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The Impact of Oncogenic EGFRvIII on the Proteome of Extracellular Vesicles Released from Glioblastoma Cells.
Mol Cell Proteomics 2018; 17(10):1948-1964MC

Abstract

Glioblastoma multiforme (GBM) is a highly aggressive and heterogeneous form of primary brain tumors, driven by a complex repertoire of oncogenic alterations, including the constitutively active epidermal growth factor receptor (EGFRvIII). EGFRvIII impacts both cell-intrinsic and non-cell autonomous aspects of GBM progression, including cell invasion, angiogenesis and modulation of the tumor microenvironment. This is, at least in part, attributable to the release and intercellular trafficking of extracellular vesicles (EVs), heterogeneous membrane structures containing multiple bioactive macromolecules. Here we analyzed the impact of EGFRvIII on the profile of glioma EVs using isogenic tumor cell lines, in which this oncogene exhibits a strong transforming activity. We observed that EGFRvIII expression alters the expression of EV-regulating genes (vesiculome) and EV properties, including their protein composition. Using mass spectrometry, quantitative proteomic analysis and Gene Ontology terms filters, we observed that EVs released by EGFRvIII-transformed cells were enriched for extracellular exosome and focal adhesion related proteins. Among them, we validated the association of pro-invasive proteins (CD44, BSG, CD151) with EVs of EGFRvIII expressing glioma cells, and downregulation of exosomal markers (CD81 and CD82) relative to EVs of EGFRvIII-negative cells. Nano-flow cytometry revealed that the EV output from individual glioma cell lines was highly heterogeneous, such that only a fraction of vesicles contained specific proteins (including EGFRvIII). Notably, cells expressing EGFRvIII released EVs double positive for CD44/BSG, and these proteins also colocalized in cellular filopodia. We also detected the expression of homophilic adhesion molecules and increased homologous EV uptake by EGFRvIII-positive glioma cells. These results suggest that oncogenic EGFRvIII reprograms the proteome and uptake of GBM-related EVs, a notion with considerable implications for their biological activity and properties relevant for the development of EV-based cancer biomarkers.

Authors+Show Affiliations

From the ‡Research Institute of the McGill University Health Centre, Glen Site, McGill University, Montreal, Quebec, H4A 3J1, Canada.From the ‡Research Institute of the McGill University Health Centre, Glen Site, McGill University, Montreal, Quebec, H4A 3J1, Canada.§Donnelly Centre and Departments of Molecular Genetics and Computer Science, University of Toronto, Toronto, Ontario, M5S 3E1, Canada. ¶Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, Toronto, Ontario, M5G 1X5, Canada.From the ‡Research Institute of the McGill University Health Centre, Glen Site, McGill University, Montreal, Quebec, H4A 3J1, Canada.§Donnelly Centre and Departments of Molecular Genetics and Computer Science, University of Toronto, Toronto, Ontario, M5S 3E1, Canada. ¶Lunenfeld-Tanenbaum Research Institute, Mount Sinai Hospital, Toronto, Ontario, M5G 1X5, Canada. ‖Canadian Institute for Advanced Research, Toronto, Ontario, M5G 1M1, Canada.From the ‡Research Institute of the McGill University Health Centre, Glen Site, McGill University, Montreal, Quebec, H4A 3J1, Canada; janusz.rak@mcgill.ca.

Pub Type(s)

Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

30006486

Citation

Choi, Dongsic, et al. "The Impact of Oncogenic EGFRvIII On the Proteome of Extracellular Vesicles Released From Glioblastoma Cells." Molecular & Cellular Proteomics : MCP, vol. 17, no. 10, 2018, pp. 1948-1964.
Choi D, Montermini L, Kim DK, et al. The Impact of Oncogenic EGFRvIII on the Proteome of Extracellular Vesicles Released from Glioblastoma Cells. Mol Cell Proteomics. 2018;17(10):1948-1964.
Choi, D., Montermini, L., Kim, D. K., Meehan, B., Roth, F. P., & Rak, J. (2018). The Impact of Oncogenic EGFRvIII on the Proteome of Extracellular Vesicles Released from Glioblastoma Cells. Molecular & Cellular Proteomics : MCP, 17(10), pp. 1948-1964. doi:10.1074/mcp.RA118.000644.
Choi D, et al. The Impact of Oncogenic EGFRvIII On the Proteome of Extracellular Vesicles Released From Glioblastoma Cells. Mol Cell Proteomics. 2018;17(10):1948-1964. PubMed PMID: 30006486.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - The Impact of Oncogenic EGFRvIII on the Proteome of Extracellular Vesicles Released from Glioblastoma Cells. AU - Choi,Dongsic, AU - Montermini,Laura, AU - Kim,Dae-Kyum, AU - Meehan,Brian, AU - Roth,Frederick P, AU - Rak,Janusz, Y1 - 2018/07/13/ PY - 2018/01/30/received PY - 2018/06/16/revised PY - 2018/7/15/pubmed PY - 2019/5/7/medline PY - 2018/7/15/entrez KW - Cancer Biology KW - EGFRvIII KW - Exosomes KW - Glioblastoma KW - Oncogenes KW - Secretome KW - extracellular vesicles KW - heterogeneity KW - invasion KW - nano-flow cytometry SP - 1948 EP - 1964 JF - Molecular & cellular proteomics : MCP JO - Mol. Cell Proteomics VL - 17 IS - 10 N2 - Glioblastoma multiforme (GBM) is a highly aggressive and heterogeneous form of primary brain tumors, driven by a complex repertoire of oncogenic alterations, including the constitutively active epidermal growth factor receptor (EGFRvIII). EGFRvIII impacts both cell-intrinsic and non-cell autonomous aspects of GBM progression, including cell invasion, angiogenesis and modulation of the tumor microenvironment. This is, at least in part, attributable to the release and intercellular trafficking of extracellular vesicles (EVs), heterogeneous membrane structures containing multiple bioactive macromolecules. Here we analyzed the impact of EGFRvIII on the profile of glioma EVs using isogenic tumor cell lines, in which this oncogene exhibits a strong transforming activity. We observed that EGFRvIII expression alters the expression of EV-regulating genes (vesiculome) and EV properties, including their protein composition. Using mass spectrometry, quantitative proteomic analysis and Gene Ontology terms filters, we observed that EVs released by EGFRvIII-transformed cells were enriched for extracellular exosome and focal adhesion related proteins. Among them, we validated the association of pro-invasive proteins (CD44, BSG, CD151) with EVs of EGFRvIII expressing glioma cells, and downregulation of exosomal markers (CD81 and CD82) relative to EVs of EGFRvIII-negative cells. Nano-flow cytometry revealed that the EV output from individual glioma cell lines was highly heterogeneous, such that only a fraction of vesicles contained specific proteins (including EGFRvIII). Notably, cells expressing EGFRvIII released EVs double positive for CD44/BSG, and these proteins also colocalized in cellular filopodia. We also detected the expression of homophilic adhesion molecules and increased homologous EV uptake by EGFRvIII-positive glioma cells. These results suggest that oncogenic EGFRvIII reprograms the proteome and uptake of GBM-related EVs, a notion with considerable implications for their biological activity and properties relevant for the development of EV-based cancer biomarkers. SN - 1535-9484 UR - https://www.unboundmedicine.com/medline/citation/30006486/The_Impact_of_Oncogenic_EGFRvIII_on_the_Proteome_of_Extracellular_Vesicles_Released_from_Glioblastoma_Cells_ L2 - http://www.mcponline.org/cgi/pmidlookup?view=long&pmid=30006486 DB - PRIME DP - Unbound Medicine ER -