Role of MLK3 in beta catenin signaling in prostate cancer

描述（由申请人提供）：在任何生物体中，细胞存活和细胞死亡途径之间的复杂平衡维持着正常的生长和体内平衡，失去这种平衡可能导致严重的病理异常。了解控制这些过程的调节机制很重要，可能使我们能够操纵它们进行治疗干预。蛋白激酶在调节细胞过程中的作用已被充分证明，其失调可导致各种病理疾病。我们的合作者早期的研究表明，一种名为混合谱系激酶3 （MLK3）的激酶的激活可以诱导癌细胞凋亡，这表明MLK3激动剂作为化疗药物的可能性。我们最近的研究表明，1)MLK3在前列腺癌细胞中表达，2)MLK3过表达导致异位p-catenin的显著稳定，这需要MLK3激酶的活性，2)SiRNA介导的MLK3敲低可降低内源性p-catenin水平，3)MLK3与p-catenin协同抑制NFicB活性，4)MLK3共表达可增加p-catenin介导的前列腺癌细胞凋亡。本研究的主要目的是确定p-catenin是否参与MLK3介导的前列腺细胞凋亡，并阐明其参与的途径。这是基于p-catenin过表达可独立于TCF/LEF转激活途径导致细胞凋亡的事实。事实上，MLK3介导的p-catenin稳定化抑制了传统的TCF/LEF介导和NFxB介导的转录。由于NFxB是前列腺癌细胞的主要存活途径，我们假设MLK3介导的p-catenin信号通路抑制NFxB途径导致细胞凋亡。为了实现我们的目标，我们提出了两个目标：(1)确定MLK3/p-catenin信号通路在前列腺癌细胞中的激活作用；(2)阐明MLK3介导的前列腺癌细胞中p-catenin信号通路。这些研究将利用各种技术进行，包括SiRNA，细胞凋亡测定，荧光素酶测定，免疫沉淀，Western Blotting，免疫组织化学。由于目前还没有关于MLK3诱导细胞凋亡的机制的信息，这些研究有望揭示这一非常新颖的途径，并可用于未来基于MLK3激活治疗前列腺肿瘤的药物的开发。

DESCRIPTION (provided by applicant): An intricate balance between cell survival and cell death pathways maintains normal growth and homeostasis in any organism, loss of which can result in serious pathological abnormalities. Understanding of the regulatory mechanisms controlling these processes is important and might allow us to manipulate them for therapeutic interventions. The role of protein kinases in regulating cellular processes has been well documented, dysregulation of which can lead to various pathological disorders. Earlier studies by our collaborators demonstrated that activation of a kinase known as mixed lineage kinase 3 (MLK3) can induce apoptosis in cancer cells, indicating the possibility of utilizing MLK3 agonists as chemotherapeutic agents. Our recent studies showed that 1) MLK3 is expressed in prostate cancer cells, 2) overexpression of MLK3 results in a significant stabilization of ectopic p-catenin, which requires MLK3 kinase activity, 2) SiRNA mediated knockdown of MLK3 reduces endogenous p-catenin levels, 3) MLK3 synergizes with p-catenin to inhibit NFicB activity, and 4) coexpression of MLK3 increases p-catenin mediated apoptosis in prostate cancer cells. The major goal of this proposal is to determine whether p-catenin is involved in MLK3 mediated prostate cell apoptosis, and to elucidate the pathway involved. This is based on the fact that overexpression of p-catenin can lead to apoptosis independent of TCF/LEF transactivation pathway. In fact, MLK3 mediated stabilization of p-catenin inhibits conventional TCF/LEF mediated and NFxB mediated transcription. Since NFxB is a major survival pathway in prostate cancer cells, we hypothesize that MLK3 mediated signaling to p-catenin inhibits NFxB pathway leading to apoptosis. To achieve our goals two aims are proposed: (1) To determine the effect of activation of MLK3/p-catenin signaling in prostate cancer cells, (2) To elucidate the pathway of MLK3 mediated p-catenin signaling in prostate cancer cells. These studies will be performed utilizing various techniques including SiRNA, apoptosis assays, luciferase assay, immunoprecipitation, Western Blotting, immunohistochemistry. Since there is no information available regarding the mechanism how MLK3 induces apoptosis, these studies are expected to shed light into this very novel pathway, which can be utilized towards the future development of drugs for the treatment of prostate tumors based on MLK3 activation.