PHIP drives glioblastoma motility and invasion by regulating the focal adhesion complex.

Abstract

The invasive behavior of glioblastoma is essential to its aggressive potential. Here, we show that pleckstrin homology domain interacting protein (PHIP), acting through effects on the force transduction layer of the focal adhesion complex, drives glioblastoma motility and invasion. Immunofluorescence analysis localized PHIP to the leading edge of glioblastoma cells, together with several focal adhesion proteins: vinculin (VCL), talin 1 (TLN1), integrin beta 1 (ITGB1), as well as phosphorylated forms of paxillin (pPXN) and focal adhesion kinase (pFAK). Confocal microscopy specifically localized PHIP to the force transduction layer, together with TLN1 and VCL. Immunoprecipitation revealed a physical interaction between PHIP and VCL. Targeted suppression of PHIP resulted in significant down-regulation of these focal adhesion proteins, along with zyxin (ZYX), and produced profoundly disorganized stress fibers. Live-cell imaging of glioblastoma cells overexpressing a ZYX-GFP construct demonstrated a role for PHIP in regulating focal adhesion dynamics. PHIP silencing significantly suppressed the migratory and invasive capacity of glioblastoma cells, partially restored following TLN1 or ZYX cDNA overexpression. PHIP knockdown produced substantial suppression of tumor growth upon intracranial implantation, as well as significantly reduced microvessel density and secreted VEGF levels. PHIP copy number was elevated in the classical glioblastoma subtype and correlated with elevated EGFR levels. These results demonstrate PHIP's role in regulating the actin cytoskeleton, focal adhesion dynamics, and tumor cell motility, and identify PHIP as a key driver of glioblastoma migration and invasion.

• Authors: David de Semir, Vladimir Bezrookove, Mehdi Nosrati, Kara R Scanlon, Eric Singer, Jonathon Judkins, Christopher Rieken, Clayton Wu, Julia Shen, Christina Schmudermayer, Altaf A Dar, James R Miller 3rd, Charles Cobbs, Garret Yount, Pierre-Yves Desprez, Robert J Debs, Nathan Salomonis, Sean McAllister, James E Cleaver, Liliana Soroceanu, Mohammed Kashani-Sabet
• Journal: Proceedings of the National Academy of Sciences of the United States of America (Proc Natl Acad Sci U S A) Vol. 117 Issue 16 Pg. 9064-9073 (04 21 2020) ISSN: 1091-6490 [Electronic] United States
• PMID: 32273388 (Publication Type: Journal Article, Research Support, N.I.H., Extramural, Research Support, Non-U.S. Gov't, Video-Audio Media)

Chemical References

• Intracellular Signaling Peptides and Proteins
• PHIP protein, human
• VCL protein, human
• Vinculin

Topics

• Actin Cytoskeleton (metabolism)
• Animals
• Brain (pathology)
• Brain Neoplasms (blood supply, genetics, pathology)
• Cell Adhesion (genetics)
• Cell Line, Tumor
• Cell Movement (genetics)
• Cohort Studies
• Disease Progression
• Female
• Focal Adhesions (pathology)
• Gene Dosage
• Gene Expression Regulation, Neoplastic
• Gene Knockdown Techniques
• Glioblastoma (blood supply, genetics, pathology)
• Humans
• Intracellular Signaling Peptides and Proteins (genetics, metabolism)
• Intravital Microscopy
• Mice
• Microscopy, Confocal
• Neoplasm Invasiveness (genetics, pathology)
• Neovascularization, Pathologic (genetics, pathology)
• Time-Lapse Imaging
• Vinculin (metabolism)
• Xenograft Model Antitumor Assays