Cell Cycle G2/M Pathway Modulated by Viral Protein R

描述(由申请人提供)：细胞从G2期的进展 细胞周期到有丝分裂是一个严格调控的过程，需要激活 它决定了所有真核细胞中有丝分裂的开始。 在裂殖酵母中，cdc2的活性是受调控的。 通过酪氨酸15(Tyr 15)在Cdc2上的磷酸化状态，即 在G2后期被Wee1和Mik1酪氨酸激酶磷酸化，并迅速 被CDC25磷酸酶去磷酸化以触发进入有丝分裂。Cdc25， Wee1和Mik1是两条特征明确的调控通路的靶标， DNA损伤和DNA复制检查点，两者都会导致细胞周期 抑制CDc2上Tyr15的磷酸化主要通过 HIV-1病毒蛋白R(VPR)诱导细胞G2期停滞 来自远亲真核生物的细胞，包括分裂酵母和人类。 与两条检查点通路类似，Vpr也在分裂中诱导G2停滞 酵母通过抑制Tyr15的磷酸化。但是，VPR不使用 检查点路径中的任何早期或后期步骤，取而代之的是它起作用 主要通过激活Wee1，蛋白磷酸酶2A(PP2A)和一种 CDC25抑制程度较小，提示VPR诱导G2期停滞 通过一种新的PP2A介导的调节途径。这项提议的目标是 使用分裂酵母作为模型系统来检验这一假说并阐明 这种新的G2/M调控途径。三个具体目标是：1)定义 VPR诱导G2期停滞时PP2A对Wee1和CDC25的调节作用 VPR与PP2A和酪蛋白激酶II的潜在蛋白质复合体的鉴定 Alpha(CKIIa)和定义这个复合体在VPR诱导的G2停滞中的作用，以及 3)鉴定四个多拷贝VPR特异性抑制子(Wos2和Wos2 Vsp24C8)/增强子(Rad25和Sum1)及其在Vp诱导的G2期停滞中的作用。 这些拟议的研究将为这个新的小组提供一个全面的框架 循环控制途径，并提供对这一新技术的基本方面的见解 真核细胞的G2/M监测系统。

DESCRIPTION (provided by applicant): Progression of cells from G2 phase of the cell cycle to mitosis is a tightly regulated process that requires activation of the Cdc2 kinase, which determines onset of mitosis in all eukaryotic cells. In fission yeast (Schizosaccharomyces pombe), the activity of Cdc2 is regulated by the phosphorylation status of tyrosine 15 (Tyr 15) on Cdc2, which is phosphorylated by Wee1 and Mik1 tyrosine kinases during late G2 and is rapidly dephosphorylated by the Cdc25 phosphatase to trigger entry into mitosis. Cdc25, Wee1 and Mik1 are the targets of two well-characterized regulatory pathways, the DNA damage and DNA replication checkpoints, which both cause cell cycle arrest by inhibitory phosphorylation of Tyr15 on Cdc2 primarily through inhibition of Cdc25.The viral protein R (Vpr) of HIV-1 induces G2 arrest in cells from distantly related eukaryotes, including fission yeast and human. Similar to the two checkpoint pathways, Vpr also induces G2 arrest in fission yeast through inhibitory phosphorylation of Tyr15. However, Vpr does not use any of the early or late steps in the checkpoint pathways, and instead it acts primarily through activation of Wee1, protein phosphatase 2A (PP2A) and to a lesser extent inhibition of Cdc25, suggesting that Vpr induces G2 arrest through a novel PP2A-mediated regulatory pathway. The goal of this proposal is to use fission yeast as a model system to test this hypothesis and to elucidate this new G2/M regulatory pathway. The three specific aims are to 1) define the regulatory effects of PP2A on Wee1 and Cdc25 when Vpr induces G2 arrest; 2) characterize a potential protein complex of Vpr with PP2A and casein kinase II alpha (CKIIa) and define the role of this complex in Vpr-induced G2 arrest, and 3) characterize four multicopy Vpr-specific suppressors (Wos2 and Vsp24C8)/enhancers (Rad25 and Sum1) and their role in Vprinduced G2 arrest. These proposed studies will provide a comprehensive framework for this new cell cycle control pathway and provide insights into fundamental aspects of this new G2/M surveillance system of eukaryotic cells.