The protein tyrosine phosphatase SHP-2 is required for EGFRvIII oncogenic transformation in human glioblastoma cells.
Oncogenic EGFRvIII is a naturally occurring oncoprotein and is expressed in about 40-50% of human glioblastomas, particularly those that arise de novo. To understand the molecular mechanisms by which this oncoprotein alters transforming phenotypes, and since our previous work indicated that SHP-2 protein tyrosine phosphatase activity modulated EGFRvIII activation and downstream signaling, we examined whether SHP-2 plays a role in EGFRvIII-induced oncogenesis by using both PTEN-deficient U87MG.EGFRvIII and PTEN-intact LN229.EGFRvIII cells. Inhibition of SHP-2 expression by Shp-2 siRNA inhibited cell growth, transformation and altered morphology of these EGFRvIII transformed GBM cells. Ectopic expression of a PTPase-inactive form of SHP-2, SHP-2 C459S, but not its wild-type SHP-2 or either of two SH2 domain mutants, abrogated transformation of EGFRvIII-expressing glioblastomas in soft agar and in nude mice. SHP-2 C459S cells grew slower and exhibited a more flattened morphology with more organized actin stress fibers under both full growth and low serum conditions. Furthermore, shp-2+/- and -/- mouse embryonic fibroblasts (MEFs) could not be transformed by EGFRvIII while shp-2+/+ MEFs displayed a fully transformed phenotype upon introduction of EGFRvIII, again indicating a requirement for functional SHP-2 in EGFRvIII transformation. Moreover, the SHP-2 PTPase activity inhibitor NSC-87877 inhibited endogenous SHP-2 activity, Erk phosphorylation and transformation in both GBM cell lines. EGFRvIII expression recruited SHP-2 to the receptor complex to transduce signals and also increased SHP-2 phosphorylation at Tyr542. Inhibition of EGFRvIII-induced cell growth and transformation by SHP-2 C459S or shp-2 siRNA was mediated by its ability to block cell cycle progression at different phases in these GBM cells. These data indicate that differential activation of SHP-2 phosphorylation at Tyr542 in these two GBM cell lines likely results in increased different PTPase activity and distinct mechanisms of cell cycle progression and SHP-2, in particular its PTPase activity, plays a critical role in EGFRvIII-mediated transformation.
Department of Neurosurgery, University of Pennsylvania School of Medicine, Philadelphia, PA 19104, USA.
Yi
Y
Zhan
George J
GJ
Counelis
Donald M
DM
O'Rourke
D000199:Actins / Q000378:metabolism; D000818:Animals; D045744:Cell Line, Tumor; D002471:Cell Transformation, Neoplastic / Q000378:metabolism*; D066246:ErbB Receptors / Q000378:metabolism*; D005347:Fibroblasts / Q000378:metabolism; D020868:Gene Silencing; D005909:Glioblastoma / Q000378:metabolism* / Q000473:pathology; D006801:Humans; D051379:Mice; D051059:PTEN Phosphohydrolase / Q000378:metabolism*; D054592:Protein Tyrosine Phosphatase, Non-Receptor Type 11 / Q000235:genetics / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000378:metabolism
D000199:Actins; D034741:RNA, Small Interfering; C410723:epidermal growth factor receptor VIII; D066246:ErbB Receptors; D054592:Protein Tyrosine Phosphatase, Non-Receptor Type 11; D051059:PTEN Phosphohydrolase