Laminin receptors and signals in Schwann cells

描述(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。层粘连蛋白突变导致人类(先天性肌营养不良，MDC1A)和小鼠(营养不良，dy)的髓鞘障碍神经病，表现为雪旺细胞-轴突相互作用受损，髓鞘改变和Ranvier形成的结节。层粘连蛋白受体Dstroglan也与周轴蛋白相互作用，在Charcot-Marie-Tooth 4F(CMT4F)中发生突变。在之前的资助期间，我们使用条件突变技术来干扰转基因小鼠雪旺细胞中主要的层粘连蛋白受体，并已确定(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，营养不良多糖对Ranvier结节的组织至关重要，并且营养不良多糖和a6-34整合素都赋予髓鞘稳定性。新出现的观点认为，层粘连蛋白受体既有特定的功能，也有重叠的功能。此外，它们控制有髓纤维的径向和纵向形态发生，从而在距离基膜一段距离的轴突上组织甚至分子。这项提议的总体目标是了解这种情况是如何发生的，定义单层粘连蛋白或受体的作用机制，以及它们激活的信号通路。我们将利用我们产生或收集的一组独特的条件等位基因和Cre转基因基因，使我们能够扰乱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和生化分析，以及对突变培养、延时显微镜和一种保存活的梳理纤维细胞结构的独特技术的研究。结合这些技术，我们将探索pi整合素的分子效应器，Ranvier结节上的dystroglan，以及dystrolycan与周轴蛋白一起参与雪旺细胞质的区域化。观察荧光分子在有髓节间隔间的运输将结合“活的挑逗”、光激活和时间推移显微镜。这一综合途径将确定不同的层粘连蛋白/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形成MDC1A、CMT4F和其他遗传性神经病的治疗策略的基础，并促进所有神经病的神经再生和重新髓鞘形成。

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.