THE CATALYTIC DOMAIN OF PROTEIN-KINASE C-DELTA IN RECIPROCAL DELTA-CHIMERA AND EPSILON-CHIMERA MEDIATES PHORBOL ESTER-INDUCED MACROPHAGE DIFFERENTIATION OF MOUSE PROMYELOCYTES

The overexpression of protein kinase C-delta (PKC-delta), but not PKC- epsilon, enables the mouse myeloid cell line 32D to differentiate into macrophages when treated with phorbol esters such as 12-O-tetradecano ylphorbol-13-acetate (TPA). To determine the domain of PKC-delta that is responsible for this isotype-specific function, cDNAs that encode r eciprocal chimeras of PKC-delta and -epsilon (PKC-delta epsilon and PK C-epsilon delta) were constructed by exchanging regulatory and kinase domains using polymerase chain reaction technology. Both chimeras were stably expressed in 32D cells using the pLTR expression vector and di splayed protein kinase activity upon TPA treatment. TPA treatment of L epsilon delta, cells that overexpressed the PKC-epsilon delta chimera , induced a dramatically increased cell volume, surface adherence, sur face expression of Mac-1 and Mac-3, lysozyme production, and phagocyto sis. These are the characteristics of the macrophage phenotype found i n TPA-treated 32D cells that overexpressed PKC-delta. In contrast, lit tle effect was seen in L delta epsilon, 32D cells that overexpressed P KC-delta epsilon, with or without TPA treatment. A PKC inhibitor direc ted toward the catalytic domain of PKC, GF109203X, and a selective inh ibitor of PKC-delta, Rottlerin, blocked the TPA-induced differentiatio n of PKC-epsilon delta-overexpressing 32D cells. These results demonst rate that the catalytic domain of PKC-delta contains the primary deter minants for its activity in phorbol ester-induced macrophage different iation.