DYRK1A signaling in control of cell growth, proliferation and DNA damage repair

描述（由申请人提供）：细胞阻止增殖和进入g1 -阻滞或G0（静止）状态的能力对肿瘤抑制至关重要。视网膜母细胞瘤（Retinoblastoma， RB）家族蛋白，包括p130，在哺乳动物细胞中介导G0/G1阻滞的机制尚不完全清楚。在进入G0/G1后，p130被招募到dna结合的DREAM蛋白复合物（DP、rb样蛋白、E2F和MuvB核心）中。5个蛋白（LIN9、LIN37、LIN52、LIN54和RBBP4）的MuvB核心在转录控制中起双重作用。当在G0/G1中与p130结合时，MuvB核心形成DREAM复合物并抑制800多种细胞周期依赖基因。当从DREAM释放时，MuvB核心与BMYB结合以激活有丝分裂基因。DYRK1A蛋白激酶通过磷酸化LIN52亚基在促进DREAM复合物的组装中发挥关键作用，而LIN52亚基是p130和MuvB核心结合所必需的。抑制DYRK1A足以阻断DREAM复合体的组装并克服G0/G1阻滞。DYRK1A基因被定位到唐氏综合征（DS）的一个关键区域，其由于21三体而导致的上调有助于唐氏综合征患者的精神障碍。建立DYRK1A在肿瘤抑制中的作用是很重要的，因为它作为改善DS患者认知功能的治疗靶点具有潜在的价值。先前的研究揭示了DYRK1A在RB家族介导的G0/G1阻滞以及致癌ras诱导的衰老中发挥重要作用。此外，生化数据表明DYRK1A在Hippo肿瘤抑制通路中位于LATS2激酶的下游。我们的初步分析发现，DYRK1A基因在高级别卵巢癌中经常发生拷贝数丢失，这表明DYRK1A失活参与了卵巢癌的发病机制。我们假设DYRK1A在卵巢癌中由于基因缺失或异常调控（如lats介导的激活缺失）而失活可能通过破坏DREAM复合物的组装和功能来促进卵巢癌的发生。我们建议通过使用以下策略表征细胞中的DYRK1A通路来验证这一假设。在Aim 1中，我们将确定DYRK1A的缺失是否有助于卵巢癌前体细胞的恶性转化，以及DYRK1A功能的获得是否可以在体外和体内抑制卵巢癌的生长。在Aim 2中，我们将确定Hippo通路在调节DYRK1A和DREAM复合体中的作用。由于DYRK1A在细胞中的功能和调控尚不完全清楚，我们鉴定了DYRK1A相互作用蛋白。在Aim 3中，我们将描述新的DYRK1A相互作用蛋白，这些蛋白因其在发育和癌症中的潜在意义而被优先考虑，并确定这些因子在DYRK1A生长抑制功能中的作用。提出的DYRK1A调控通路的生化和功能表征将推进我们对静止的认识，提高我们对卵巢癌发病机制的理解，并确定DYRK1A是否可以安全地靶向治疗DS。

DESCRIPTION (provided by applicant): The ability of cells to arrest proliferation and to enter a G1-arrested or G0 (quiescent) state is critical for tumor suppression. Retinoblastoma (RB) family proteins, including p130, mediate G0/G1 arrest in mammalian cells by mechanisms that are not fully understood. Upon entry into G0/G1, p130 is recruited into the DNA-binding DREAM protein complex (DP, RB-like, E2F, and MuvB core). The MuvB core of 5 proteins (LIN9, LIN37, LIN52, LIN54, and RBBP4) plays a dual role in transcriptional control. When bound to p130 in G0/G1, the MuvB core forms the DREAM complex and represses more than 800 cell cycle-dependent genes. When released from DREAM, the MuvB core binds to BMYB to activate mitotic genes. DYRK1A protein kinase plays a critical role in promoting assembly of the DREAM complex by phosphorylating the LIN52 subunit, which is required for binding between the p130 and MuvB core. Inhibition of DYRK1A is sufficient to block assembly of the DREAM complex and to override G0/G1 arrest. The DYRK1A gene is mapped to a Down syndrome (DS) critical region, and its upregulation due to trisomy 21 contributes to mental impairment in people with DS. It is important to establish the role of DYRK1A in tumor suppression because of its potential value as a therapeutic target to improve the cognitive functions of people with DS. Previous studies have revealed an essential role of DYRK1A in G0/G1 arrest, mediated by RB family, as well as in oncogenic Ras-induced senescence. Furthermore, biochemical data suggest that DYRK1A is downstream of LATS2 kinase in the Hippo tumor suppressor pathway. Our preliminary analysis found that the DYRK1A gene undergoes frequent copy number loss in high-grade ovarian carcinoma, suggesting that inactivation of DYRK1A contributes to the pathogenesis of ovarian cancer. We hypothesize that inactivation of DYRK1A in ovarian cancer due to genetic losses or aberrant regulation (such as loss of LATS-mediated activation) could promote ovarian carcinogenesis by disrupting the assembly and function of the DREAM complex. We propose to test this hypothesis by characterizing DYRK1A pathways in cells using the following strategy. In Aim 1, we will determine whether loss of DYRK1A contributes to the malignant transformation of the ovarian cancer precursor cells and whether gain of DYRK1A function can suppress the growth of ovarian cancers in vitro and in vivo. In Aim 2, we will determine the role of the Hippo pathway in regulating DYRK1A and the DREAM complex. Since the function and regulation of DYRK1A in cells is not fully understood, we identified DYRK1A-interacting proteins. In Aim 3, we will characterize novel DYRK1A-interacting proteins that have been prioritized by their potential significance in development and cancer, and establish the role of these factors in the growth arrest functions of DYRK1A. The proposed biochemical and functional characterization of DYRK1A-regulated pathways will advance our knowledge of quiescence, improve our understanding of the pathogenesis of ovarian cancer, and establish whether DYRK1A can be safely targeted to treat DS.