Protein kinase C isozymes in human megakaryoblastic leukemia cell line, MEG-01: possible involvement of the isozymes in the differentiation process of MEG-01 cells.
The expression of the different protein kinase C (PKC) isozymes in various states of differentiation of the human megakaryoblastic leukaemia cell line MEG-01 were analysed using thermocycle amplification of mRNA and immunoblotting. MEG-01 expressed mRNAs of PKC alpha, -beta I, -beta II, -delta, -epsilon, -eta, -theta and -zeta, but not PKC gamma. At the protein molecule level, MEG-01 was observed to express PKC alpha, -beta I, -beta II,- epsilon, -theta and -zeta, but lack -gamma, -delta and -eta. When differentiation of MEG-01 was induced by 100 nm 12-O-tetradecanoyl-phorbol-13-acetate (TPA), rapid translocation from cytosol to membrane fraction and down-regulation of PKC alpha, -epsilon and -theta was observed in 1-2h. On the other hand, PKC beta I and -beta II were observed to translocate not only to the membrane fraction but also to the cytoskeletal fraction in a different manner, and their down-regulation, especially beta II, was very slow. The myristoylated, alanine-rich C kinase substrate (MARCKS) in the membrane fraction of MEG-01 cells was observed to decrease gradually throughout the differentiation process. Additionally, time-course study of TPA treatment indicated that incubation of the cells for 30 min is sufficient for differentiation. These results strongly suggest that the activation of PKC alpha, -epsilon and -theta is involved in the initiation of differentiation, and that PKC beta I and -beta II have important roles in the maintenance of differentiation. Although PKC zeta was resistant to TPA treatment and its translocation was reduced, the amount of this isozyme in the cytosol fraction decreased throughout the differentiation process.
Nagata
K I
KI
Department of Molecular Pathobiochemistry, Gifu University School of Medicine, Japan.
Okano
Y
Y
Nozawa
Y
Y
D001483:Base Sequence; D001792:Blood Platelets / Q000201:enzymology; D002454:Cell Differentiation; D004273:DNA, Neoplasm / Q000378:metabolism; D015536:Down-Regulation; D006801:Humans; D015151:Immunoblotting; D007527:Isoenzymes / Q000378:metabolism*; D007947:Leukemia, Megakaryoblastic, Acute / Q000201:enzymology* / Q000473:pathology; D008533:Megakaryocytes / Q000201:enzymology / Q000473:pathology*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction; D011493:Protein Kinase C / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D014407:Tumor Cells, Cultured
D004273:DNA, Neoplasm; D007527:Isoenzymes; D012333:RNA, Messenger; D011493:Protein Kinase C