Signal Strength Dictates Phosphoinositide 3-Kinase Contribution to Ras/Extracellular Signal-Regulated Kinase 1 and 2 Activation via Differential Gab1/Shp2 Recruitment: Consequences for Resistance to Epidermal Growth Factor Receptor Inhibition (original) (raw)
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Molecular and Cellular Biology, 2008
Phosphoinositide 3-kinase (PI3K) participates in extracellular signal-regulated kinase 1 and 2 (ERK1-2) activation according to signal strength, through unknown mechanisms. We report herein that Gab1/Shp2 constitutes a PI3K-dependent checkpoint of ERK1-2 activation regulated according to signal intensity. Indeed, by up-and down-regulation of signal strength in different cell lines and through different methods, we observed that Gab1/Shp2 and Ras/ERK1-2 in concert become independent of PI3K upon strong epidermal growth factor receptor (EGFR) stimulation and dependent on PI3K upon limited EGFR activation. Using Gab1 mutants, we observed that this conditional role of PI3K is dictated by the EGFR capability of recruiting Gab1 through Grb2 or through the PI3K lipid product PIP 3 , according to a high or weak level of receptor stimulation, respectively. In agreement, Grb2 siRNA generates, in cells with maximal EGFR stimulation, a strong dependence on PI3K for both Gab1/Shp2 and ERK1-2 activation. Therefore, Ras/ERK1-2 depends on PI3K only when PIP 3 is required to recruit Gab1/Shp2, which occurs only under weak EGFR mobilization. Finally, we show that, in glioblastoma cells displaying residual EGFR activation, this compensatory mechanism becomes necessary to efficiently activate ERK1-2, which could probably contribute to tumor resistance to EGFR inhibitors.
Reactivation of ERK signaling causes resistance to EGFR kinase inhibitors
Cancer discovery, 2012
The clinical efficacy of epidermal growth factor receptor (EGFR) kinase inhibitors is limited by the development of drug resistance. The irreversible EGFR kinase inhibitor WZ4002 is effective against the most common mechanism of drug resistance mediated by the EGFR T790M mutation. Here, we show, in multiple complementary models, that resistance to WZ4002 develops through aberrant activation of extracellular signal-regulated kinase (ERK) signaling caused by either an amplification of mitogen-activated protein kinase 1 (MAPK1) or by downregulation of negative regulators of ERK signaling. Inhibition of MAP-ERK kinase (MEK) or ERK restores sensitivity to WZ4002 and prevents the emergence of drug resistance. We further identify MAPK1 amplification in an erlotinib-resistant EGFR-mutant non-small cell lung carcinoma patient. In addition, the WZ4002-resistant MAPK1-amplified cells also show an increase both in EGFR internalization and a decrease in sensitivity to cytotoxic chemotherapy. Our...
Activation of extracellular-regulated kinases by normal and mutant EGF receptors
Biochimica et Biophysica Acta (BBA) - Molecular Cell Research, 2001
Glioblastoma cells express a mutant EGF receptor (EGFRvIII) that has constitutive tyrosine kinase activity and enhances their tumorigenicity. Here we show that EGFRvIII promotes constitutive phosphorylation of extracellular regulated kinases (ERKs) in glioblastoma cells in the absence of EGF. EGFRvIII also promoted constitutive activation of phosphoinositide 3kinase in these cells, as assessed by phosphorylation of protein kinase B/akt. As expected, phosphorylation of protein kinase B/akt was blocked by the phosphoinositide 3-kinase inhibitors wortmannin and LY294002. Less expectedly, we found that this treatment also blocked EGFRvIII-induced phosphorylation of ERKs. In contrast, ERK phosphorylation induced by EGF-activated normal EGF receptor in the same cells was largely unaffected by treatment with phosphoinositide 3-kinase inhibitors. This difference in behavior between the normal receptor and EGFRvIII was not due to differences in the levels of activated EGFRvIII and wild-type EGF receptor, as the two types of receptor were tyrosine phosphorylated to a similar extent under the experimental conditions used. EGFRvIII activation of ERKs was also sensitive to the phospholipase C inhibitor U73122, whereas ERK activation by normal EGF receptor was not. These results show that EGFRvIII and wildtype EGF receptor preferentially use different signaling pathways to induce ERK phosphorylation. The different mechanisms of ERK activation used by normal and mutant EGF receptors may be important in understanding the potent tumorigenic activity of EGFRvIII. ß
Journal of Biological Chemistry, 2008
