Role of EPHA2 Serine 897 phosphorylation in thyroid cancer: molecular mechanisms and biological properties of a novel player in thyroid tumorigenesis

EPH (Erythropoietin-Producing Hepatocellular carcinoma cell line) receptor tyrosine kinases (RTK) belong to the largest subfamily of RTKs counting 14 genes in humans. Among them, EPHA2 is often overexpressed in a variety of human cancers, including thyroid carcinoma. Thyroid carcinomas are commonly driven by genetic lesions targeting the MAPK signaling cascade including rearrangements of several RTKs, such as RET and NTRK, or point mutations in RAS or BRAF. We have previously demonstrated, through a siRNA-based genetic screen of the human kinome, that EPHA2 expression is essential for viability of thyroid cancer cells in culture. To gain insight into the EPHA2 function in thyroid tumorigenesis, we studied the role of the intracellular domain of EPHA2 and, in particular, of its phosphorylation on Serine 897 (pSer897). Ser897 phosphorylation has been previously reported to mediate EPHA2 oncogenic activity. Ser897 is embedded in the consensus phosphorylation sequence for AGC (PKA, PKG, PKC) family kinases, including p90RSK, a direct MAPK target. Here we show that in thyroid cancer cells bearing oncogenic lesions in the MAPK signaling cascade, EPHA2 is robustly phosphorylated on Ser897. Treatment with chemical inhibitors targeting p90RSK or other MAPK pathway components blunts Ser897 phosphorylation of EPHA2. Recombinant p90RSK phosphorylates in vitro EPHA2 Ser897. Finally, RNA interference-mediated knock-down combined with rescue experiments demonstrate that Ser897 phosphorylation of EPHA2 mediates thyroid cancer cell proliferation and motility. Collectively, these findings point to EPHA2 pSer897 as a novel crucial mediator of the oncogenic MAPK signaling cascade, and in particular of p90RSK, in thyroid cancer.