REGULATION OF MYELIN P2 GENE

髓鞘形成和再髓鞘形成过程的调节很差。 明白了。我们的长期目标是更好地理解这些过程。 然而，眼前的目标是理解单一的监管 蛋白质-P2蛋白质。我们将研究该基因的表达调控。 人类和小鼠的P2基因。为了完成这些研究，我们克隆了 小鼠和人的P2基因，并产生了大鼠雪旺细胞系(MT4H1)和 将试图产生一种人类雪旺细胞系。在具体目标1中，我们 将确定小鼠P2的重要近端调节区 将该基因导入大鼠雪旺细胞。事实上，在《初选》中 研究发现，我们发现了一个富含TC的区域，它下调了P2的表达 表达和第二个上调P2表达的区域。 此外，我们已经确定了三种转录因子，它们可以改变 P2表达：c-jun/c-fos，CAAT/增强结合蛋白(C/EBP)，属于 BZip转录因子家族，以及HNF-1，aPOU结构域 蛋白。这三个转录因子如何影响P2的表达 将被详细审查。此外，启动子的区域是 将确定组织特异性表达的必要条件和充分性 通过将5‘-缺失构建体导入Schwann细胞和其他 细胞类型，以及产生转基因动物和检查组织 用来表达β-半乳糖苷酶。 在具体目标2中，我们将使用以前用于生成RAT的策略 人雪旺细胞系产生人雪旺细胞系。这些细胞系 将获得的细胞进行免疫细胞化学研究，形态观察 外貌、生化特性和在联合培养条件下形成髓鞘的能力 用背根神经节神经突起培养。只有那些细胞最多 类似于未转基因的人类雪旺细胞将被用于进一步 学习。虽然我们已经克隆了人类P2基因，并知道 限制性内切酶图谱，我们还必须对启动子进行测序。一旦序列被 已知，类似于对小鼠P2启动子进行的研究将 被引导了。这些研究应该有助于理解其中的相似之处 以及人类和小鼠髓鞘P2基因的调控差异。 有关这一规定的信息可能会提供对这一过程的洞察 髓鞘形成和脱髓鞘，然后可能被用来帮助 设计可能的疗法以帮助由以下原因引起的疾病的康复 脱髓鞘或髓鞘异常症。

The regulation of the processes of myelination and remyelination are poorly understood. Our long-term goals are to better understand these processes. However, the immediate goals are to understand the regulation of a single protein-the P2 protein. We will study the regulation of expression of the human and mouse P2 genes. To accomplish these studies, we have cloned the mouse and human P2 genes and generated a rat Schwann cell line (MT4H1) and will attempt to generate a human Schwann cell line. In Specific Aim #1, we will identify the important proximal regulatory regions of the mouse P2 gene by transfection into rat Schwann cells. In fact, in the "Preliminary Studies," we have identified a TC-rich region which down-regulates P2 expression and a second region that up-regulates P2 expression. Furthermore, we have identified three transcriptional factors which alter P2 expression: c-jun/c-fos, CAAT/enhance binding protein (C/EBP), belonging to the bZip family of transcriptional factors, and HNF-1, aPOU domain protein. How these three transcriptional factors affect P2 expression will be examined in detail. Furthermore, the regions of the promoter that are necessary and sufficient for tissue-specific expression will be determined by transfecting the 5'-deletion constructs into Schwann cells and other cell types, as well as, producing transgenic animals and examining tissue for beta-galactosidase expression. In Specific Aim #2, we will use strategies previously used to generate rat Schwann cell lines to produce human Schwann cell lines. The cell lines obtained will be characterized by immunocytochemical studies, morphological appearance, biochemical properties, and ability to form myelin when co- cultured with dorsal root ganglia neurites. Only those cells most resembling untransfected human Schwann cells will be used for further studies. Although we have cloned the human P2 gene and know the restriction map, we must still sequence the promoter. Once the sequence is known, studies similar to those undertaken for the mouse P2 promoter will be conducted. These studies should help in understanding the similarities and differences in the regulation of the human and mouse myelin P2 genes. Information about this regulation may provide insights into the processes of myelination and demyelination, which then may be used to help in designing possible therapies to aid recovery in diseases resulting from demyelination or dysmyelination.