Downregulation of FIP200 induces apoptosis of glioblastoma cells and microvascular endothelial cells by enhancing Pyk2 activity

• Published in: PloS one 2011-05, Vol.6 (5), p.e19629-e19629
• Main Authors: Wang, Dongyan, Olman, Mitchell A, Stewart, Jr, Jerry, Tipps, Russell, Huang, Ping, Sanders, Paul W, Toline, Eric, Prayson, Richard A, Lee, Jeongwu, Weil, Robert J, Palmer, Cheryl A, Gillespie, G Yancey, Liu, Wei Michael, Pieper, Russell O, Guan, Jun-Lin, Gladson, Candece L
• Format: Article
• Language: English
• Subjects: Adhesion; Angiogenesis; Apoptosis; Apoptosis - drug effects; Astrocytes; Biology; Brain; Brain - blood supply; Brain - pathology; Brain cancer; Brain tumors; Cell adhesion & migration; Cell culture; Cell cycle; Cell growth; Cell survival; Competition; Cytokines; Down-Regulation - genetics; Endothelial cells; Endothelial Cells - cytology; Endothelial Cells - enzymology; Endothelium; Focal adhesion kinase; Focal Adhesion Kinase 2 - analysis; Focal Adhesion Kinase 2 - metabolism; Fusion protein; Gene expression; Glial cells; Glioblastoma; Glioblastoma - enzymology; Glioblastoma - pathology; Glioblastoma cells; Glioblastomas; Humans; Kinases; Ligands; Medicine; Microcirculation; Microvasculature; Nervous system; Neuronal-glial interactions; Neurons; Neurons - enzymology; Neuropathology; Neurosurgery; Pathology; Proline; Proline-rich tyrosine kinase 2; Protein Binding; Protein-tyrosine kinase; Protein-Tyrosine Kinases - analysis; Protein-Tyrosine Kinases - genetics; Protein-Tyrosine Kinases - metabolism; Proteins; Ribonucleic acid; RNA; RNA, Small Interfering - pharmacology; Signal transduction; siRNA; Stem cells; Tissues; Transcription factors; Tumor cell lines; Tumor cells; Tumor Cells, Cultured; Tumors; Tyrosine
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0019629

The expression of focal adhesion kinase family interacting protein of 200-kDa (FIP200) in normal brain is limited to some neurons and glial cells. On immunohistochemical analysis of biopsies of glioblastoma tumors, we detected FIP200 in the tumor cells, tumor-associated endothelial cells, and occasional glial cells. Human glioblastoma tumor cell lines and immortalized human astrocytes cultured in complete media also expressed FIP200 as did primary human brain microvessel endothelial cells (MvEC), which proliferate in culture and resemble reactive endothelial cells. Downregulation of endogenous expression of FIP200 using small interfering RNA resulted in induction of apoptosis in the human glioblastoma tumor cells, immortalized human astrocytes, and primary human brain MvEC. It has been shown by other investigators using cells from other tissues that FIP200 can interact directly with, and inhibit, proline-rich tyrosine kinase 2 (Pyk2) and focal adhesion kinase (FAK). In the human glioblastoma tumor cells, immortalized human astrocytes, and primary human brain MvEC, we found that downregulation of FIP200 increased the activity of Pyk2 without increasing its expression, but did not affect the activity or expression of FAK. Coimmunoprecipitation and colocalization studies indicated that the endogenous FIP200 was largely associated with Pyk2, rather than FAK, in the glioblastoma tumor cells and brain MvEC. Moreover, the pro-apoptotic effect of FIP200 downregulation was inhibited significantly by a TAT-Pyk2-fusion protein containing the Pyk2 autophosphorylation site in these cells. In summary, downregulation of endogenous FIP200 protein in glioblastoma tumor cells, astrocytes, and brain MvECs promotes apoptosis, most likely due to the removal of a direct interaction of FIP200 with Pyk2 that inhibits Pyk2 activation, suggesting that FIP200 expression may be required for the survival of all three cell types found in glioblastoma tumors.