Abstract
We have reported previously that the expression of focal adhesion kinase (FAK) is elevated in glioblastomas and that expression of FAK promotes the proliferation of glioblastoma cells propagated in either soft agar or in the C.B.17 severe combined immunodeficiency (scid) mouse brain. We therefore determined the effect of FAK on cell cycle progression in these cells. We found that overexpression of wild-type FAK promoted exit from G(1) in monolayer cultures of glioblastoma cells, enhanced the expression of cyclins D1 and E while reducing the expression of p27(Kip1) and p21(Waf1), and enhanced the kinase activity of the cyclin D1-cyclin-dependent kinase-4 (cdk4) complex. Transfection of the monolayers with a FAK molecule in which the autophosphorylation site is mutated (FAK397F) inhibited exit from G(1) and reduced the expression of cyclins D1 and E while enhancing the expression of p27(Kip1) and p21(Waf1). Small interfering RNA (siRNA)-mediated down-regulation of cyclin D1 inhibited the enhancement of cell cycle progression observed on expression of wild-type FAK, whereas siRNA-mediated down-regulation of cyclin E had no effect. siRNA-mediated down-regulation of p27(Kip1) overcame the inhibition of cell cycle progression observed on expression of FAK397F, whereas down-regulation of p21(Waf1) had no effect. These results were confirmed in vivo in the scid mouse brain xenograft model in which propagation of glioblastoma cells expressing FAK397F resulted in a 50% inhibition of tumor growth and inhibited exit from G(1). Taken together, our results indicate that FAK promotes proliferation of glioblastoma cells by enhancing exit from G(1) through a mechanism that involves cyclin D1 and p27(Kip1).
Highlights
We have reported previously that the expression of focal adhesion kinase (FAK) is elevated in glioblastomas and that expression of FAK promotes the proliferation of glioblastoma cells propagated in either soft agar or in the C.B.17 severe combined immunodeficiency mouse brain
These results are consistent with those of Zhao et al [8], who found that serum starvation did not significantly alter the effect of FAK on cell cycle progression in NIH3T3 cells and indicate that FAK promotes exit from G1 in glioblastoma cells in monolayer culture, This conclusion was supported by the results obtained on analysis of BrdUrd labeling of cells cultured in complete medium, which showed that expression of wild-type FAK resulted in a 58% higher level of BrdUrd labeling than in cells that were not treated with doxycycline (p ϭ 0.001) (Fig. 3) and that expression of FAK397F resulted in a 70% reduction in BrdUrd labeling compared with the cells not treated with doxycycline (p Ͻ 0.001) (Fig. 3)
In this paper we demonstrate that FAK promotes cell cycle progression of glioblastoma cells when propagated in vitro as a monolayer and in vivo in the scid mouse brain by promoting exit from G1
Summary
We have reported previously that the expression of focal adhesion kinase (FAK) is elevated in glioblastomas and that expression of FAK promotes the proliferation of glioblastoma cells propagated in either soft agar or in the C.B.17 severe combined immunodeficiency (scid) mouse brain. We report that wild-type FAK promotes the cell cycle progression of glioblastoma cells propagated both in vitro as a monolayer and in vivo in the C.B.17 scid mouse brain.
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