Here we generate fluorescence resonance energy transfer biosensors for guanine exchange factors (GEFs) by inserting a fluorescent protein pair in a structural 'hinge' common to many GEFs. Fluorescent biosensors can map the activation of signaling molecules in space and time, but it has not been possible to quantify how different activation events affect one another or contribute to a specific cell behavior. By imaging the GEF biosensors in the same cells as red-shifted biosensors of Rho GTPases, we can apply partial correlation analysis to parse out the extent to which each GEF contributes to the activation of a specific GTPase in regulating cell movement. Through analysis of spontaneous cell protrusion events, we identify when and where the GEF Asef regulates the GTPases Cdc42 and Rac1 to control cell edge dynamics. This approach exemplifies a powerful means to elucidate the real-time connectivity of signal transduction networks.

中文翻译：

---

多重GTPase和GEF生物传感器成像可实现网络连接分析。

在这里，我们通过将荧光蛋白对插入许多GEF共有的结构“铰链”中，生成鸟嘌呤交换因子（GEF）的荧光共振能量转移生物传感器。荧光生物传感器可以绘制信号分子在空间和时间上的激活图，但无法量化不同的激活事件如何相互影响或促进特定的细胞行为。通过在与Rho GTPases的红移生物传感器相同的细胞中对GEF生物传感器进行成像，我们可以应用部分相关分析来分析每个GEF在调节细胞运动中对特定GTPase的激活贡献的程度。通过分析自发的细胞突起事件，我们可以确定GEF Asef在何时何地调节GTPases Cdc42和Rac1来控制细胞边缘动力学。