Spatiotemporal aspects of ERK activation are stimulus-specific and dictate cellular consequences. They are dependent upon dual specificity phosphatases (DUSPs) that bind ERK via docking domains and can both inactivate and anchor ERK in cellular compartments. Using high throughput fluorescence microscopy in combination with a system where endogenous ERKs are removed and replaced with wild-type or mutated ERK2-green fluorescent protein (GFP), we show that ERK2 activation responses to epidermal growth factor (EGF) and protein kinase C (PKC) are transient and sustained, respectively. PKC-mediated ERK2 activation is associated with prolonged nuclear localization in the dephosphorylated form, whereas EGF-stimulated ERK2 activation mediates only transient nuclear accumulation. By using short inhibitory RNAs to nuclear inducible DUSP1, -2, or -4 (alone or in combination), we demonstrate that all three of these enzymes contribute to the dephosphorylation of PKC (but not EGF)-activated ERK2 in the nucleus but that they have opposing effects on localization. DUSP2 and -4 inactivate and anchor ERK2, whereas DUSP1 dephosphorylates ERK in the nucleus but allows its traffic back to the cytoplasm. Overexpression of DUSP1, -2, or -4 prevented ERK2 activation, but only DUSP2 and -4 caused ERK2-GFP nuclear accumulation or could be immunoprecipitated with ERK2. Furthermore, protein synthesis inhibition or replacement of wild-type ERK2-GFP with docking domain mutants selectively increased PKC effects on ERK activity and altered ERK2-GFP localization. These mutations also impaired the ability of ERK2-GFP to bind DUSP2 and -4. Together, our data reveal a novel, stimulus-specific, and phosphatase-specific mechanism of ERK2 regulation in the nucleus by DUSP1, -2, and -4. Extracellular signal-regulated kinases 1 and 2 (ERK1/2, referred to as ERK 2 herein) are the prototypic members of the mitogen-activated protein kinase (MAPK) family, and are activated by a wide variety of stimuli. The biological outcome of ERK signaling is dependent upon the magnitude, duration, and localization of its activation (1-3). ERK is typically associated with its upstream activator MAPK/ERK kinase (MEK) in the cytosol of quiescent cells. Upon phosphorylation by MEK, ERKs dissociate and typically translocate to the nucleus where they can phosphorylate transcription factors and immediate early gene products, leading to altered
A TNF–JNK–Axl–ERK signaling axis mediates primary resistance to EGFR inhibition in glioblastoma
Nature Neuroscience, 2017
Aberrant EGFR signaling is widespread in cancer, making the EGFR an important target for therapy. EGFR gene amplification and mutation are common in glioblastoma (GBM), but EGFR inhibition has not been effective in treating this tumor. Here, we propose that primary resistance to EGFR inhibition in glioma cells results from a rapid compensatory response to EGFR inhibition that mediates cell survival. We show that in glioma cells expressing either EGFR wild type or the mutant EGFRvIII, EGFR inhibition triggers a rapid adaptive response driven by increased TNF Users may view, print, copy, and download text and data-mine the content in such documents, for the purposes of academic research, subject always to the full Conditions of use:
Inhibition of DYRK1A destabilizes EGFR and reduces EGFR-dependent glioblastoma growth
Journal of Clinical Investigation, 2013
Glioblastomas (GBMs) are very aggressive tumors that are resistant to conventional chemo-and radiotherapy. New molecular therapeutic strategies are required to effectively eliminate the subpopulation of GBM tumorinitiating cells that are responsible for relapse. Since EGFR is altered in 50% of GBMs, it represents one of the most promising targets; however, EGFR kinase inhibitors have produced poor results in clinical assays, with no clear explanation for the observed resistance. We uncovered a fundamental role for the dual-specificity tyrosine phosphorylation-regulated kinase, DYRK1A, in regulating EGFR in GBMs. We found that DYRK1A was highly expressed in these tumors and that its expression was correlated with that of EGFR. Moreover, DYRK1A inhibition promoted EGFR degradation in primary GBM cell lines and neural progenitor cells, sharply reducing the self-renewal capacity of normal and tumorigenic cells. Most importantly, our data suggest that a subset of GBMs depends on high surface EGFR levels, as DYRK1A inhibition compromised their survival and produced a profound decrease in tumor burden. We propose that the recovery of EGFR stability is a key oncogenic event in a large proportion of gliomas and that pharmacological inhibition of DYRK1A could represent a promising therapeutic intervention for EGFR-dependent GBMs.
Oncotarget, 2015
We identified a synthetic lethality between PLK1 silencing and the expression of an oncogenic Epidermal Growth Factor Receptor, EGFRvIII. PLK1 promoted homologous recombination (HR), mitigating EGFRvIII induced oncogenic stress resulting from DNA damage accumulation. Accordingly, PLK1 inhibition enhanced the cytotoxic effects of the DNA damaging agent, temozolomide (TMZ). This effect was significantly more pronounced in an Ink4a/Arf(-/-) EGFRvIII glioblastoma model relative to an Ink4a/Arf(-/-) PDGF-β model. The tumoricidal and TMZ-sensitizing effects of BI2536 were uniformly observed across Ink4a/Arf(-/-) EGFRvIII glioblastoma clones that acquired independent resistance mechanisms to EGFR inhibitors, suggesting these resistant clones retain oncogenic stress that required PLK1 compensation. Although BI2536 significantly augmented the anti-neoplastic effect of EGFR inhibitors in the Ink4a/Arf(-/-) EGFRvIII model, durable response was not achieved until TMZ was added. Our results sugg...