Regulation and Functional Dissection of YAP in Mitosis

描述(申请人提供)：河马途径通过磷酸化和失活下游癌蛋白YAP(YAP)来控制细胞接触抑制、干细胞自我更新和肿瘤形成。PI和其他研究表明，YAP通过刺激细胞增殖和抑制细胞凋亡来促进肿瘤发生。YAP在许多类型的癌症中过度表达或过度激活。目前涉及YAP的研究主要集中在确定其在不同器官/组织中的整体致癌作用，以及其在与其他信号通路的串扰中的作用。然而，虽然这些研究为YAP的致癌特性提供了重要的见解，但YAP发挥致癌功能的潜在分子机制却知之甚少。该项目的长期目标是阐明Hippo-Yap信号通路在有丝分裂细胞周期控制和致癌转化中的调控机制，从而提供潜在的治疗靶点。我们的初步研究表明，在有丝分裂过程中，YAP在新的位点上被磷酸化，并以依赖于CDK1的方式激活。重要的是，有丝分裂磷酸化是YAP驱动的细胞转化所必需的。我们发现，在有丝分裂过程中，纺锤体检查点的激活需要YAP。此外，YAP的过度表达，但不是不可磷酸化的突变体，过度激活纺锤体检查点并导致有丝分裂缺陷。在这些初步研究的基础上，我们推测CDK1介导的YAP有丝分裂磷酸化在调节纺锤体检查点激活和随后的致癌转化中具有生物学意义。我们的中心假设将通过以下三个具体目标得到验证：目标1：确定有丝分裂过程中YAP调节/激活的分子机制；目标2：确定YAP及其磷酸化在有丝分裂进程、纺锤体检查点和非整倍体中的作用；目标3：确定YAP磷酸化在其靶点上的功能意义。这些研究的成功完成不仅将揭示YAP在有丝分裂和基因组不稳定中的新作用，而且还将揭示YAP驱动的肿瘤发生的机制。

DESCRIPTION (provided by applicant): The Hippo pathway controls cell contact inhibition, stem cell self -renewal, and tumorigenesis through phosphorylating and inactivating the downstream oncoprotein, yes-associated protein (YAP). The PI, along with others, has demonstrated that YAP promotes oncogenesis by stimulating cell proliferation and inhibiting apoptosis. YAP is overexpressed or hyperactivated in many types of cancers. Current studies involving YAP focus on determining its overall oncogenic role in various organs/tissues as well as its role in crosstalk with other signaling pathways. While these studies provide important insight into the oncogenic properties of YAP, however, the underlying molecular mechanisms through which YAP exerts its oncogenic function are poorly understood. The long-term goal of this project is to elucidate the regulatory mechanisms of the Hippo-YAP signaling pathway in mitotic cell-cycle control and oncogenic transformation, thus providing potential therapeutic targets. Our preliminary studies demonstrate that during mitosis YAP is phosphorylated on novel sites and activated in a CDK1-dependent manner. Importantly, mitotic phosphorylation is required for YAP-driven cellular transformation. We have found that YAP is required for the activation of the spindle checkpoint during mitosis. Furthermore, overexpression of YAP, but not of the non - phosphorylatable mutant, hyper-activates the spindle checkpoint and causes mitotic defects. Based on these preliminary studies, we hypothesize that CDK1-mediated mitotic phosphorylation of YAP is biologically significant in the regulation of the spindle checkpoint activation and subsequent oncogenic transformation. Our central hypothesis will be tested through the following three specific aims: Aim 1: Determine the molecular mechanism of YAP regulation/activation during mitosis; Aim 2: Determine the role of YAP and its phosphorylation in mitotic progression, the spindle checkpoint, and aneuploidy; Aim 3: Determine the functional significance of YAP phosphorylation on its targets. Successful completion of these studies will not only reveal novel roles of YAP in mitosis and genome instability, but will also shed light on the mechanisms involved in YAP-driven oncogenesis.