The activation of PI3-K and PKC zeta in PMA-induced differentiation of HL-60 cells

The human myelocytic leukemia cell line HL-60 is a useful model for the stu dy of cellular differentiation. Phorbol 12-myristate 13-acetate (PMA) induc es the monocyte/macrophage-like differentiation of HL-60 cells and results in growth arrest, increasing adherence. In PMA-induced differentiation of H L-60 cells, phosphoinositide 3-kinase (PI 3-K) activity was measured as pho sphatidylinositol(3)P recovery from phosphatidylinositol by in vitro kinase assay. PI 3-K activity was increased in HL-60 cells that were stimulated b y 20 nM PMA and the activity was inhibited by pretreatment with 20 muM LY29 4002, a specific inhibitor of PI 3-K. Members of the protein kinase C (PKC) family have been suggested to be one of the downstream targets of PI 3-K. PKC zeta is one of the atypical PKCs, non-diacylglycerol-responsive PKCs, a nd the activity was measured by the ability of phosphorylation onto myelin basic protein. PMA also induced the activation of PKC zeta during monocytic differentiation of HL-60 cells, and LY294002-pretreated cells failed to in duce PKC zeta activation. The activity of PI 3-K is essential for PKC activ ation, and LY294002 blocks both monocytic differentiation of HL-60 cells an d activation of PKC zeta during PMA-induced cell differentiation. This impl ies that activated PI 3-K subsequently stimulates the PKC zeta in the proce ss of PMA-induced monocytic differentiation. (C) 2001 Elsevier Science Irel and Ltd. All rights reserved.