Expression of protein kinase C isoforms in HL60 and phorbol ester resistant HL525 cells.
Isoform protein expression and activity were investigated to evaluate whether differential expression of protein kinase C (PKC) accounts for the inability of HL525 cells to respond to phorbol 12-13,myristate acetate (PMA). Immunoblotting analysis of lysates from both the parental HL60 and PMA-resistant HL525 cells revealed expression of alpha-, beta-, delta- and zeta-PKC isoforms. Diminished expression of delta-PKC was observed along with increased expression of zeta-PKC in HL525 cells. Lysates from HL60 cells were resolved into the following 4 peaks of PKC activity using hydroxylapatite: peak 1, beta; peak 2, delta; peak 3, alpha; and peak 4, zeta. A similar elution pattern was noted for PKC isoforms expressed in HL525 cells although no peak 2-delta activity was recovered. PMA treatment of HL60 cells abolished activity of peaks 2, 3, 4; diminished alpha-PKC and increased zeta-PKC expression. In contrast, PMA treatment of HL525 cells did not abolish activity of peaks 1, 3 or 4, compared with the parental HL60 cells and failed to stimulate changes in zeta-PKC isoform expression. These data suggest that the inability of PMA to induce differentiation of HL525 cells may reside in diminished expression of delta-PKC along with alterations in zeta-PKC. The findings support the hypothesis that differential expression of PKC isoforms in HL525 cells contributes to the lack of morphological changes in response to PMA. The data also demonstrate that PMA alters the level of histone-dependent phosphotransferase activity as well as PKC cofactor dependence.
Wooten
M W
MW
Department of Zoology, Alabama Agricultural Experiment Station, Auburn University 36849.
Seibenhener
M L
ML
Soh
Y
Y
D015153:Blotting, Western; D015536:Down-Regulation / Q000187:drug effects / Q000502:physiology; D004351:Drug Resistance; D006801:Humans; D007536:Isomerism; D007942:Leukemia, Experimental / Q000201:enzymology* / Q000473:pathology*; D007951:Leukemia, Myeloid / Q000201:enzymology* / Q000473:pathology*; D011493:Protein Kinase C / Q000032:analysis* / Q000737:chemistry / Q000502:physiology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D014407:Tumor Cells, Cultured
D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate