ROLE OF THE NUCLEAR MATRIX PROTEIN NRB/B IN BRAIN

描述(逐字摘自申请者摘要)：定义了核矩阵 作为核的不可溶的骨架，并被牵连到调节 基因表达、细胞周期和核结构完整性 细胞骨架的中间丝。我们已经发现和 一种新的神经元核基质蛋白NRP/B(核)的特征 限制蛋白质/脑)，它包含两个主要结构元素：BTB 在预测的N-末端有类似结构域的结构，在 预测的C-末端结构域。NRP/B基因(5.5kb)主要表达于 人类胎儿和成人的大脑。在小鼠胚胎发育过程中，NRP/B基因的表达 在神经系统中被上调。NRP/B蛋白也在 大鼠原代海马神经元，而不是原代星形胶质细胞。在.期间 小鼠神经母细胞瘤细胞株NRP/B的分化 蛋白质表达上调。NRP/B在这些细胞中的过表达 显著地导致生长停滞并增强神经元突起的形成， 反义NRP/B寡核苷酸抑制轴突生长 大鼠原代海马神经元的发育及神经元突起 SH-SY5Y和Neuro 2A神经母细胞瘤细胞分化过程中的形成 此外，我们还观察到微量注射抗NRP/B抗体 抑制神经生长因子刺激PC12细胞突起生长 微量注射NRP/B基因可促进这些细胞的突起生长。 体内和体外实验表明，NRP/B可以 磷酸化并结合到具有功能活性的亚磷酸化形式 P110RB在SH-SY5Y神经母细胞瘤细胞诱导分化中的作用 通过维甲酸。NRP/B与p110RB的相互作用是通过 NRP/B的BTB/POZ结构域。此外，NRP/B与核的直接结合 肌动蛋白被观察到，这种联系是通过“海带”重复来调节的。 这些数据表明，NRP/B是一种新的核基质蛋白， 在初级神经元中特异表达，与p110RB相互作用，并 参与神经元突起形成的调节。 我们的目标是进一步表征这种新的神经元核基质蛋白。 关于它在控制神经元的调节通路中的作用 差异化。我们建议具体解决：(1)生物 Nrp/B蛋白的功能(S)；(2)Nrp/B蛋白的DNA结合特性和 其结构/功能关系；(3)NRP/B的特征 蛋白质在神经元生长和分化过程中的作用。这些研究应该 为这种核基质蛋白在神经元中的作用提供了新的见解 差异化。

DESCRIPTION(Verbatim from Applicant's Abstract): The nuclear matrix is defined as the insoluble framework of the nucleus and has been implicated in modulating gene expression, the cell cycle and nuclear structural integrity via linkage to intermediate filaments of the cytoskeleton. We have discovered and characterized a novel neuronal nuclear matrix protein, NRP/B (nuclear restricted protein/brain), which contains two major structural elements: a BTB domain-like structure in the predicted N-terminus, and a "kelch motif" in the predicted C-terminal domain. NRP/B mRNA (5.5 kb) is predominantly expressed in human fetal and adult brain. During mouse embryogenesis, NRP/B mRNA expression is upregulated in the nervous system. The NRP/B protein is also expressed in rat primary hippocampal neurons, but not in primary astrocytes. During the differentiation of murine Neuro 2A and human SH-SY5Y neuroblastoma cells, NRP/B protein expression is upregulated. Overexpression of NRP/B in these cells significantly induces growth arrest and augments neuronal process formation, while treatment with antisense NRP/B oligodeoxynucleotides inhibits neurite development of rat primary hippocampal neurons as well as neuronal process formation during differentiation of SH-SY5Y and Neuro 2A neuroblastoma cells. In addition, we have observed that microinjection of anti-NRP/B antibodies inhibited neurite outgrowth of PC12 cells in response to NGF stimulation, while microinjection of NRP/B cDNA promoted the neurite outgrowth of these cells. Both in vivo and in vitro experiments demonstrate that NRP/B can be phosphorylated and bind to the functionally active hypophosphorylated form of p110RB during neuronal differentiation of SH-SY5Y neuroblastoma cells induced by retinoic acid. The interaction of NRP/B with p110RB is mediated via the BTB/POZ domain of NRP/B. Furthermore, direct association of NRP/B with nuclear actin was observed, and this association is mediated via the "kelch" repeats. These data demonstrate that NRP/B is a novel nuclear matrix protein, specifically expressed in primary neurons, that interacts with p110RB and participates in the regulation of neuronal process formation. We aim to further characterize this novel neuronal nuclear matrix protein with regard to its role in regulatory pathways that control neuronal differentiation. We propose to specifically address: (1) the biological function(s) of the NRP/B protein; (2) the DNA binding properties of NRP/B and its structure/function relationships; and (3) the characterization of NRP/B protein during neuronal growth and differentiation. These studies should provide novel insights into the role of this nuclear matrix protein in neuronal differentiation.