The role of the NIMA kinase in mitotic regulation

描述（由申请人提供）：不正确的有丝分裂调节有助于癌症和出生缺陷的进展，有关有丝分裂调节的发现对医学研究和疾病治疗产生了积极的影响。然而，新的见解和对医学的进一步影响的潜力显然是存在的。例如，有丝分裂过程中最引人注目但又最不为人所知的过程之一是核孔复合物（npc）的拆卸和重组，这种巨大的结构提供了跨越核膜的调节通道。这个过程是如何调节的，并与其他有丝分裂事件整合，可能涉及可逆磷酸化，我们计划的实验将有助于验证这一假设。我们已经证明，中性曲霉的NIMA有丝分裂激酶通过促进NPC分解来启动有丝分裂。NIMA是促进NPC分解的必要条件和充分条件，并显著影响脊椎动物细胞的有丝分裂，这表明存在保守的有丝分裂底物。因此，a . nidulans提供了一个强大而复杂的模型系统，用于理解NPCs的有丝分裂调节。该应用程序的目的包括使用亲和纯化和质谱技术来定义NPC相互作用组的动态有丝分裂变化，以鉴定新的NPC蛋白（Nups）以及参与有丝分裂调节的Nup相关蛋白。此外，我们还将重点研究一个新的Nup相关蛋白（AN0162），它包含一个跨膜结构域，并可能编码第四个跨膜Nup。我们的数据表明AN0162具有有丝分裂功能，在有丝分裂期间维持核仁周围核膜上的外周mRNA输出因子Gle1，我们将验证这一假设。我们的第三个重点是Nup2及其相关蛋白NupA，输入蛋白1和输入蛋白2的有丝分裂功能。Nup2和NupA在有丝分裂期间从npc转移到染色质，同时它们也被磷酸化。我们将验证NupA介导Nup2的有丝分裂染色质位置的假设，Nup2反过来控制输入蛋白1和2调节有丝分裂进程的功能。由于磷酸化似乎是这一过程不可或缺的一部分，最终目的是绘制Nup2、NupA和importin 2中有丝分裂磷酸化位点，然后对特定磷酸化位点进行体内突变分析，以确定其功能。

DESCRIPTION (provided by applicant): Incorrect regulation of mitosis contributes to progression of cancer and birth defects and discoveries regarding the regulation of mitosis have positively impacted medical research and disease treatments. However, the potential for new insights and further impact on medicine clearly exists. For example, one of the most dramatic yet least understood mitotic processes is the disassembly and reassembly of nuclear pore complexes (NPCs), massive structures providing regulated gateways across the nuclear envelope. How this process is regulated and integrated with other mitotic events potentially involves reversible phosphorylation and our planned experiments will help test this hypothesis. We have shown the NIMA mitotic kinase of Aspergillus nidulans initiates mitosis by promoting NPC disassembly. NIMA is both required and sufficient to promote NPC disassembly and markedly affects mitosis in vertebrate cells suggesting the existence of conserved mitotic substrates. A. nidulans therefore provides a powerful and sophisticated model system in which to understand mitotic regulation of NPCs. The aims of this application include defining the dynamic mitotic changes in the NPC interactome using affinity purifications and mass spectrometry to identify new NPC proteins (Nups) as well as Nup associated proteins involved in mitotic regulation. In addition we will also focus studies on a new Nup associated protein (AN0162) which contains a transmembrane domain and potentially encodes a fourth transmembrane Nup. Our data suggest that AN0162 has a mitotic function to maintain the peripheral mRNA export factor Gle1 on the nuclear envelope surrounding the nucleolus during mitosis and we will test this hypothesis. Our third focus is on the mitotic functions of Nup2 and its associated proteins NupA, importin 1 and importin 2. Nup2 and NupA translocate from NPCs to chromatin during mitosis when they also become phosphorylated. We will test the hypothesis that NupA mediates the mitotic chromatin location of Nup2, which in turn controls the functions of importin 1 and 2 to regulate mitotic progression. As phosphorylation appears integral to this process, the final aim is to map mitotic phosphorylation sites within Nup2, NupA and importin 2 and then carry out in vivo mutational analysis of the specific phosphorylation sites to determine their function. PUBLIC HEALTH RELEVANCE: How the cell cycle is regulated is of fundamental medical importance because when this regulation goes awry disease states such as birth defects and cancer can result. In addition, many current cancer treatments target different aspects of cell cycle regulation to preferentially kill cells actively passing through the cell cycle. Therefore in terms of public health, the more we understand how the cell cycle is regulated the better positioned we will be to develop drugs against new chemotherapeutic targets to combat diseased states associated with cell cycle defects. The experiments proposed will in addition further our understanding of the NIMA kinase human orthologues of which are involved in specific disease states (kidney disease). Finally, A. nidulans is the model organism of the Aspergilli. Insights to the biology of A. nidulans will impact research on the opportunistic Aspergillus pathogens as well as members being used to generate new pharmacologically active compounds.