Mutant Laminin Chains and Nephrotic Syndrome

层粘连蛋白链突变与肾病综合征

基本信息

批准号：
8140525
负责人：
Ying Maggie Chen
金额：
$ 12.87万
依托单位：
WASHINGTON UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2010
资助国家：
美国
起止时间：
2010-09-15 至 2015-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8140525
关键词：
Affect Amino Acid Sequence Animal Model Attenuated Biological Assay Cells Chemicals Childhood Congenital Nephrotic Syndrome Cultured Cells Data Defect Deposition Destinations Disease Endoplasmic Reticulum Engineering Exhibits Extracellular Matrix Goals Human In Vitro Kidney Kidney Diseases Kidney Failure Laminin Lead Missense Mutation Molecular Molecular Chaperones Molecular Weight Mus Mutant Strains Mice Mutation Nephrotic Syndrome Neurologic Pathogenesis Patients Peptide Sequence Determination Phenotype Point Mutation Production Protein Conformation Proteins Proteinuria Reporting Research Scheme Syndrome Taurodeoxycholic Acid Testing Therapeutic Therapeutic Uses Transgenes Transgenic Mice Variant WT1 gene endoplasmic reticulum stress glomerular basement membrane human disease improved in vivo lamin B2 mouse model mutant nephrin null mutation podocyte polymerization promoter protein folding protein misfolding public health relevance response scaffold trafficking

项目摘要

DESCRIPTION (provided by applicant): Laminin (2 (LAMB2) is a component of Laminin-521, the major laminin trimer of the kidney glomerular basement membrane (GBM). Null mutations in LAMB2 cause Pierson syndrome, characterized by congenital nephrotic syndrome with severe ocular and neurological defects. In contrast, most missense LAMB2 mutations, such as R246Q, cause isolated congenital nephrotic syndrome. To investigate the mechanisms whereby missense mutations cause proteinuria, we have generated transgenic mice in which R246Q mutant (2 is expressed in podocytes at different levels and replaces normal (2 in the GBM. Our preliminary data show that these mice exhibit proteinuria that can be attenuated by increased deposition of mutant R246Q-LAMB2 into the GBM. Moreover, in vitro, secretion of mutant LAMB2 proteins is severely inhibited compared to wild-type. Therefore, we hypothesize that pathogenic LAMB2 point mutations cause protein misfolding, ER stress, and secretion defects both in vitro and in vivo. Chemical chaperones, such as taurodeoxycholic acid (TUDCA), are low molecular weight compounds that can stabilize protein conformation and rescue trafficking-defective misfolded proteins. We have shown that TUDCA treatment of cells expressing the mutants significantly increases the secretion of mutant LAMB2 proteins into the medium. We propose, therefore, that chemical chaperones can improve the folding of mutant LAMB2 proteins, reduce ER stress, and improve the secretion of mutant laminin trimers into the GBM, thereby attenuating proteinuria in Lamb2-/-; mutLAMB2 transgenic mice. To test these hypotheses, we will: 1. Examine whether LAMB2 point mutations cause protein misfolding and ER stress both in vitro and in vivo. 2. Determine whether chemical chaperones can improve the secretion of LAMB2 point mutants and reduce ER stress in vitro. 3. Investigate whether chemical chaperones improve secretion of mutant laminin (2 and attenuate proteinuria in Lamb2-/-; mutLAMB2 mice. These studies will help us to understand the molecular mechanisms underlying the pathogenesis of congenital nephrotic syndrome and assay the feasibility for therapeutic use of chemical chaperones in a relevant animal model. Our results may have significant therapeutic implications for the treatment of congenital nephrotic syndrome in patients carrying missense LAMB2 mutations. PUBLIC HEALTH RELEVANCE: Congenital nephrotic syndrome is an important childhood kidney disease which causes proteinuria and progressive renal failure. We are studying whether the administration of pharmacologically active chaperones can ameliorate proteinuria in an animal model. The results may indicate a new treatment scheme for renal proteinuric diseases.

描述（由申请人提供）：层粘连蛋白 (2 (LAMB2) 是 Laminin-521 的组成部分，Laminin-521 是肾脏肾小球基底膜 (GBM) 的主要层粘连蛋白三聚体。LAMB2 的无效突变会导致皮尔森综合征，其特征是患有严重的先天性肾病综合征相反，大多数错义 LAMB2 突变（例如 R246Q）会导致孤立性先天性肾病综合征。突变导致蛋白尿，我们培育了转基因小鼠，其中 R246Q 突变体 (2 在不同水平的足细胞中表达，并取代 GBM 中的正常 (2)。我们的初步数据表明，这些小鼠表现出的蛋白尿可以通过增加突变体 R246Q 的沉积来减弱-LAMB2 进入 GBM 此外，在体外，与野生型相比，突变型 LAMB2 蛋白的分泌受到严重抑制，因此，我们假设致病性 LAMB2 点突变导致蛋白质。体外和体内的错误折叠、内质网应激和分泌缺陷。化学伴侣，例如牛磺脱氧胆酸 (TUDCA)，是低分子量化合物，可以稳定蛋白质构象并拯救运输缺陷的错误折叠蛋白质。我们已经表明，TUDCA 处理表达突变体的细胞显着增加了突变体 LAMB2 蛋白向培养基中的分泌。因此，我们认为化学伴侣可以改善突变型 LAMB2 蛋白的折叠，减少内质网应激，并改善突变型层粘连蛋白三聚体向 GBM 的分泌，从而减轻 Lamb2-/- 中的蛋白尿； mutLAMB2 转基因小鼠。为了检验这些假设，我们将： 1. 检查LAMB2点突变是否导致蛋白质错误折叠和ER应激体外和体内。 2.确定化学伴侣是否可以改善LAMB2点的分泌突变体并减少体外 ER 应激。 3. 研究化学伴侣是否能改善突变层粘连蛋白的分泌（2 和减轻 Lamb2-/- 中的蛋白尿； mutLAMB2 小鼠。这些研究将帮助我们了解先天性肾病综合征发病机制的分子机制，并分析化学伴侣在相关动物模型中治疗的可行性。我们的结果可能对携带错义 LAMB2 突变的先天性肾病综合征患者的治疗具有重要的治疗意义。公共卫生相关性：先天性肾病综合征是一种重要的儿童肾脏疾病，可导致蛋白尿和进行性肾功能衰竭。我们正在研究在动物模型中施用具有药理活性的伴侣是否可以改善蛋白尿。该结果可能为肾蛋白尿疾病提供新的治疗方案。