TY - JOUR
T1 - Ras/ERK1/2-mediated STAT3 Ser727 Phosphorylation by Familial Medullary Thyroid Carcinoma-associated RET Mutants Induces Full Activation of STAT3 and Is Required for c-fos Promoter Activation, Cell Mitogenicity, and Transformation
AU - Plaza-Menacho, Iván
AU - Sluis, Tineke van der
AU - Hollema, Harry
AU - Gimm, Oliver
AU - Buys, Charles H.C.M.
AU - Magee, Anthony I.
AU - Isacke, Clare M.
AU - Hofstra, Robert M.W.
AU - Eggen, Bart J.L.
N1 - Relation: http://www.rug.nl/gbb/
date_submitted:2007
Rights: University of Groningen, Groningen Biomolecular Sciences and Biotechnology Institute
PY - 2007
Y1 - 2007
N2 - The precise role of STAT3 Ser727 phosphorylation in RET-mediated cell transformation and oncogenesis is not well understood. In this study, we have shown that familial medullary thyroid carcinoma (FMTC) mutants RETY791F and RETS891A induced, in addition to Tyr705 phosphorylation, constitutive STAT3 Ser727 phosphorylation. Using inhibitors and dominant negative constructs, we have demonstrated that RETY791F and RETS891A induce STAT3 Ser727 phosphorylation via a canonical Ras/ERK1/2 pathway and that integration of the Ras/ERK1/2/ELK-1 and STAT3 pathways was required for up-regulation of the c-fos promoter by FMTC-RET. Moreover, inhibition of ERK1/2 had a more severe effect on cell proliferation and cell phenotype in HEK293 cells expressing RETS891A compared with control and RETWT-transfected cells. The transforming activity of RETY791F and RETS891A in NIH-3T3 cells was also inhibited by U0126, indicating a role of the ERK1/2 pathway in RET-mediated transformation. To investigate the biological significance of Ras/ERK1/2-induced STAT3 Ser727 phosphorylation for cell proliferation and transformation, N-Ras-transformed NIH-3T3 cells were employed. These cells displayed elevated levels of activated ERK1/2 and Ser727-phosphorylated STAT3, which were inhibited by treatment with U0126. Importantly, overexpression of STAT3, in which the Ser727 was mutated into Ala (STAT3S727A), rescued the transformed phenotype of N-Ras-transformed cells. Immunohistochemistry in tumor samples from FMTC patients showed strong nuclear staining of phosphorylated ERK1/2 and Ser727 STAT3. These data show that FMTC-RET mutants activate a Ras/ERK1/2/STAT3 Ser727 pathway, which plays an important role in cell mitogenicity and transformation.
AB - The precise role of STAT3 Ser727 phosphorylation in RET-mediated cell transformation and oncogenesis is not well understood. In this study, we have shown that familial medullary thyroid carcinoma (FMTC) mutants RETY791F and RETS891A induced, in addition to Tyr705 phosphorylation, constitutive STAT3 Ser727 phosphorylation. Using inhibitors and dominant negative constructs, we have demonstrated that RETY791F and RETS891A induce STAT3 Ser727 phosphorylation via a canonical Ras/ERK1/2 pathway and that integration of the Ras/ERK1/2/ELK-1 and STAT3 pathways was required for up-regulation of the c-fos promoter by FMTC-RET. Moreover, inhibition of ERK1/2 had a more severe effect on cell proliferation and cell phenotype in HEK293 cells expressing RETS891A compared with control and RETWT-transfected cells. The transforming activity of RETY791F and RETS891A in NIH-3T3 cells was also inhibited by U0126, indicating a role of the ERK1/2 pathway in RET-mediated transformation. To investigate the biological significance of Ras/ERK1/2-induced STAT3 Ser727 phosphorylation for cell proliferation and transformation, N-Ras-transformed NIH-3T3 cells were employed. These cells displayed elevated levels of activated ERK1/2 and Ser727-phosphorylated STAT3, which were inhibited by treatment with U0126. Importantly, overexpression of STAT3, in which the Ser727 was mutated into Ala (STAT3S727A), rescued the transformed phenotype of N-Ras-transformed cells. Immunohistochemistry in tumor samples from FMTC patients showed strong nuclear staining of phosphorylated ERK1/2 and Ser727 STAT3. These data show that FMTC-RET mutants activate a Ras/ERK1/2/STAT3 Ser727 pathway, which plays an important role in cell mitogenicity and transformation.
KW - SERINE PHOSPHORYLATION
KW - NEUROTROPHIC FACTOR
KW - CANCER-CELLS
KW - SER-727 PHOSPHORYLATION
KW - KINASE
KW - PHENOTYPE
KW - PATHWAY
KW - MUTATIONS
KW - 2B
U2 - 10.1074/jbc.M608952200
DO - 10.1074/jbc.M608952200
M3 - Article
VL - 282
SP - 6415
EP - 6424
JO - The Journal of Biological Chemistry
JF - The Journal of Biological Chemistry
SN - 0021-9258
IS - 9
ER -