Role of Protein Methylation in Cell Mitosis and Glioblastoma

组蛋白和非组蛋白上的甲基化是一种常见而重要的翻译后修饰， 在调节癌症致瘤性中控制蛋白质功能和活性的修饰（PTM）， 对治疗的反应。在本计画中，我们将探讨蛋白质精氨酸（R）甲基化的机制 通过阐明蛋白质精氨酸甲基转移酶6（PRMT 6）对细胞的作用来调节GBM致瘤性 有丝分裂通过特异性精氨酸甲基化的调节染色体凝聚1（RCC 1），从而 调节GBM表型和对治疗的应答。我们提供了新的证据，PRMT 6调节RCC 1 通过RCC 1在R214处甲基化的活性，R214是RCC 1激活Ran GT3所需的精氨酸残基 通过与染色体中的组蛋白结合，从而影响细胞有丝分裂过程、GBM致瘤性和 对治疗的反应。我们证明，酪蛋白激酶2（CK 2），一个普遍表达和组成型， 在癌症中重要的活性丝氨酸/苏氨酸激酶刺激PRMT 6甲基转移酶活性， 磷酸化PRMT 6蛋白的苏氨酸21。此外，基因耗竭或小分子靶向 PRMT 6、RCC 1或CK 2在体外和体内抑制GBM致瘤性，并通过以下方式增强抗肿瘤作用： 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：CK 2激活的PRMT 6诱导RCC 1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。 三个具体目标：1）确定PRMT 6诱导的RCC 1的R214 aDMA如何影响细胞有丝分裂过程 作为体外和体内GBM肿瘤生物学特性; 2）定义CK 2调节PRMT 6的机制 活性，以及CK 2相关的PRMT 6磷酸化，RCC 1功能，细胞有丝分裂， GBM肿瘤表型，以及与GBM患者结果的关联;以及3）确定PRMT 6和PRMT 7是否与GBM患者结果相关。 CK 2导向的联合治疗比相应的单一治疗更有效，以及这种联合治疗是否 联合治疗增强了RT和替莫唑胺（TMZ）治疗的抗肿瘤活性。在 完成这个项目，我们将学习如何PRMT 6-catayed aDMA的RCC 1形状的细胞有丝分裂，GBM 表型和对治疗的反应。我们提出的研究将是第一个解决PRMT活动的作用 在细胞有丝分裂、GBM肿瘤生物学和对治疗的反应中对RCC 1的非组蛋白aDMA的影响。这些知识， 反过来，将提供包括PRMT 6抑制和PRMT 7抑制的组合的潜在益处的明确指示。 CK 2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.