Regulation of protein kinase C isoform proteins in phorbol ester-stimulated Jurkat T lymphoma cells.
Activation of protein kinase C (PKC) in T cells leads to a variety of responses including IL-2 production and IL-2 receptor expression. PKC consists of several isoforms that exhibit some different in vitro properties. We have set up a Western blotting system to explore the regulation of PKC isoforms during T cell activation. In Jurkat T lymphoma cells, PKC alpha, beta, delta, epsilon, and zeta were detected. PKC alpha and beta existed primarily in the cytosol, translocated to the membrane fraction after 10 minutes of treatment with PMA, and almost completely disappeared within 16 h. A larger fraction of PKC delta and epsilon existed in the membrane fraction compared to PKC alpha or beta, and PKC epsilon translocated to the membrane fraction rapidly. Translocation of PKC delta was not apparent after 1 h treatment with PMA, but total PKC delta protein was reduced within 4 to 6 h of treatment. Consistent with this, overnight treatment with PMA caused down-regulation of both PKC delta and epsilon, but to a lesser degree than was observed with PKC beta. Anti-PKC zeta antibody detected two bands at 82 and 75 kDa. The 75-kDa band existed mostly in the cytosol fraction and showed no translocation or down regulation after PMA. We present evidence that this 75-kDa band represents PKC zeta. Similar PMA-induced translocation responses were observed in murine thymocytes showing that the responses are not unique to PKC isoforms in Jurkat. These results demonstrate that it is possible for the PKC isoforms to be differentially regulated during T cell activation.
Tsutsumi
A
A
Department of Molecular Immunology, Syntex Discovery Research, Palo Alto, CA 94304.
Kubo
M
M
Fujii
H
H
Freire-Moar
J
J
Turck
C W
CW
Ransom
J T
JT
D000818:Animals; D001692:Biological Transport; D015536:Down-Regulation; D005260:Female; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism*; D008213:Lymphocyte Activation; D016399:Lymphoma, T-Cell / Q000201:enzymology; D051379:Mice; D008807:Mice, Inbred BALB C; D011493:Protein Kinase C / Q000276:immunology / Q000378:metabolism*; D013601:T-Lymphocytes / Q000201:enzymology* / Q000276:immunology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*; D014407:Tumor Cells, Cultured
D007527:Isoenzymes; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate