The DNA synthesis of leukemic (L 2C) guinea pig B lymphocytes involves a permanent activation of protein kinase C without corresponding phosphoinositide hydrolysis

• Published in: Leukemia research Vol. 13; no. 7; pp. 583 - 594
• Main Authors: Vial, Henri J., Parant, Marc R., Marie, Josette Sainte, Laurent, Anne M., Le Peuch, Christian J.
• Format: Journal Article
• Language: English
• Published: Oxford Elsevier Ltd 1989; Elsevier Science
• Subjects: Animals; B lymphocyte; B-Lymphocytes - enzymology; B-Lymphocytes - metabolism; Biological and medical sciences; Burkitt Lymphoma - enzymology; Burkitt Lymphoma - genetics; Burkitt Lymphoma - metabolism; Cell Line; Chromatography, DEAE-Cellulose; Cyclic AMP - antagonists & inhibitors; DNA - biosynthesis; Enzyme Activation; Female; Fundamental and applied biological sciences. Psychology; Fundamental immunology; Guinea Pigs; Hydrolysis; Immunobiology; L 2C; Leukemia; Lymphoid cells: ontogeny, maturation, markers, receptors, circulation and recirculation; Phosphatidic Acids - biosynthesis; Phosphatidylinositol 4,5-Diphosphate; Phosphatidylinositols - biosynthesis; Phosphatidylinositols - metabolism; phosphoinositide; Phospholipids - analysis; polymyxin B; protein kinase C; Protein Kinase C - antagonists & inhibitors; Protein Kinase C - metabolism; Tumor Cells, Cultured - enzymology; Tumor Cells, Cultured - metabolism; PmB, polymyxin B; PMBN, polymyxin B nonapeptide; PKC, protein kinase C; Leukemia; L 2C; polymyxin B; DAG, diacylglycerol; B lymphocyte; TLC, thin layer chromatography; IP 1, IP 2, IP 3 , inositol mono- , di- , tris-phosphate; PPI, polyphosphoinositide; phosphoinositide; TPA, 12- O-tetradecanoyl-phorbol-13-acetate; PMSF, phenylmethylsulfonyl fluoride; mIG, membrane immunoglobulin; L 2C , leukemic strain-2 cells; Hepes, 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid; PIP 1, PIP 2 , phosphatidylinositol 4- , 4,5-phosphate; protein kinase C; Phosphoinositide; Protein kinase C; Radiolabelling; Enzyme; Leukocyte; Rodentia; Enzyme inhibitor; DNA synthesis; Activation; Hemopathy; B-Lymphocyte; Blood; Vertebrata; Blood cell; Mammalia; Guinea pig
• ISSN: 0145-2126; 1873-5835
• DOI: 10.1016/0145-2126(89)90125-2

L 2C B lymphocytes have a constant high DNA synthesis due to their continuous proliferative state. The addition of polymyxin B (PmB), a rather selective inhibitor of protein kinase C, stopped ( 3H)thymidine incorporation with an IC 50 of 10 μM when added 18 h before measuring DNA synthesis. Interestingly, PmB inhibition of DNA synthesis was suppressed when 4 nM 12- O-tetradecanoylphorbol-13-acetate was added along with PmB, indicating that PmB may act through inhibition of protein kinase C. In the node and spleen lymphocytes of normal guinea pigs, protein kinase C activity was entirely cytosolic and was eluted at 0.12 M NaCl when adsorbed on DEAE-cellulose. In L 2C leukemic lymphocytes, total protein kinase C activity was of the same order of magnitude, but 20% of it was associated with the membrane fraction. The lipid-dependent activity, eluted at 0.12 M NaCl from cytosolic and membrane fractions, was suppressed by staurosporine with an IC 50 of 10–40 nM and by polymyxin B with an IC 50 of 2–6 μM. Phosphoinositide metabolism was studied in the transformed cells. Incorporation of 32Pi into polyphosphoinositides was considerable, whereas much more time was required for a tiny incorporation of inositol. We detected no release of radioactive inositol triphosphate. Taken together, these results suggest that protein kinase C function is indispensible for triggering L 2C leukemic lymphocyte proliferation. The causes of this permanent activation merit further investigation.