Nup50 control of neurogenesis and cancer stem cell transformation

描述(申请人提供)：核孔复合体(NPC)是由~30个单独的核孔蛋白(NUP)的多个拷贝组成的大蛋白通道。NPC跨越核膜(NE)，促进所有核的运输。在对NPC作出贡献的大约30个核子中，有两个主要类型--(1)稳定地与东北方向联系的核子和(2)穿梭到核内部的动态核子。赫策实验室和其他机构最近的研究结果发现，移动核仁与远离核细胞的染色质相互作用。这是第一批核蛋白直接影响转录的报道。特别是，NUP98和NUP50结合在活性基因的近端，包括许多发育调节的基因座。令人惊讶的是，NUPS的敲除减少了靶基因的表达，并表明它们在复杂的基因网络中发挥着未被重视的调节作用。初步数据表明，NUP98与染色质修饰物结合。考虑到NUP98和NUP50结合基因的重叠(~75%)，推测NUP50也参与染色质重塑。这里提出的实验重点是假设NUP50靶向组蛋白甲基化复合体到特定的位置，从而促进活跃的转录。初步数据进一步表明，Nup50影响神经干细胞分化。包括Nup98在内的许多核蛋白与多种癌症相关，尽管人们对它们在肿瘤生长过程中的机制知之甚少。阐明NUPS与正常发育和癌细胞转化相关的机制将为在许多癌症中观察到的一类新的蛋白质提供立足点。这项提案旨在阐明NUP50如何调控脊椎动物的发育和肿瘤。初步实验将表征NUP50在体外神经发生过程中的特征，包括其蛋白质伙伴和染色质占有率的变化。表征NUP50在分化过程中的变化将包括详细研究ERK磷酸化如何调节NUP50与染色质相互作用的能力。最后，对Nup50的需求将在体外和体内神经发生过程中进行检测。体内分析将利用在将结果转化为哺乳动物系统时开发的现有工具，以便 提供生理相关性和洞察个体核素如何协调哺乳动物的发育。

DESCRIPTION (provided by applicant): Nuclear pore complexes (NPC) are large protein channels composed of multiple copies of ~30 individual nucleoporins (Nups). NPC span the nuclear envelope (NE) and facilitate all nuclear transport. Among the ~30 Nups that contribute to the NPC, there are two primary categories - (1) Nups that stably associate with the NE and (2) dynamic Nups that shuttle into the nuclear interior. Recent results from the Hetzer laboratory and others found that mobile Nups interact with chromatin away from NPCs. These were the first reports that Nups directly influence transcription. In particular, NUP98 and NUP50 bind proximal to active genes, including many developmentally regulated loci. Amazingly, knockdown of Nups reduces target gene expression and suggests that they serve unappreciated regulatory roles in complex gene networks. Preliminary data indicates that NUP98 binds to chromatin modifiers. Given the overlap (~75%) of NUP98 and NUP50 bound genes, it is speculated that NUP50 is also involved in chromatin remodeling. The experiments proposed here focus on the hypothesis that NUP50 targets histone methylation complexes to specific loci thereby promoting active transcription. Preliminary data further suggests that Nup50 influences neural stem cell differentiation. Many Nups, including Nup98, are correlated with a diverse array of cancers although their mechanisms during tumor growth are poorly understood. Elucidating mechanisms relating Nups to normal development and cancer cell transformation would provide a foothold into a new class of proteins observed in many cancers. This proposal aims to clarify how NUP50 regulates vertebrate development and neoplasia. Initial experiments will characterize NUP50 during in vitro neurogenesis, including changes in its protein partners and chromatin occupancy. Characterizing changes in NUP50 during differentiation will include a detailed study of how Erk phosphorylation regulates the ability of NUP50 to interact with chromatin. Lastly, the requirement for Nup50 will be assayed during in vitro and in vivo neurogenesis. In vivo analysis will utilize existing tools developed while translating results into a mammalian system in order to provide physiological relevance and insight into how individual Nups orchestrate mammalian development.