Nup50 control of neurogenesis and cancer stem cell transformation

描述（由申请人提供）：核孔复合物（NPC）是由约30个单个核孔蛋白（Nups）的多个拷贝组成的大蛋白通道。NPC跨越核膜（NE）并促进所有核运输。在对NPC有贡献的~30种Nup中，有两种主要类别-（1）与NE稳定缔合的Nup和（2）穿梭进入核内部的动态Nup。最近来自Hetzer实验室和其他人的结果发现，移动的Nups与远离NPC的染色质相互作用。这是第一次报道Nups直接影响转录。特别地，NUP 98和NUP 50与活性基因近端结合，包括许多发育调节基因座。令人惊讶的是，Nups的敲低降低了靶基因的表达，并表明它们在复杂的基因网络中起着未被重视的调节作用。初步数据表明NUP 98与染色质修饰剂结合。鉴于NUP 98和NUP 50结合基因的重叠（~75%），推测NUP 50也参与染色质重塑。这里提出的实验集中在NUP 50将组蛋白甲基化复合物靶向特定位点从而促进活性转录的假设上。初步数据进一步表明，Nup 50影响神经干细胞分化。许多Nup，包括Nup 98，与多种癌症相关，尽管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蛋白质提供立足点。该提案旨在阐明NUP 50如何调节脊椎动物的发育和肿瘤形成。最初的实验将表征NUP 50在体外神经发生过程中的特征，包括其蛋白质伴侣和染色质占有率的变化。表征分化过程中NUP 50的变化将包括详细研究Erk磷酸化如何调节NUP 50与染色质相互作用的能力。最后，将在体外和体内神经发生期间测定对Nup 50的需求。体内分析将利用在将结果转化到哺乳动物系统中时开发的现有工具， 提供生理相关性和洞察如何个别Nups编排哺乳动物的发展。

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.