Membrane Proteome Analysis of Glioblastoma Cell Invasion

• Published in: Journal of neuropathology and experimental neurology Vol. 74; no. 5; pp. 425 - 441
• Main Authors: Mallawaaratchy, Duthika M., Buckland, Michael E., McDonald, Kerrie L., Li, Cheryl C.Y., Ly, Linda, Sykes, Erin K., Christopherson, Richard I., Kaufman, Kimberley L.
• Format: Journal Article
• Language: English
• Published: England American Association of Neuropathologists, Inc 01.05.2015; by American Association of Neuropathologists, Inc
• Subjects: Annexin A1 - genetics; Annexin A1 - metabolism; Antigens, CD - genetics; Antigens, CD - metabolism; Brain Neoplasms - pathology; Cell Line, Tumor; Databases, Protein - statistics & numerical data; Gene Expression Regulation, Neoplastic; Glioblastoma - pathology; Humans; Male; Membrane Proteins - genetics; Membrane Proteins - metabolism; Middle Aged; Neoplasm Invasiveness - pathology; Proteomics - methods; RNA, Small Interfering - genetics; RNA, Small Interfering - metabolism; Tandem Mass Spectrometry; Transfection; CD97; Epithelial-mesenchymal transition; Membrane protein; Cell invasion; Glioblastoma; Proteomics; Invadopodia; Integrin α5
• ISSN: 0022-3069; 1554-6578; 1554-6578
• DOI: 10.1097/NEN.0000000000000187

Glioblastoma multiforme (GBM) tumor invasion is facilitated by cell migration and degradation of the extracellular matrix. Invadopodia are actin-rich structures that protrude from the plasma membrane in direct contact with the extracellular matrix and are proposed to participate in epithelial-mesenchymal transition. We characterized the invasiveness of 9 established GBM cell lines using an invadopodia assay and performed quantitative mass spectrometry–based proteomic analyses on enriched membrane fractions. All GBM cells produced invadopodia, with a 65% difference between the most invasive cell line (U87MG) and the least invasive cell line (LN229) (p = 0.0001). Overall, 1,141 proteins were identified in the GBM membrane proteome; the levels of 49 proteins correlated with cell invasiveness. Ingenuity Pathway Analysis predicted activation “cell movement” (z-score = 2.608, p = 3.94E−04) in more invasive cells and generated a network of invasion-associated proteins with direct links to key regulators of invadopodia formation. Gene expression data relating to the invasion-associated proteins ITGA5 (integrin α5), CD97, and ANXA1 (annexin A1) showed prognostic significance in independent GBM cohorts. Fluorescence microscopy demonstrated ITGA5, CD97, and ANXA1 localization in invadopodia assays, and small interfering RNA knockdown of ITGA5 reduced invadopodia formation in U87MG cells. Thus, invasion-associated proteins, including ITGA5, may prove to be useful anti-invasive targets; volociximab, a therapeutic antibody against integrin α5β1, may be useful for treatment of patients with GBM.