INDUCTION OF DIFFERENTIATION IN NORMAL HUMAN KERATINOCYTES BY ADENOVIRUS-MEDIATED INTRODUCTION OF THE ETA-ISOFORMS AND DELTA-ISOFORMS OF PROTEIN-KINASE-C

Protein kinase C (PKC) plays a crucial role(s) in regulation of growth and differentiation of cells. In the present study, we examined possi ble roles of the alpha, delta, eta, and zeta isoforms of PKC in squamo us differentiation by overexpressing these genes in normal human kerat inocytes. Because of the difficulty of introducing foreign genes into keratinocytes, we used an adenovirus vector system, Ax, which allows e xpression of these genes at a high level in almost all the cells infec ted for at least 72 h. Increased kinase activity,vas demonstrated in t he cells overexpressing the alpha, delta, and eta isoforms. Overexpres sion of the eta isoform inhibited the growth of keratinocytes of human s and mice in a dose (multiplicity of infection [MOI])-dependent manne r, leading to G(1) arrest. The eta-overexpressing cells became enlarge d and flattened, showing squamous cell phenotypes. Expression and acti vity of transglutaminase 1, a key enzyme of squamous cell differentiat ion, were induced in the eta-overexpressing cells in dose (MOI)- and t ime dependent manners. The inhibition of growth and the induction of t ransglutaminase 1 activity were found only in the cells that express t he eta isoform endogenously, i.e., in human and mouse keratinocytes bu t not in human and mouse fibroblasts or COS1 cells. A dominant-negativ e eta isoform counteracted the induction of transglutaminase 1 by diff erentiation inducers such as a phorbol ester, 1a,25-dihydroxyvitamin D -3, and a high concentration of Ca2+. Among the isoforms examined, the delta isoform also inhibited the growth of keratinocytes and induced transglutaminase 1, but the alpha and zeta isoforms did not. These fin dings indicate that the eta and delta isoforms of PKC are involved cru cially in squamous cell differentiation.