Department of Molecular Cell Biology, University of Groningen, 9751 NN Haren, The Netherlands.
Mol Biol Cell. 2010 Mar 15;21(6):936-45. doi: 10.1091/mbc.e09-03-0177. Epub 2010 Jan 20.
GbpD, a Dictyostelium discoideum guanine exchange factor specific for Rap1, has been implicated in adhesion, cell polarity, and chemotaxis. Cells overexpressing GbpD are flat, exhibit strongly increased cell-substrate attachment, and extend many bifurcated and lateral pseudopodia. Phg2, a serine/threonine-specific kinase, mediates Rap1-regulated cell-substrate adhesion, but not cell polarity or chemotaxis. In this study we demonstrate that overexpression of GbpD in pi3k1/2-null cells does not induce the adhesion and cell morphology phenotype. Furthermore we show that Rap1 directly binds to the Ras binding domain of PI3K, and overexpression of GbpD leads to strongly enhanced PIP3 levels. Consistently, upon overexpression of the PIP3-degradating enzyme PTEN in GbpD-overexpressing cells, the strong adhesion and cell morphology phenotype is largely lost. These results indicate that a GbpD/Rap/PI3K pathway helps control pseudopod formation and cell polarity. As in Rap-regulated pseudopod formation in Dictyostelium, mammalian Rap and PI3K are essential for determining neuronal polarity, suggesting that the Rap/PI3K pathway is a conserved module regulating the establishment of cell polarity.
GbpD，一种专用于 Rap1 的盘基网柄菌鸟嘌呤交换因子，与黏附、细胞极性和趋化性有关。过表达 GbpD 的细胞是平的，表现出强烈增加的细胞-基质附着，并延伸许多分叉和侧生伪足。Phg2，一种丝氨酸/苏氨酸特异性激酶，介导 Rap1 调节的细胞-基质黏附，但不调节细胞极性或趋化性。在这项研究中，我们证明了在 pi3k1/2-/-细胞中过表达 GbpD 不会诱导黏附和细胞形态表型。此外，我们还表明 Rap1 直接结合到 PI3K 的 Ras 结合域，并且过表达 GbpD 导致强烈增强的 PIP3 水平。一致地，在过表达 GbpD 过表达细胞中的 PIP3 降解酶 PTEN 后，强烈的黏附和细胞形态表型大部分丢失。这些结果表明，GbpD/Rap/PI3K 途径有助于控制伪足形成和细胞极性。与盘基网柄菌中 Rap 调节的伪足形成一样，哺乳动物 Rap 和 PI3K 对于确定神经元极性是必不可少的，这表明 Rap/PI3K 途径是一个保守的模块，调节细胞极性的建立。
