Mechanisms of effector activation by the RAS oncogene

Ras癌基因经常与人类癌症相关，在实验系统中，Ras的活化形式具有强大的转化能力。Ras蛋白通过多种效应物控制多种信号通路。这些效应物被Ras激活的机制尚不清楚。我们研究了Ras与其效应/调节因子p120 GAP之间的相互作用。我们发现Ras可以调节p120 GAP的催化区和调控区之间的分子间相互作用。我们也在研究Ras蛋白激活Raf激酶和Nore1类效应物的分子机制。这些研究的最终目的是允许设计高度特异性的ras介导转化的小分子抑制剂。我们发现Ras通过结合Raf上的两个不同位点激活Raf。这两种结合相互作用对于产生完全激活的Raf分子至关重要。我们已经确定了第二个Ras结合域内的特定残基，这些残基对Ras相互作用至关重要。此外，我们还发现14-3-3和磷脂酰丝氨酸的相互作用是ras介导的Raf激活的辅助因子。我们现在有证据表明，与p120 GAP一样，Ras可以将Raf的c端激酶结构域从Raf的n端调控结构域的抑制性分子内结合接触中释放出来。这种相互作用似乎是由14-3-3介导的，需要脂质辅助因子（PS）的结合才能完全表现出来。此外，抑制分子内相互作用的释放似乎允许相同的结合位点调节活性所必需的激酶结构域二聚化事件。我们目前正在研究新的Ras效应物Nore1，以确定Ras是否也能破坏分子内结合事件的稳定性或影响二聚化。

Ras oncogenes are frequently associated with human cancer and activated forms of ras are powerfully transforming in experimental systems. Ras proteins control multiple signaling pathways via multiple effectors. The mechanisms by which those effectors are activated by Ras remain unclear. We have investigated the interaction between Ras and its effector/regulator p120 GAP. We have found that Ras serves to modulate an interamolecular interaction between the catalytic and the regulatory regions of p120 GAP. We are also investigating the molecular mechanisms by which the Ras proteins activate the Raf kinase and Nore1 class of effectors. The ultimate aim of these studies is to allow the design of highly specific small molecule inhibitors of Ras-mediated transformation.We have found that Ras activates Raf by binding to two distinct sites on Raf. Both binding interactions are crtitical in generating a fully activated Raf molecule. We have identified specific residues within the second Ras binding domain that are essential for Ras interaction. Moreover, we have characterized the interplay of 14-3-3 and phosphatidylserine as co-factors in the Ras-mediated activation of Raf. We now have evidence that, as with p120 GAP, Ras serves to release the C-terminal kinase domain of Raf from inhibitory, intramolecular binding contacts in the N-terminal, regulatory domain of Raf. This interaction appears to mediated by 14-3-3 and requires the binding of a lipid co-factor (PS) for full manifestation. Moreover, it appears that the release of the inhibitory intramolecular interaction allows the same binding sites to modulate a kinase domain dimerization event essential for activity.We are currently studying the novel Ras effector Nore1 to determine if Ras also acts to destabilize intramolecular binding events or to influence dimerization.