KSR and regulators of RAS signaling in C elegans

描述(由申请人提供)：RAS刺激的Raf/MEK/ERK激酶级联在正常动物发育和癌症中发挥关键作用。多个鲜为人知的机制确保了这个级联只有在它应该在何时何地才是活动的。越来越多的证据表明，支架对信号蛋白的分隔是这种调控的重要组成部分。我们的研究使用线虫作为遗传模型系统来识别Raf/MEK/ERK信号的新调控因子，并了解它们是如何发挥作用的。在之前的资助期间(04/01/00到3/31/05)，我们展示了支架KSR在MEK/ERK激活中的重要和广泛的作用，我们还证明了支架CNK促进Raf激活的特定步骤。我们还发现了许多其他保守的、但以前未被研究过的基因产物，它们与KSR-1合作，促进对这一信号级联的某些下游反应。在下一个资助阶段，我们将集中在几组这些基因产物上，以了解信号蛋白的区隔如何控制信号的传播和特异性。在目标1中，我们将测试KSR-1和KSR-2对映体之间的内在功能差异是否有助于组织特异性反应。在目标2中，我们将确定支架CNK-1及其结合伙伴KIC-1在Raf激活和定位中的作用。膜转位和内吞是Raf激活和失活的关键步骤，我们将检验CNK-1和KIC-1参与这些过程的模型。在目标3中，我们将确定保守的泛素结合(E2)酶在RAS信号和支架功能中的作用。我们将测试这个E2调节KSR或CNK支架或其他信号蛋白的运输和定位的模型，并确定它的作用机制。在目标4中，我们将继续对新的RAS途径调节因子进行正向遗传和基于RNAi的筛选，并选择那些最可能与KSR、CNK和/或我们的E2蛋白一起发挥功能的调节因子进行进一步研究。我们的研究有望揭示RAS途径调控的新方面，并阐明信号区隔的机制和生物学意义。RAS或RAF信号的异常是人类癌症最常见的原因之一。从长远来看，对Raf/MEK/ERK调控的详细了解将有助于合理设计治疗此类癌症的治疗方法。

DESCRIPTION (provided by applicant): The Ras-stimulated Raf/MEK/ERK kinase cascade plays critical roles in normal animal development and cancer. Multiple poorly understood mechanisms ensure that this cascade is active only when and where it should be. Increasing evidence suggests that compartmentalization of signaling proteins by scaffolds is an important component of such regulation. Our research uses C. elegans as a genetic model system to identify new regulators of Raf/MEK/ERK signaling and understand how they function. During the previous funding period (04/01/00 to 3/31/05), we demonstrated the essential and widespread role of the scaffold KSR in MEK/ERK activation, and we showed that the scaffold CNK promotes a specific step of Raf activation. We also identified multiple other conserved, but previously unstudied, gene products that cooperate with KSR-1 to promote certain downstream responses to this signaling cascade. In the next funding period, we will focus on several sets of these gene products to understand how compartmentalization of signaling proteins controls signal propagation and specificity. In Aim 1, we will test if inherent functional differences between the KSR-1 and KSR-2 paralogs contribute to tissue-specific responses. In Aim 2, we will determine the role of the scaffold CNK-1 and its binding partner KIC-1 in Raf activation and localization. Membrane translocation and endocytosis are key steps in Raf activation and de-activation, and we will test the model that CNK-1 and KIC-1 are involved in these processes. In Aim 3, we will determine the role of a conserved ubiquitin conjugating (E2) enzyme in Ras signaling and scaffold function. We will test the model that this E2 regulates trafficking and localization of the KSR or CNK scaffolds, or of other signaling proteins, and we will identify the mechanism by which it acts. In Aim 4, we will continue both forward genetic and RNAi-based screens for new Ras pathway regulators, and choose for further study those mostly likely to function with KSR, CNK and/or our E2 protein. Our studies are expected to reveal new aspects of Ras pathway regulation and clarify the mechanisms and biological relevance of signaling compartmentalization. Aberrant signaling by Ras or Raf is 1 of the most frequent causes of human cancers. In the long term, a detailed understanding of Raf/MEK/ERK regulation will allow the rational design of therapeutic approaches to treat such cancers.