Chronic expression of RET/PTC 3 enhances basal and insulin-stimulated PI3 kinase/AKT signaling and increases IRS-2 expression in FRTL-5 thyroid cells.
The RET/PTC3 oncogene is a genetically rearranged and constitutively activated tyrosine kinase receptor that is common in papillary thyroid cancer. Because RET/PTC3 is chronically overexpressed in these thyroid cancer cells, and RET/PTC3-expressing tumors are associated with overactivity of tyrosine kinase signaling pathways and a more aggressive clinical course, we questioned whether chronic RET/PTC3 expression enhances cellular responses to thyroid mitogens in vitro. We stably transfected FRTL-5 cells with the RET/PTC3 gene; transfected and control cell lines were cultured without insulin, TSH, or serum. Thymidine incorporation into DNA was enhanced in the RET/PTC3 cells, but transformation was not observed. RET/PTC3 cells demonstrated higher basal and insulin-stimulated levels of activated Akt, both of which were reduced by LY294002, a PI3 kinase inhibitor, but not PD98059, a MEK inhibitor. By contrast, mitogen activated protein kinase (MAP kinase) was only minimally activated in RET/PTC3 cells before and after stimulation. Consistent with preferential activation of PI3 kinase, increased levels of total and phosphorylated IRS2 protein, relative activation of PDK-1, and enhanced IRS2-p85 interactions were identified in RET/PTC3-expressing cells. RET/PTC3 cells were also sensitized to insulin-induced thymidine incorporation; this effect was blocked by PI3 kinase (LY294002) rather than MEK 1/2 (PD98059) inhibitors. In summary, we have demonstrated that RET/PTC3 expression enhances basal and insulin-stimulated DNA synthesis through PI3 kinase, cooperatively activates Akt with insulin via PI3 kinase, and preferentially activates the Akt rather than MAP kinase pathway in FRTL-5 cells.
Miyagi
Eri
E
Section of Endocrinology and Laboratory of Molecular Endocrinology, Washington Hospital Center/MedStar Research Institute, Washington, DC, USA.
Braga-Basaria
Milena
M
Hardy
Elena
E
Vasko
Vasily
V
Burman
Kenneth D
KD
Jhiang
Sissy
S
Saji
Motoyasu
M
Ringel
Matthew D
MD
D000818:Animals; D002460:Cell Line; D002471:Cell Transformation, Neoplastic; D004247:DNA / Q000096:biosynthesis; D007328:Insulin / Q000494:pharmacology; D055504:Insulin Receptor Substrate Proteins; D047908:Intracellular Signaling Peptides and Proteins; D015513:Oncogene Proteins / Q000378:metabolism; D015514:Oncogene Proteins, Fusion; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism*; D010749:Phosphoprotein Phosphatases / Q000378:metabolism; D010750:Phosphoproteins / Q000378:metabolism*; D000071636:Protein Phosphatase 2C; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D011505:Protein-Tyrosine Kinases; D011506:Proteins / Q000378:metabolism*; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D051057:Proto-Oncogene Proteins c-akt; D051381:Rats; D015398:Signal Transduction; D013936:Thymidine; D013961:Thyroid Gland; D013964:Thyroid Neoplasms; D014162:Transfection
D004247:DNA; D007328:Insulin; D055504:Insulin Receptor Substrate Proteins; D047908:Intracellular Signaling Peptides and Proteins; D015513:Oncogene Proteins; D015514:Oncogene Proteins, Fusion; D019869:Phosphatidylinositol 3-Kinases; D010749:Phosphoprotein Phosphatases; D010750:Phosphoproteins; D000071636:Protein Phosphatase 2C; D017346:Protein Serine-Threonine Kinases; D011505:Protein-Tyrosine Kinases; D011506:Proteins; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; D013936:Thymidine; C526299:Irs2 protein, rat; C490558:RET-PTC oncoprotein, rat; C494920:Akt1 protein, rat