The activation of the phosphotyrosine phosphatase eta (r-PTP eta) is responsible for the somatostatin inhibition of PC Cl3 thyroid cell proliferation.
The aim of this study was the characterization of the intracellular effectors of the antiproliferative activity of somatostatin in PC Cl3 thyroid cells. Somatostatin inhibited PC Cl3 cell proliferation through the activation of a membrane phosphotyrosine phosphatase. Conversely, PC Cl3 cells stably expressing the v-mos oncogene (PC mos) were completely insensitive to the somatostatin antiproliferative effects since somatostatin was unable to stimulate a phosphotyrosine phosphatase activity. In PC mos cells basal phosphotyrosine phosphatase activity was also reduced, suggesting that the expression of a specific phosphotyrosine phosphatase was impaired in these transformed cells. We suggested that this phosphotyrosine phosphatase could be r-PTP eta whose expression was abolished in the PC mos cells. To directly prove the involvement of r-PTP eta in somatostatin's effect, we stably transfected this phosphatase in PC mos cells. This new cell line (PC mos/PTP eta) recovered somatostatin's ability to inhibit cell proliferation, showing dose-dependence and time course similar to those observed in PC Cl3 cells. Conversely, the transfection of a catalytically inactive mutant of r-PTP eta did not restore the antiproliferative effects of somatostatin. PC mos/PTP eta cells showed a high basal phosphotyrosine phosphatase activity which, similarly to PC Cl3 cells, was further increased after somatostatin treatment. The specificity of the role of r-PTP eta in somatostatin receptor signal transduction was demonstrated by measuring its specific activity after somatostatin treatment in an immunocomplex assay. Somatostatin highly increased r-PTP eta activity in PCCl3 and PC mos/PTP eta (+300%, P < 0.01) but not in PCmos cells. Conversely, no differences in somatostatin-stimulated SHP-2 activity, (approximately +50%, P < 0.05), were observed among all the cell lines. The activation of r-PTP eta by somatostatin caused, acting downstream of MAPK kinase, an inhibition of insulin-induced ERK1/2 activation with the subsequent blockade of the phosphorylation, ubiquitination, and proteasome degradation of the cyclin-dependent kinase inhibitor p27(kip1). Ultimately, high levels of p27(kip1) lead to cell proliferation arrest. In conclusion, somatostatin inhibition of PC Cl3 cell proliferation requires the activation of r-PTP eta which, through the inhibition of MAPK activity, causes the stabilization of the cell cycle inhibitor p27(kip1).
Florio
T
T
Pharmacology and Neuroscience, National Institute for Cancer Research (IST) and Advanced Biotechnology Center (CBA) Genova 16132, Italy.
Arena
S
S
Thellung
S
S
Iuliano
R
R
Corsaro
A
A
Massa
A
A
Pattarozzi
A
A
Bajetto
A
A
Trapasso
F
F
Fusco
A
A
Schettini
G
G
D015152:Blotting, Northern; D018797:Cell Cycle Proteins / Q000378:metabolism; D002455:Cell Division / Q000187:drug effects*; D002460:Cell Line; D050760:Cyclin-Dependent Kinase Inhibitor p27; D004357:Drug Synergism; D004789:Enzyme Activation / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005455:Fluorescent Antibody Technique; D019204:GTP-Binding Proteins / Q000502:physiology; D015870:Gene Expression; D007328:Insulin / Q000494:pharmacology; D007700:Kinetics; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D009154:Mutation; D010766:Phosphorylation; D017027:Protein Tyrosine Phosphatases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000032:analysis; D017481:Receptors, Somatostatin / Q000502:physiology; D015398:Signal Transduction; D013004:Somatostatin / Q000494:pharmacology*; D013961:Thyroid Gland / Q000166:cytology*; D013972:Thyrotropin / Q000494:pharmacology; D014162:Transfection; D025521:Tumor Suppressor Proteins / Q000378:metabolism; D014638:Vanadates / Q000494:pharmacology
D018797:Cell Cycle Proteins; D004791:Enzyme Inhibitors; D007328:Insulin; D012333:RNA, Messenger; D017481:Receptors, Somatostatin; D025521:Tumor Suppressor Proteins; D050760:Cyclin-Dependent Kinase Inhibitor p27; D014638:Vanadates; D013004:Somatostatin; D013972:Thyrotropin; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D017027:Protein Tyrosine Phosphatases; D019204:GTP-Binding Proteins