEMENT OF PROTEIN-KINASE C-EPSILON IN THE STIMULATION OF ANIONICAMINO-ACID-TRANSPORT IN CULTURED HUMAN FIBROBLASTS

Protein kinase C (PKC) activation stimulates transport system X(AG)(-) for anionic amino acids in cultured human fibroblasts (Franchi-Gazzol a, R., Visigalli, R., Bussolati, O., and Gazzola, G. C. (1994) FEBS Le tt. 352, 109-112), To identify which PKC isoform is responsible for th is effect, aspartate transport through system X(AG)(-), PKC activity, and the subcellular distribution of PKC isoforms have been studied bef ore and after treatment with phorbol 12,13-dibutyrate (PDBu) in fibrob lasts maintained at low serum for 1 (control cells) or 7 days (quiesce nt cells), In control cells aspartate transport and PKC activity in th e particulate fraction were stimulated by short term PDBu treatment; b oth stimulatory effects were down-regulated by a prolonged exposure to the phorbol. In contrast, in quiescent cells aspartate transport and particulate PKC activity were higher than control under basal conditio ns, unaffected by a short term PDBu treatment, and lowered by a prolon ged incubation with the phorbol, In both control and quiescent cells a short term PDBu treatment modified PKC alpha distribution, increasing its membrane associated fraction. PKC delta was mostly in the soluble fraction and scarcely sensitive to PDBu. A brief exposure to PDBu inc reased membrane-associated PKC epsilon in control but not in quiescent cells, In these cells epsilon isoform was found exclusively in the pa rticulate fraction even in PDBu-untreated cells. A prolonged PDBu trea tment caused a partial down-regulation of membrane-associated PKC epsi lon in control cells and its marked decrease in quiescent cells, It is concluded that PKC-dependent changes in system Ii,, activity parallel the behavior of PKC epsilon, thus suggesting a specific role for this isoform in system X(AG)(-) regulation.