Malignant mesothelioma is a deadly disease with limited therapeutic options. EphB4 is an oncogenic tyrosine kinase receptor expressed in malignant mesothelioma as well as in a variety of cancers. It is involved in tumor microenvironment mediating angiogenesis and invasive cellular effects via both EphrinB2 ligand-dependent and independent mechanisms. The molecular network underlying EphB4 oncogenic effects is still unclear. Here we show that EphB4 expression in malignant mesothelioma cells is markedly decreased upon neutralization of cancer-secreted IGF-II. In particular, we demonstrate that EphB4 protein expression in malignant mesothelioma cells depend upon a degradation rescue mechanism controlled by the autocrine IGF-II-insulin receptor-A specific signaling axis. We show that the regulation of EphB4 expression is linked to a competing post-translational modification of its carboxy-terminal tail via phosphorylation of its tyrosine 987 by the Insulin receptor isoform-A kinase-associated activity in response to the autocrine IGF-II stimuli. Neutralization of this autocrine-induced EphB4-phosphorylation by IGF-II associates with the increased ubiquitination of EphB4 carboxy-terminal tail and with its rapid degradation. We also describe a novel Ubiquitin binding motif in the targeted region as part of the identified EphB4 phosphodegron and provide 3D modeling data supporting a possible model for the acute EphB4 PTM-driven regulation by IGF-II. Altogether, these findings disclose a novel molecular mechanism for the maintenance of EphB4-expression in malignant mesothelioma cells and other IGF-II-secreting cancers (IGF2omas).
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We thank Dr. Steven Albelda (University of Pennsylvania, USA) for providing the MSTO211H cell line and his constructive criticism during the manuscript preparation; Dr Veronica Vella (University of Catania, Italy) for IR system expression data in colorectal cancer cell lines; Dr Parkash S Gill (USC, Los Angeles, CA) for critical insights in manuscript revision. We also thank ASEM srl (Treviso, Italy) and the GianAmico Alessandrini Family for supporting PS research.
PS conceptualized the study, designed and performed experiments, performed sequence comparison, conserved domains search and prepared manuscript; GP prepared cDNAs from cancer cells, provided unpublished data used for Table 1 and contributed to the manuscript preparation; EC performed 3D modeling analysis, AG provided conceptual insight and logistic support; SJW performed experiments, contributed to manuscript preparation and managed technical and administrative tasks.
Conflict of interest
The authors declare that they have no conflict of interest.
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