Spinal Muscular Atrophy: SMNs role in MRNA localization and local translation

描述(申请人提供)：脊髓性肌萎缩症(SMA)是一种常见的遗传性神经退行性疾病，由SMN1基因突变或缺失引起，SMN1基因编码运动神经元蛋白(SMN)的无处不在的生存，SMN对所有细胞中剪接体SnRNP复合体的组装至关重要。一个尚未回答的主要问题是SMN的缺失如何导致神经元功能障碍和SMA。要确定SMN在神经元中的功能，还需要进一步的工作。免疫组织化学研究显示，SMN定位于脊髓切片中的轴突和树突。利用培养神经元的高分辨率免疫荧光，我们发现SMN在生长锥体中形成与RNA和核糖体共存的颗粒。利用活细胞成像，我们发现EGFP-SMN颗粒在活神经元的神经元突起和生长锥体中显示出快速的双向运动。从转基因SMA小鼠模型中培养的原代运动神经元显示出轴突缺陷，包括生长锥体中β-肌动蛋白mRNA的丢失，这表明SMN在mRNA定位机制中具有新的功能。我们的初步数据表明，SMN与mRNA结合蛋白ZBP1、ZBP2和HUD有关，这些蛋白参与了神经元mRNA的定位和稳定性。我们假设这些信使核糖核酸结合蛋白与SMN相互作用，促进信使核糖核酸在生长过程中的定位和生长锥体内的稳定。本文提出的实验将进一步表征SMN、HUD、ZBP与RNA运输颗粒内相关mRNAs之间的相互作用、分子相互作用和动态调节。SMN缺失的运动神经元将被用来识别在其定位和翻译过程中发生改变的特定mRNAs。我们将调查SMN缺陷型神经元内局部mRNA调节功能障碍是否导致生长锥运动改变。这项研究将为SMN的神经元功能提供新的见解，mRNP复合体的组装、定位和/或翻译可能存在的缺陷可能参与SMA的疾病过程。

DESCRIPTION (provided by applicant): Spinal Muscular Atrophy (SMA) is a common inherited neurodegenerative disease caused by mutations or deletions in the SMN1 gene that encodes for the ubiquitous Survival of Motor Neuron Protein (SMN), which is essential for the assembly of spliceosomal snRNP complexes in all cells. A major unanswered question is how the loss of SMN leads to neuronal dysfunction and SMA. Further work is greatly needed to identify functions for SMN in neurons. Immunohistochemical studies have revealed localization of SMN to axons and dendrites from sections of spinal cord. Using high-resolution immunofluorescence of cultured neurons, we have shown that SMN forms granules which colocalize with RNA and ribosomes in growth cones. Using live cell imaging, we showed that EGFP-SMN granules exhibited rapid and bi-directional movements in neuronal processes and growth cones of live neurons. Primary motor neurons cultured from a transgenic mouse model of SMA displayed axonal defects that include loss of beta-actin mRNA from growth cones, suggesting a novel function for SMN in the mechanism of mRNA localization. Our preliminary data indicate that SMN associates with the mRNA binding proteins, ZBP1, ZBP2 and HuD, which are known to be involved in neuronal mRNA localization and stability. We hypothesize that these mRNA binding proteins interact with SMN to facilitate mRNA localization in processes and stabilization within growth cones. Experiments proposed here will further characterize the association, molecular interactions and dynamic regulation between SMN, HuD, ZBPs and associated mRNAs within RNA transport granules. SMN deficient motor neurons will be used to identify specific mRNAs that are altered in their localization and translation. We will investigate whether dysfunction of local mRNA regulation within SMN-deficient neurons contributes to altered growth cone motility. This research will provide new insight into neuronal functions for SMN, whereby possible defects in the assembly, localization and/or translation of mRNP complexes may contribute to the disease process in SMA.