Roles of SN27 in regulating glutamate receptors during neurodegeneration

描述（由申请人提供）：在各种神经退行性疾病如阿尔茨海默病（AD）和唐氏综合征（DS）中观察到谷氨酸受体（GluR）失调。然而，负责在GluR的变化和GluR失调的神经退行性变的贡献的潜在机制仍然在很大程度上难以捉摸。在我们的初步研究中，我们发现分选连接蛋白27（SNX 27）蛋白可以与GluRs相互作用，并控制其运输到质膜进行再循环。SNX 27在大脑中大量表达，SNX 27纯合敲除（KO）小鼠表现出严重的神经病理学，类似于AD和DS患者中发现的神经病理学。SNX 27杂合KO小鼠具有记忆缺陷，并且它们的神经元具有降低的GluR水平和突触功能障碍。DS是由21号染色体的额外拷贝引起的，这导致该染色体上的基因过量。我们发现，在DS患者的大脑中，21号染色体上的微小RNA基因（miR-155）上调，该基因负调节转录因子C/EBP ²，同时伴随着C/EBP ²和SNX 27的减少。我们还表明，C/EBP？正调控SNX 27的表达。这些结果揭示了导致DS中GluR失调和突触功能障碍的miR-155/C/EBP？/SNX 27通路。由于晚期DS患者发展为AD样病理和认知缺陷，我们研究了SNX 27对<$-淀粉样蛋白（A <$）生成的影响，A <$-淀粉样蛋白是AD发病机制的主要罪魁祸首，并通过<$-和<$-分泌酶的连续切割从<$-淀粉样蛋白前体蛋白衍生而来。我们发现SNX 27的过表达降低了A <$的产生以及<$-分泌酶的水平和活性。因此，我们假设SNX 27在调节GluRs和突触功能中起重要作用，并且SNX 27缺陷可能导致DS和AD的神经变性。在本研究中，我们将通过以下方法进一步确定SNX 27在DS中的参与：（1）证实SNX 27及其调节剂对GluRs的调节作用（miR 155和C/EBP）在原代神经元中的作用~（2）利用病毒系统调节miR 155、C/EBP和SNX 27缺陷小鼠和Ts 65 Dn小鼠中的SNX 27水平，并研究这些小鼠中受损的表型是否可以改善，以及（3）产生脑特异性SNX 27转基因（Tg）小鼠并将它们与Ts 65 Dn小鼠杂交以研究通过SNX 27过表达的任何病理学改善。此外，我们将探讨SNX 27在AD中的作用。我们将：（1）调节SNX 27的水平，并确定是否以及如何影响分泌酶水平/运输/活性和APP加工/A？生成~（2）研究SNX 27是否受A？介导的神经毒性的影响并参与A？介导的神经毒性，并检查AD脑中SNX 27的任何变化~（3）将SNX 27 Tg小鼠与Tg 2576 AD小鼠杂交，并尝试挽救AD小鼠中受损的表型。这些结果将阐明SNX 27在调节DS和AD中GluRs和突触功能中的作用，为疾病发病机制/病理学提供新的分子机制。

DESCRIPTION (provided by applicant): Dysregulation of glutamate receptors (GluRs) has been observed in various neurodegerative diseases such as Alzheimer's disease (AD) and Down Syndrome (DS). However, the underlying mechanisms responsible for the changes in GluRs and the contribution of GluR dysregulation to neurodegeneration have remained largely elusive. In our preliminary studies, we found that the Sorting Nexin 27 (SNX27) protein can interact with GluRs and govern their trafficking to the plasma membrane for recycling. SNX27 is abundantly expressed in the brain and SNX27 homozygous knockout (KO) mice exhibit severe neuropathologies resembling those found in AD and DS patients. SNX27 heterozygous KO mice have memory deficits and their neurons have reduced GluR levels and synaptic dysfunction. DS is caused by an extra copy of chromosome 21 which results in over- dosage of the genes on this chromosome. We found that in DS patient brains, a microRNA gene on chromosome 21, miR-155, which negatively regulates the transcription factor C/EBP¿, was upregulated, accompanied by a concomitant reduction of C/EBP¿ and SNX27. We also show that C/EBP¿ positively regulates SNX27 expression. These results reveal a miR-155/C/EBP¿/SNX27 pathway that leads to GluR dysregulation and synaptic dysfunction in DS. Since late stage DS patients develop AD-like pathologies and cognitive deficits, we studied the effect of SNX27 on the generation of ¿-amyloid (A¿), which is the primary culprit in AD pathogenesis and is derived from ¿-amyloid precursor protein through sequential cleavages by ¿- and ¿-secretases. We found that overexpression of SNX27 reduces A¿ production and the level and activity of ¿-secretase as well. Therefore, we hypothesize that SNX27 plays an important role in regulating GluRs and thus synaptic function, and that SNX27 deficiency may contribute to neurodegeneration in DS and AD. In this proposal, we will further ascertain the participation of SNX27 in DS by: (1) corroborating the regulation of GluRs by SNX27 and its modulators (miR155 and C/EBP¿) in primary neurons~ (2) using virus systems to modulate miR155, C/EBP¿, and SNX27 levels in SNX27 deficient mice and Ts65Dn mice, and study whether the impaired phenotypes in these mice can be ameliorated~ and (3) generating brain-specific SNX27 transgenic (Tg) mice and crossing them with Ts65Dn mice to study any amelioration of pathology through SNX27 overexpression. Moreover, we will explore the role of SNX27 in AD. We will: (1) modulate the levels of SNX27 and determine whether and how ¿-secretase level/trafficking/activity and APP processing/A¿ generation are affected~ (2) study whether SNX27 is affected by and involved in A¿-mediated neurotoxicity and examine any changes in SNX27 in AD brains~ and (3) cross SNX27 Tg mice with Tg2576 AD mice and attempt to rescue impaired phenotypes in AD mice. These results will elucidate the role of SNX27 in regulating GluRs and synaptic functions in DS and AD, providing novel molecular mechanisms for disease pathogenesis/pathology.