Laminin receptors and signals in Schwann cells

描述（由申请人提供）：层粘连蛋白及其受体几乎控制周围神经发育和髓鞘形成的所有方面。层粘连蛋白突变引起人类（先天性肌营养不良症，MDC1 a）和小鼠（营养不良症，dy）的髓鞘异常神经病变，表现为雪旺细胞-轴突相互作用受损，髓鞘形成和兰维耶形成淋巴结改变。一种层粘连蛋白受体，三聚糖聚糖，也与环轴蛋白相互作用，在Charcot-Marie-Tooth 4F （CMT4F）中发生突变。在之前的资助期内，我们使用条件诱变来破坏转基因小鼠雪旺细胞中的主要层粘连蛋白受体，并确定（31）整合素在径向分选过程中对雪旺细胞-轴突相互作用至关重要，糖歧义蛋白对Ranvier节点的组织很重要，糖歧义蛋白和6-34整合素都赋予髓磷脂稳定性。新出现的观点是层粘连蛋白受体具有特异性和重叠功能。此外，它们控制有髓纤维的径向和纵向形态发生，从而在远离基底层的轴突上组织均匀的分子。本提案的总体目标是了解这是如何发生的，定义单个层粘连蛋白或受体的作用机制，以及它们激活的信号通路。我们将利用我们生产或收集的独特的条件等位基因组和Cre转基因，使我们能够破坏转基因小鼠雪旺细胞中的层粘连蛋白受体和相关信号分子。我们将结合体内的形态学和生化分析、突变培养研究、延时显微镜和一种独特的技术来保存活的戏弄纤维的细胞结构。结合这些技术，我们将探索pi整合素的分子效应，朗维耶淋巴结的糖异常蛋白，糖异常蛋白与周轴蛋白参与雪旺细胞质的区隔化。观察荧光分子在有髓鞘节间室的运输将结合“活戏弄”与光激活和延时显微镜。该综合方法将确定不同层粘菌蛋白/受体/信号通路在周围神经中的作用，从而阐明MDC1A和CMT4F突变的发病机制。这些实验产生的信息将共同为制定MDC1A、CMT4F等遗传性神经病的治疗策略奠定基础，并促进所有神经病的神经再生和髓鞘再生。

DESCRIPTION (provided by applicant): Laminins and their receptors control nearly all aspects of peripheral nerve development and myelination. Laminin mutants cause a dysmyelinating neuropathy in man (congenital muscular dystrophy, MDC1 A) and mouse (dystrophic, dy) that manifests impaired Schwann cell-axon interactions, altered myelination and nodes of Ranvier formation. A laminin receptor, dystroglycan, also interacts with periaxin, mutated in Charcot-Marie-Tooth 4F (CMT4F). In the previous grant period we used conditional mutagenesis to disrupt singly or multiply the major laminin receptors in Schwann cells of transgenic mice, and have determined that (31 integrins are crucial for Schwann cell-axon interactions during radial sorting, dystroglycan is important for organization of nodes of Ranvier, and both dystroglycan and a6-34 integrin confer myelin stability. The emerging view is that laminin receptors have both specific and overlapping functions. In addition they control both radial and longitudinal morphogenesis of myelinated fibers, and thereby organize even molecules on the axon, at a distance from the basal lamina. The overall goal of this proposal is to understand how this occurs, defining the mechanism of action of single laminins or receptors, and the signaling pathways that they activate. We will take advantage of the unique group of conditional alleles and Cre transgenes that we produced or collected, that allow us to disrupt laminin receptors and relevant signaling molecules in Schwann cells of transgenic mice. We will combine morphological and biochemical analysis in vivo with studies of mutant cultures, time-lapse microscopy and a unique technique that preserves cytoarchitecture of live teased fibers. Combining these techniques, we will probe molecular effectors of pi integrin, of dystroglycan at nodes of Ranvier, and participation of dystroglycan with periaxin in compartmentalization of the Schwann cell cytoplasm. Observation of transport of fluorescent molecules in compartments of myelinated internodes will combine "live teasing" with photoactivation and time lapse microscopy. This comprehensive approach will establish the role of the different laminins/receptors/signaling pathways in peripheral nerve, and thereby clarify the pathogenesis of MDC1A and CMT4F mutations. The information produced by these experiments will collectively form a basis for developing treatment stategies of MDC1A, CMT4F and other hereditary neuropathies, and to promote nerve regeneration and remyelination in all neuropathies.