Cross talk between oncogene and ant-oncogene via the Crk oncogene product

Crk癌基因产物激活Ras和Rap1。Ras是一种众所周知的致癌基因，而Rap1可以拮抗Ras依赖性的肿瘤发生。为了了解Crk激活癌基因和抗癌基因的机制，我们开发了一种基于fret的单分子探针，可以监测活细胞中Crk的激活。使用这种探针，我们看到了egf刺激细胞中Crk的快速激活。然而，由于探针的快速扩散，我们无法确定细胞中Crk的激活是从哪里开始的。为了克服这一缺陷，我们将Ki-Ras蛋白的CAAX盒与探针融合，期望探针能锚定在质膜上。事实上，通过使用这种caax融合探针，我们发现Crk从细胞的外周区域被激活。此外，我们构建了一种重组腺病毒来表达探针，使我们能够在多种细胞中快速有效地表达探针。因此，新的Crk活性探针的开发，可视化了Crk癌基因产物在活细胞中的时空调控，为我们理解Crk癌基因产物在细胞生长和转化中的调控作用提供了基础。

Crk oncogene product activates Ras and Rap1. Ras is a well-known oncogene, whereas Rap1 antagonize Ras-dependent oncogenesis. To understand the mechanism by which Crk activates both oncogene and anti-oncogene, we developed a FRET-based single-molecule probe that can monitor the activation of Crk in a living cell. Using this probe, we visualized rapid Crk activation in an EGF-stimulated cell. However, with the prototype probe, we could not determine where in the cell the activation of Crk starts, due to rapid diffusion of the probe. To overcome this flaw, we fused CAAX box of Ki-Ras protein to the probe, expecting the anchoring of probe to the plasma membrane. Indeed, by the use of this CAAX-fused probe, we found that Crk was activated from the peripheral region of the cell. Furthermore, we generated a recombinant adenovirus for the expression of the probe, which enabled us rapid and efficient expression of the probe in a variety of cells. Thus, the development of new probe for the Crk activity visualized the spatio temporal regulation of Crk oncogene products in living cells and promised us for the understanding of the role of Crk oncogene products in the regulation of cell growth and transformation.