Protein Amyloidogenesis in the Epididymis: Mechanisms and Biological Significance

附睾中蛋白质淀粉样蛋白生成：机制和生物学意义

基本信息

批准号：
7992363
负责人：
Gail A Cornwall
金额：
$ 29.89万
依托单位：
TEXAS TECH UNIVERSITY HEALTH SCIS CENTER
依托单位国家：
美国
项目类别：
财政年份：
2008
资助国家：
美国
起止时间：
2008-12-01 至 2013-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7992363
关键词：
Address Adopted Affect Agglutination Alzheimer&apos s Disease Amyloid Amyloid Fibrils Antibodies Apical Biochemical Biological Biological Assay Biological Process Calcium Cell Culture Techniques Cell physiology Cells Complex Crowding Cystatins Defect Degenerative Disorder Deposition Detection Disease Disease Marker Dyes Electron Microscopy Endocytosis Enzymes Epididymis Epithelium Excision Exhibits Exposure to Gel Goals Health Human Immunohistochemistry Impairment In Vitro Incubated Infertility L68Q cystatin C Lead Link Liquid substance Mechanics Messenger RNA Modeling Molecular Molecular Chaperones Molecular Conformation Molecular Models Molecular Sieve Chromatography Mus Mutant Strains Mice Nature Negative Staining Neurobiology Neurodegenerative Disorders Outcome Parkinson Disease Patients Process Protein Family Protein Precursors Protein Secretion Proteins Proteolysis Quality Control Recombinants Reproduction Role Sperm Agglutination Sperm Maturation Sperm Motility Structure Techniques Testing Testis Transglutaminases Trypsin Tubular formation Wild Type Mouse amyloid formation amyloidogenesis base biological systems cerebral artery crosslink cystatin As cytotoxic cytotoxicity extracellular in vivo male molecular mass molecular modeling monomer mutant novel post gamma-globulins prevent protein aggregate protein aggregation protein crosslink protein structure reproductive research study sperm analysis sperm cell sperm function

项目摘要

DESCRIPTION (provided by applicant): The long range objective of our studies is to determine the biological significance of amyloid-type protein aggregation and mechanisms for its control in the epididymal lumen using the cystatins as molecular models. The abnormal accumulation of aggregated protein, also known as amyloid, is common in degenerative diseases including Alzheimer's disease. Amyloid in the testis and epididymis has also been implicated in human infertility. Proteins, including the cystatins, which can self-aggregate and form amyloid adopt a common cytotoxic structure during their aggregation. Because of the active secretion of proteins and profound removal of fluid by the epithelium, macromolecular crowding is likely to occur in the tubular lumen of the epididymis causing amyloid-type protein aggregation. However, because of its critical role in sperm maturation, surveillance/clearance mechanisms must be in place to control this process and prevent a pathological accumulation of cytotoxic aggregates. We have established that the cystatins CRES and cystatin C are present in the caput lumen as high molecular mass oligomeric complexes. We have also shown that CRES is associated with defined structures in the epididymal lumen. Furthermore, in vitro CRES and cystatin C form soluble amyloid precursors, which may be cytotoxic, as well as amyloid fibrils. We have also determined that male mice expressing the mutant L68Q cystatin C, an unstable and highly amyloidogenic form, are infertile possibly due to excess cystatin C oligomeric complexes in the lumen. These novel findings emphasize the critical nature of controlling protein aggregation in the epididymis. One mechanism by which the epididymis may control aggregation is by transglutaminase (TG) crosslinking resulting in protein aggregates in a nontoxic conformation. In support we have shown TG activity in the lumen, that CRES is a substrate for TG, and that TG will form CRES oligomers in caput fluid. Based on these studies we propose that amyloid-type protein aggregation occurs in the epididymal lumen and that quality control mechanisms, such as TG crosslinking, prevent the accumulation of toxic protein aggregates thereby maintaining normal epididymal function. We also propose that conditions that impair these protective mechanisms can negatively impact sperm maturation and function. We will address this hypothesis by: 1) characterizing amyloid-type aggregation in the epididymal lumen; 2) examine the pathological consequences of excessive amyloid aggregation; and 3) examine mechanisms of extracellular quality control in the epididymis. PUBLIC HEALTH RELEVANCE: The objective of our studies is to determine the biological significance of amyloid-type protein aggregation and mechanisms for its control in the epididymal lumen using the cystatins as molecular models. A completion of our aims will provide valuable information for our understanding of amyloid formation not only in the reproductive tract and its potential role in infertility but in general and as such may lead to new therapies and/or markers for diseases associated with extracellular aggregated proteins such as Alzheimer's disease.

描述（由申请人提供）：我们的研究的远距离目标是确定使用囊蛋白作为分子模型的淀粉样蛋白型蛋白聚集的生物学意义和在附属管道中对其控制的机制。聚集蛋白的异常积累，也称为淀粉样蛋白，在包括阿尔茨海默氏病在内的退行性疾病中很常见。睾丸和附睾中的淀粉样蛋白也与人类不育症有关。蛋白质，包括囊肿，可以自我聚集并形成淀粉样蛋白在聚集过程中采用常见的细胞毒性结构。由于蛋白质的积极分泌和上皮的深刻去除液，大分子人群可能会发生在附睾的管状腔中，从而导致淀粉样蛋白型蛋白质聚集。但是，由于其在精子成熟中的关键作用，必须建立监视/清除机制来控制这一过程并防止细胞毒性骨料的病理积累。我们已经确定，囊蛋白CRES和胱抑素C存在于高分子质量寡聚复合物中。我们还表明，CRE与附睾腔中定义的结构有关。此外，体外CRE和半胱氨酸蛋白酶C形成可溶性淀粉样蛋白前体，可能是细胞毒性的，以及淀粉样蛋白的原纤维。我们还确定，表达突变体L68Q胱抑素C的雄性小鼠是一种不稳定且高度淀粉样蛋白生成的形式，可能是由于管腔中过量的胱抑素C寡聚复合物所致。这些新颖的发现强调了控制附睾中蛋白质聚集的关键性质。附睾可能控制聚集的一种机制是通过转谷氨酰胺酶（TG）交联，导致蛋白质聚集物以无毒构象形成。为了支持，我们已经显示了管腔中的TG活性，CRE是TG的底物，TG将在盖帽流体中形成CRES低聚物。基于这些研究，我们建议淀粉样蛋白蛋白聚集发生在附睾腔中，并且质量控制机制（例如TG交联）可以防止有毒蛋白聚集物的积累，从而维持正常的附子函数。我们还建议损害这些保护机制的条件会对精子成熟和功能产生负面影响。我们将通过以下方式解决这一假设：1）表征附属腔中的淀粉样蛋白型聚集； 2）检查过度淀粉样蛋白聚集的病理后果； 3）检查附睾细胞外质量控制的机制。公共卫生相关性：我们研究的目的是确定淀粉样蛋白蛋白聚集的生物学意义以及使用伴半胱氨酸蛋白酶作为分子模型的附睾腔中对其控制的机制。我们的目标的完成将为我们的理解提供有价值的信息，以理解淀粉样蛋白的形成，不仅在生殖道及其在不孕症中的潜在作用，而且通常也可能导致新的疗法和/或标记与诸如阿尔茨海默氏病等细胞外聚集蛋白有关的疾病。