Contrasting effects of two tumour promoters, phorbol myristate acetate and okadaic acid, on T-cell responses and activation of p42 MAP-kinase/ERK-2.
The induction of T-cell growth by the T-cell antigen receptor (TcR) is dependent on a co-ordinated process of phosphorylation and dephosphorylation of intracellular proteins. An intermediary in this signalling pathway is the serine kinase, p42 mitogen-activated protein kinase (p42MAPK), also known as microtubule-associated protein-2 kinase (MAP-2K). MAP-kinase is activated upon the acquisition of tyrosine as well as threonine phosphate groups and removal of either by specific tyrosine or serine/threonine phosphatases abrogates kinase activity. Okadaic acid (OA), a tumour promoter and potent inhibitor of type 1 and 2A serine/threonine protein phosphatases (PP1 and PP2A), induced MAP-kinase activity in Jurkat T cells in a dose-dependent fashion with optimal effect at 1 microM. Compared to rapid activation (peak < 10 min) of MAP-kinase by another tumour promoter, the phorbol ester, PMA, the effect of OA was delayed (> 30 min) and more sustained. In spite of activating a growth-promoting kinase, OA differed from PMA by its lack of mitogenic activity and failure to induce CD25 [interleukin-2R alpha (IL-2R alpha)] expression in normal human T cells. This implies that PP1 and PP2A also act downstream of MAP-kinase to facilitate later cell cycle events. PMA induced a 42,000 MW tyrosine phosphoprotein which co-electrophoresed and co-chromatographed with ERK-2, a p42 MAP-kinase. Although OA induced an identical Mono-Q peak, there was less avid tyrosine phosphorylation of p42. OA also differed from PMA to the extent by which it induced mobility shift of the tyrosine protein kinase, p56lck, which has been implicated in p42MAPK activation in T cells. Taken together, these results indicate that OA and PMA exert both overlapping as well as divergent effects on lymphocyte growth pathways.
Amaral
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Department of Medicine, UCLA School of Medicine 90024-1680.
Casillas
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D002478:Cells, Cultured; D004988:Ethers, Cyclic / Q000494:pharmacology*; D006801:Humans; D015151:Immunoblotting; D066298:In Vitro Techniques; D007700:Kinetics; D019950:Mitogen-Activated Protein Kinase 1; D019319:Okadaic Acid; D010749:Phosphoprotein Phosphatases / Q000037:antagonists & inhibitors*; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism*; D011505:Protein-Tyrosine Kinases / Q000378:metabolism*; D015375:Receptors, Interleukin-2 / Q000032:analysis; D011994:Recombinant Proteins / Q000378:metabolism; D013601:T-Lymphocytes / Q000187:drug effects* / Q000276:immunology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology*
D004988:Ethers, Cyclic; D015375:Receptors, Interleukin-2; D011994:Recombinant Proteins; D019319:Okadaic Acid; D011505:Protein-Tyrosine Kinases; D017346:Protein Serine-Threonine Kinases; D019950:Mitogen-Activated Protein Kinase 1; D010749:Phosphoprotein Phosphatases; D013755:Tetradecanoylphorbol Acetate