Role of Protein Methylation in Cell Mitosis and Glioblastoma

组蛋白和非组蛋白的甲基化是一种常见而重要的翻译后 修饰(PTM)，控制蛋白质的功能和活性，调节癌症的致瘤性和 对治疗的反应。在这个项目中，我们将研究蛋白质精氨酸(R)甲基化的机制 蛋白精氨酸甲基转移酶6(PRMT6)对基底膜致瘤性的调控 通过染色体凝聚调节因子1(RCC1)的精氨酸甲基化进行有丝分裂，从而 调节GBM表型和对治疗的反应。我们提供了PRMT6调控RCC1的新证据 RCC1通过R214甲基化的活性，RCC1激活RAN GTP酶所需的精氨酸残基 通过与染色体上的组蛋白结合，从而影响细胞有丝分裂过程，基底膜的致瘤性和 对治疗的反应。我们证明了酪蛋白激酶2(CK2)是一种普遍表达的结构性蛋白。 在癌症中重要的活性丝氨酸/苏氨酸激酶通过以下方式刺激PRMT6甲基转移酶活性 将PRMT6蛋白的苏氨酸21磷酸化。此外，基因枯竭或小分子靶向 PRMT6、RCC1或CK2抑制GBM的体内外致瘤性并增强其抗肿瘤作用 放射治疗(RT)。这些强有力的科学前提为这一总体假设提供了基础 建议：CK2激活的PRMT6诱导RCC1的不对称二甲基化(ADMA)，从而调节细胞 有丝分裂、GBM致瘤性以及GBM对RT的反应。我们将在以下背景下讨论这一假设 3个具体目标：1)确定PRMT6诱导的RCC1的R214 ADMA如何影响细胞有丝分裂过程 2)明确CK2调控PRMT6的机制 活性，以及CK2相关的PRMT6磷酸化、RCC1功能、细胞有丝分裂、 GBM肿瘤表型及其与GBM患者预后的关系；以及3)决定PRMT6和PRMT6和 CK2导向的联合治疗比相应的单一治疗更有效，以及是否如此 联合治疗可增强RT和替莫唑胺(TMZ)治疗的抗肿瘤活性。在 这个项目的完成，我们将学习PRMT6-催化RCC1的ADMA如何形成细胞有丝分裂，GBM 表型和对治疗的反应。我们提议的研究将是第一个解决PRMT活动的作用的研究 RCC1的非组蛋白ADMA在细胞有丝分裂、基底膜肿瘤生物学和治疗反应中的作用。这种知识， 反过来，将提供明确的指示，包括抑制PRMT6和PRMT6的组合的潜在好处 CK2联合RT和TMZ作为GBM患者治疗策略的一部分。

Methylation on histone and non-histone proteins is a common and important post-translational modification (PTM) that controls protein functions and activities in modulating cancer tumorigenicity and responses to therapies. In this project, we will investigate the mechanism for protein arginine (R) methylation regulation of GBM tumorigenicity by elucidating the role of protein arginine methyltransferase 6 (PRMT6) on cell mitosis through specific arginine methylation of regulator of chromosome condensation 1 (RCC1), thereby regulating GBM phenotype and responses to therapies. We provide novel evidence that PRMT6 regulates RCC1 activity through methylation of RCC1 at R214, the arginine residue required for RCC1 activation of Ran GTPase through association with histones in chromosome, thereby affecting cell mitotic process, GBM tumorigenicity and responses to therapies. We demonstrate that casein kinase 2 (CK2), a ubiquitously expressed and constitutively active serine/threonine kinase that is important in cancers stimulates PRMT6 methyltransferase activity by phosphorylating threonine 21 of PRMT6 protein. Additionally, genetic depletions or small molecule targeting of PRMT6, RCC1 or CK2 inhibited GBM tumorigenicity in vitro and in vivo, and enhanced anti-tumor effects by radiation therapy (RT). These strong scientific premises provide the basis for the overarching hypothesis of this proposal: CK2-activated PRMT6 induces asymmetric dimethylation (aDMA) of RCC1, thereby regulating cell mitosis, GBM tumorigenicity as well as GBM responses to RT. We will address this hypothesis in the context of three specific aims: 1) Determine how PRMT6-induced R214 aDMA of RCC1 affects cell mitotic process as well as GBM tumor biologic properties in vitro and in vivo; 2) Define mechanisms by which CK2 regulates PRMT6 activity, and the relationships between CK2-associated phosphorylation of PRMT6, RCC1 function, cell mitosis, GBM tumor phenotypes, and association with GBM patient outcome; and 3) Determine whether PRMT6 and CK2 directed combination therapy is more effective than corresponding monotherapies, and whether such combination therapy enhances the anti-tumor activity of RT and temozolomide (TMZ) treatment. At the completion of this project, we will learn how PRMT6-catayzed aDMA of RCC1 shapes of cell mitosis, GBM phenotype and response to therapy. Our proposed studies will be the first to address the role of PRMT activity on non-histone aDMA of RCC1 in cell mitosis, GBM tumor biology and responses to therapies. This knowledge, in turn, will provide clear indication of the potential benefit of including combination of inhibition of PRMT6 and CK2 with RT and TMZ as part of treatment strategy for patient with GBM.