Selective role for β-protein kinase C in signaling for O2radical anion generation but not degranulation or adherence in differentiated HL60 cells

A role for protein kinase C (PKC) isotypes is implicated in the activation of phagocytic cell functions. An antisense approach was used to selectively deplete beta-PRC, both beta I- and beta II-PKC, but not alpha-PKC, delta-PKC, or zeta-PKC in HL60 cells differentiated to a neutrophil-like phenotype (dHL60 cells). Depletion of beta-PKC in dHL60 cells elicited selective inhibition of O-2(radical anion) generation triggered by fMet-Leu-Phe, immune complexes, or phorbol myristate acetate, an activator of PKC. In contrast, neither ligand-elicited beta-glucuronidase (azurophil granule) release nor adherence to fibronectin was inhibited by beta-PKC depletion. Ligand-induced phosphorylation of a subset of proteins was reduced in beta-PKC-depleted dHL60 cells. Phosphorylation of p47(phox) and translocation of p47(phox) to the membrane are essential for activation of the NADPH oxidase and generation of O-2(radical anion). beta-PKC depletion had no effect on the level of p47(phox) in dHL60 cells but did significantly decrease ligand-induced phosphorylation of this protein. Furthermore, translocation of p47(phox) to the membrane in response to phorbol myristate acetate or fMet-Leu-Phe was reduced in beta-PKC-depleted cells. These results indicate that beta-PKC is essential for signaling for O-2(radical anion) generation but not cell adherence or azurophil degranulation. Depletion of beta-PKC inhibited ligand-induced phosphorylation of p47phox, translocation of p47(phox) to the membrane, and activation of O-2(radical anion) generation.