Identifying and characterizing gene(s) involved in beta-amyloid production.

描述（由申请人提供）：阿尔茨海默病（AD）的特征是由b-淀粉样蛋白（A？）肽和由微管相关蛋白tau的过度磷酸化的成对螺旋丝组成的细胞内神经元缠结（NFT）。多线的证据表明，生产过剩的A？在脑中的过度磷酸化是AD的主要原因，而过度磷酸化的tau已经在许多称为tau蛋白病的神经退行性疾病中发现。因此，识别参与这些过程的新基因/蛋白质是AD研究的主要目标，并可能提供潜在的治疗靶点。在上一个资助期，我们应用随机纯合基因扰动（RHGP）技术筛选参与A？并鉴定了一种新的小鼠基因Rps 23 r1（在最初的提交中以前被命名为Fg 01）。RPS 23 R1蛋白的过表达可显著降低A？产生、tau磷酸化和GSK-3活性。进一步的体外研究表明，RPS 23 R1与腺苷酸环化酶相互作用，增加cAMP合成，从而上调PKA活性。已知PKA的激活和GSK-3的失活均抑制A？生成和GSK-3失活也抑制tau磷酸化。Rps 23 r1的功能也在表达脑特异性Rps 23 r1的转基因小鼠中得到证实。此外，APP/PS1/tau三重转基因AD小鼠的AD样病理学得到改善，并且在将小鼠与Rps 23 r1转基因小鼠杂交后，突触标记蛋白的水平增加。虽然Rps 23 r1是一个独特的小鼠基因，但Rps 23 r1在包括人类在内的各种物种的细胞中产生相同的表型。Rps 23 r1起源于小鼠Rps 23（核糖体蛋白S23）mRNA逆转录，这是一个mRNA重新整合到基因组中导致基因复制/进化的过程。由于Rps 23逆转录也发生在人类中，因此人类可能具有功能性Rps 23 r1同源物。我们的研究结果表明，RPS 23 R1和功能性人类RPS 23 R1同源介导的信号通路可能在AD和其他疾病的发病机制中发挥重要作用，如癌症和糖尿病，其中PKA和GSK-3是至关重要的参与。因此，在这项竞争性更新提案中，我们的目标是确定RPS 23 R1的功能结构域，鉴定功能性人类Rps 23 R1同源物，进一步表征RPS 23 R1及其人类同源物，并破译其作用机制，特别是抑制A生成，tau磷酸化和GSK-3活性的调节。我们将脑特异性Rps 23 r1转基因小鼠与AD转基因（Tg 2576和APP/PS1/tau三联体）小鼠杂交，以确定Rps 23 r1是否可以改善AD样病理和行为/认知缺陷。我们还将产生Rps 23 r1条件性敲除小鼠，以探索Rps 23 r1的其他生理功能。这些研究的结果有望为制定防治AD和其他疾病的新策略提供重要信息。

DESCRIPTION (provided by applicant): Alzheimer's disease (AD) is characterized by extraneuronal plaques consisting of b-amyloid (A?) peptides and intracellular neurofibrillary tangles (NFTs) composed of hyperphosphorylated, paired helical filaments of the microtubule-associated protein, tau. Multiple lines of evidence suggest that overproduction of A? in the brain is a major cause of AD, whereas hyperphosphorylated tau has been found in many neurodegenerative diseases known as tauopathies. Hence, identification of new genes/proteins involved in these processes is a major goal in AD research and may provide potential therapeutic targets. In the previous funding period, we applied the random homozygous gene perturbation (RHGP) technology to screen for genes involved in A? generation and identified a novel mouse gene, Rps23r1(previously named Fg01 in the original submission). Overexpression of the RPS23R1 protein can significantly reduce A? generation, tau phosphorylation and GSK-3 activity. Further in vitro studies showed that RPS23R1 interacts with adenylate cyclases, increasing cAMP synthesis, which upregulates PKA activity. Both activation of PKA and inactivation of GSK-3 are known to inhibit A? generation and GSK-3 inactivation also inhibits tau phosphorylation. The function of Rps23r1 was also demonstrated in transgenic mice expressing brain-specific Rps23r1. Furthermore, the AD-like pathologies of the APP/PS1/tau triple transgenic AD mice were ameliorated and levels of synaptic marker proteins were increased after crossing the mice with the Rps23r1 transgenic mice. Although Rps23r1 is a unique mouse gene, Rps23r1 produces the same phenotype in cells of various species including human. Rps23r1 originated through mouse Rps23 (ribosomal protein S23) mRNA retroposition, a process where mRNA is reintegrated into the genome resulting in gene duplication/evolution. Since Rps23 retroposition also occurred in humans, it is possible that humans possess functional Rps23r1 homologs. Our results suggest that RPS23R1- and functional human RPS23R1 homolog-mediated signaling pathways may play important roles in the pathogenesis of AD and other diseases, such as cancer and diabetes, in which PKA and GSK-3 are crucially involved. Therefore, in this competitive renewal proposal we aim to determine the functional domain(s) of RPS23R1, identify functional human Rps23r1 homologs, further characterize RPS23R1 and its human homologs and decipher mechanisms underlying their effects, specifically inhibition of A generation, tau phosphorylation and modulation of GSK-3 activity. We will cross brain-specific Rps23r1 transgenic mice with AD transgenic (Tg2576 and APP/PS1/tau triple) mice to determine whether Rps23r1 can ameliorate AD-like pathologies and behavioral/cognitive deficits. We will also generate Rps23r1 conditional knockout mice to explore other physiological functions of Rps23r1. The results of these studies are expected to provide important information for developing new strategies to combat AD and other diseases.