Involvement of EphA2-mediated tyrosine phosphorylation of Shp2 in Shp2-regulated activation of extracellular signal-regulated kinase

Abstract

1Shp2 is a positive regulator for Erk activation downstream of receptor tyrosine kinases for growth factors. It has been controversial how Shp2 induces Erk activation. We here demonstrate that EphA2 is responsible for Shp2-mediated Erk activation by phosphorylating Tyr542 and Tyr580 of Shp2 in the cells stimulated with growth factors. In NMuMG mammary epithelial cells stimulated with hepatocyte growth factor (HGF), HGF-dependent Erk phosphorylation was prolonged only in the presence of EphA2. This Erk activation paralleled the phosphorylation of Tyr542/580 of Shp2 and the association of Grb2 with Shp2, suggesting the positive signal involving Grb2 signal to activate Ras-Erk pathway. Immunohistochemical studies of mammary cancer specimens revealed that the cancer progression was associated with both Tyr580 phosphorylation of Shp2 and increased expression of EphA2, which were also correlated with increased Erk phosphorylation. Overexpression of either Shp2Thr468Met (a phosphatase-defective mutant found in Lentigines, Electrocardiographic abnormalities, Ocular hypertelorism, Pulmonary stenosis, Abnormal genitalia, Retardation of growth and sensorineural Deafness (LEOPARD) syndrome) or Shp2Asn308Asp (a phosphatase-active mutant found in Noonan syndrome) with EphA2 exhibited comparable activation of Erk and stronger activation than wild-type Shp2, suggesting the phosphatase-independent Erk activation. Expression of Shp2Thr468Met with Tyr542/580Phe mutations resulted in the suppression of Erk activation. Phosphatase-active and -inactive, and wild-type Shp2s bound equally to Grb2, suggesting that phosphorylation of Tyr542/580 of Shp2 was essential but not sufficient for Shp2-mediated Erk activation. We found that Gab1 (Grb2-associated binder 1) was involved in the mutant Shp2-mediated Erk activation. Zebrafish injected with Shp2Thr468Met mRNA showed cardiac edema, whereas those depleted of EphA2b showed less phenotype, suggesting that EphA2 might partly account for the phenotype of LEOPARD syndrome. Collectively, tyrosine phosphorylation of Shp2 by EphA2 contributes to the phosphatase-independent Shp2-mediated activation of Erk and might be involved in Shp2-associated diseases.

PIM1 kinase regulates cell death, tumor growth and chemotherapy response in triple-negative breast cancer

“…By example we observed increased expression of CDKN1B , encoding cell cycle inhibitor p27, shown to be regulated by PIM1 37 . We also showed for the first time that PIM1 regulates PTPN11 , encoding SHP2, known to be important for maintenance of tumor initiating cells and clonogenic survival in TNBC 15 and its regulatory kinase EPHA2 38 . These results were independently validated at the protein level (Fig.…”

Phosphorylation of Rab-coupling protein by LMTK3 controls Rab14-dependent EphA2 trafficking to promote cell:cell repulsion

“…Ligand-engagement of EphA2 is known to promote phosphorylation of Tyr 588 in its cytodomain (Supplementary Fig. 5b) which triggers ligand-dependent downstream signalling24. Consistently, occupation of EphA2 with ephrin-A1 drives complete redistribution of the receptor from the cell surface to endosomes, and this is opposed by substitution of Tyr 588 with Phe (Supplementary Fig.…”

Shp2 Plays a Critical Role in IL-6-Induced EMT in Breast Cancer Cells

“…As shown in Figure 6A, the expression of the two gain-of-function Shp2 mutants, but not the Shp2 T468M mutant, enhanced Erk phosphorylation, and resulted in a significant decrease in E-cadherin, when compared with that in the wild-type Shp2 cells. Several reports indicate that the tyrosine phosphorylation of Shp2 at Tyr542 and Tyr580 is the most important post-translational modification [4]. To determine if tyrosine phosphorylation of Shp2 regulates IL-6-induced EMT in breast cancer cells, a phospho-mimicking mutant form of Shp2 (Shp2 2YE ) and a phospho-deficient mutant form of Shp2 (Shp2 2YF ), were introduced into the Shp2 knockdown cells.…”

“…The binding of phosphotyrosyl residues to SH2 domains relieves this autoinhibitory effect and activates the phosphatase function of Shp2 [3]. In addition, the tyrosine phosphorylation of Shp2 in its C-terminal region (Tyr542 and Tyr580), regulates the phosphatase activity and changes its substrate specificity [4]. The phosphatase ability of Shp2 can dephosphorylate large quantities of important signal molecules.…”