Biosynthesis and Trafficking of Surfactant Protein C In Health and Disease

表面活性剂蛋白 C 在健康和疾病中的生物合成和运输

• 批准号: 8731968
• 负责人: MICHAEL FRANCIS BEERS
• 金额: $ 39.42万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-09-09 至 2018-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8731968
• 关键词: Abnormal Epithelial Cell; Address; Adult; Affect; Alveolar; Anabolism; Apoptosis; Autophagocytosis; Binding; Binding Proteins; Bioenergetics; Biological Models; Cell membrane; Cell model; Cell physiology; Cells; Cellular biology; Cessation of life; Child; Cytoprotection; Data; Development; Disease; Distal; Epithelial; Epithelial Cells; Evaluation; Fibrosis; Functional disorder; Funding; Generations; Genes; Goals; Golgi Apparatus; Health; Homeostasis; Homologous Gene; Human; Impairment; In Vitro; Indium; Interstitial Lung Diseases; Laboratories; Ligands; Lung; Lung diseases; Mediating; Metabolism; Mitochondria; Modeling; Molecular; Monoubiquitination; Mutation; N-terminal; Organelles; Pathogenesis; Pathway interactions; Patients; Peptides; Phenotype; Phospholipids; Play; Process; Protein Isoforms; Proteins; Publishing; Pulmonary Fibrosis; Pulmonary Surfactant-Associated Protein C; Quality Control; Reporting; Respiratory Failure; Role; Signal Transduction; Stress; Structure; Surface; adapter protein; alveolar lamellar body; cytotoxicity; design; gain of function; in vitro Model; in vivo; mouse model; mutant; new therapeutic target; novel; programs; protein protein interaction; protein transport; prototype; public health relevance; recombinase; response; surfactant; trafficking; ubiquitin-protein ligase; uptake

DESCRIPTION (provided by applicant): Surfactant protein C (SP-C), a lung-specific hydrophobic peptide that enhances the biophysical activity of surfactant phospholipid, is synthesized as a 21 kD propeptide (proSP-C21) and post-translationally processed to yield a 3.7 kD surface active alveolar form. The importance of SP-C to lung health and disease has been underscored by observations that heterozygous expression of mutations in the SP-C gene (SFTPC) in humans is associated with interstitial lung disease (ILD). Recent studies from our laboratory have: (i) identified a novel PPDY motif in the proSP-C NH2 terminus and its ligand the E3 ligase Nedd4-2 as crucial elements in the targeting of proSP-C to the distal secretory pathway; (ii) functionally characterized the consequences of improper folding of the proSP-C COOH terminus for epithelial cell dysfunction seen with aggregation prone SFTPC mutations ("BRICHOS" domain) found in patients with accompanying ILD; (iii) uncovered a second class of phenotypically distinct ILD-associated SFTPC mutations ("Non-BRICHOS") that, while not ER retained, are instead mistrafficked to non-native organelles. Our published and new preliminary data also indicate that non-BRICHOS SP-C mutants disrupt both endosomal/ lysosomal function as well as macroautophagy producing cytotoxicity by ER stress independent pathways. This project seeks to build upon our long-standing discovery program studying cellular and molecular mechanisms underlying the biosynthetic metabolism of SP-C by further focusing on 3 emerging themes critical to lung cell biology: protein trafficking, organelle homeostasis, and cytoprotection. Specific Aim 1 will focus on defining the role of 4 key factors, the SP-C NH2 PPDY motif, Nedd4-2 WW domains, proSP-C monoubiquitination, and the adapter protein Golgi-associated, ?-adaptin homologues, Arf-binding protein 3 (GGA3) in anterograde trafficking of proSP-C. In addition, the consequences of proximal retention of proSP-C initiated by disruption of this targeting machinery will be studied using both well-characterized in vitro model systems and a new generation inducible, Cre-recombinase-driven mouse model of Nedd4-2 deficiency to evaluate quality control responses in vivo. In Specific Aim 2, mechanisms underlying AT2 cell dysfunction and cytotoxicity resulting from expression of mistrafficked non-BRICHOS SP-C mutant isoforms will be evaluated and compared with responses to aggregation prone COOH folding mutants. Results from the proposed studies will enhance our understanding of the molecular mechanisms underlying not only the pathophysiology of AT2 cell dysfunction that accompanies fibrotic lung remodeling in ILD but will provide a framework and model systems for "personalized" manipulation of proteostasis and cellular signaling for other disorders associated with parenchymal lung disease and epithelial dysfunction.

描述（由申请人提供）：表面活性剂蛋白 C (SP-C) 是一种肺特异性疏水性肽，可增强表面活性剂磷脂的生物物理活性，合成为 21 kD 前肽 (proSP-C21)，并经过翻译后处理，产生 3.7 kD 表面活性肺泡形式。人类 SP-C 基因 (SFTPC) 突变的杂合表达与间质性肺疾病 (ILD) 相关，这一观察结果强调了 SP-C 对肺部健康和疾病的重要性。我们实验室最近的研究发现：（i）在 proSP-C NH2 末端发现了一个新的 PPDY 基序及其配体 E3 连接酶 Nedd4-2，作为 proSP-C 靶向远端分泌途径的关键元件； (ii) 在功能上表征了 proSP-C COOH 末端的不当折叠对上皮细胞功能障碍的影响，这些功能障碍是在伴有 ILD 的患者中发现的易于聚集的 SFTPC 突变（“BRICHOS”结构域）所见； (iii) 发现了第二类表型不同的 ILD 相关 SFTPC 突变（“非 BRICHOS”），这些突变虽然没有保留 ER，但却被错误地输送到非天然细胞器。我们发表的和新的初步数据还表明，非 BRICHOS SP-C 突变体破坏内体/溶酶体功能以及巨自噬，通过内质网应激独立途径产生细胞毒性。该项目旨在建立在我们研究 SP-C 生物合成代谢背后的细胞和分子机制的长期发现计划的基础上，进一步关注对肺细胞生物学至关重要的 3 个新兴主题：蛋白质运输、细胞器稳态和细胞保护。具体目标 1 将重点定义 4 个关键因素的作用：SP-C NH2 PPDY 基序、Nedd4-2 WW 结构域、proSP-C 单泛素化以及接头蛋白高尔基体相关、β-适应素同源物、Arf 结合蛋白 3 (GGA3) 在 proSP-C 顺行运输中的作用。此外，将使用特征良好的体外模型系统和新一代可诱导、Cre重组酶驱动的 Nedd4-2 缺陷小鼠模型来研究因破坏该靶向机制而引发的 proSP-C 近端保留的后果，以评估体内质量控制反应。在具体目标 2 中，将评估因错误运输的非 BRICHOS SP-C 突变亚型表达而导致的 AT2 细胞功能障碍和细胞毒性的机制，并与易于聚集的 COOH 折叠突变体的反应进行比较。拟议研究的结果将增强我们对分子机制的理解，不仅是伴随 ILD 纤维化肺重塑的 AT2 细胞功能障碍的病理生理学，而且还将为与实质肺疾病和上皮功能障碍相关的其他疾病的蛋白质稳态和细胞信号传导的“个性化”操作提供框架和模型系统。