The Biology of the ABCA3 Lipid Transporter in Health and Disease

 描述(申请人提供)：ABCA3作为一种脂类和磷脂转运体发挥作用，对板层小体的生物发生至关重要。随着我们在肺泡II型(AT2)细胞中发现ABCA3，大量令人信服的报告将ABCA3的突变与各种肺部疾病联系起来，包括新生儿致命的表面活性物质缺乏和年龄较大的儿童和成人的慢性间质性肺疾病(ILD)，突显了ABCA3对肺部健康和疾病的重要性。在我们先前工作的基础上，描述了ABCA3的基本细胞生物学，这个项目将使用整合和还原的方法来确定AT2细胞表达与人类遗传性肺部疾病相关的两类不同类型的ABCA3蛋白突变异构体对肺的细胞和器官型影响的机制和途径。我们已经鉴定了几个ABCA3突变，它们在体外表现出功能性(转运体)或运输性缺陷，后者诱导内质网应激和细胞毒性。此外，我们还开发了一种新的转基因敲入(KI)小鼠模型，表达最常见的ABCA3临床变体(ABCA3E292V)，该变体可发展成年龄相关的实质肺重塑。我们的初步数据表明，这个功能性的ABCA3突变体破坏了细胞的宏观自噬，通过溶酶体依赖和内质网应激非依赖的途径产生细胞毒性。我们的工作模型提出，ILD相关的ABCA3突变引起一组特定于类别的独特的AT2细胞反应，然后驱动异常的肺损伤/重塑。该项目包含三个主题相互关联的具体目标，以解决对肺细胞生物学至关重要的新兴主题： 蛋白质质量控制、细胞器动态平衡和细胞保护。特定目的1将描述AT2细胞功能障碍的机制，这些机制是通过体内表达运输缺陷的ABCA3E292V突变来实现的。具体目的2将研究ABCA3E292V肺对外源性“二次冲击”的易感性，并确定伴随的内质网应激在促进和增强异常肺重塑中的作用。特殊目的3将利用我们最近开发的第二个新的ILD KI小鼠模型，该模型携带最常见的表面活性蛋白C(SP-C)基因的ILD相关突变，SP-CI73T，我们最近已经证明，这种突变也深刻地扰乱了包括自噬在内的细胞质量控制。利用这一新的双基因模型，我们将评估SP-CI73T共表达在调控ABCA3E292V突变相关病理中的后果。以AT2细胞罕见的基因突变为模型的研究结果不仅将增进我们对家族性ILD病理生理学基础的分子机制的认识，还将更广泛地促进对AT2细胞功能障碍诱导的异常肺重塑的意义的理解，并为未来针对散发性肺纤维化的靶向治疗提供策略。

 DESCRIPTION (provided by applicant): ABCA3 functions as a lipid and phospholipid transporter and is critical for the biogenesis of lamellar bodies. Following our discovery of ABCA3 in alveolar type II (AT2) cells, its importance to lung health and disease has been highlighted by an abundance of compelling reports linking mutations in ABCA3 to various lung disorders, including fatal surfactant deficiency in newborns and chronic interstitial lung disease (ILD) in older children and adults. Building upon our prior work characterizing the basic cell biology of ABCA3, this project will use both integrative and reductionist approaches to define mechanisms and pathways underlying the cellular and organotypic consequences for the lung from the expression by AT2 cells of two distinct classes of mutant isoforms of ABCA3 proteins associated with inherited lung disorders in humans. We have identified several ABCA3 mutations that in vitro exhibit either functional (transporter) or trafficking defects with the later inducing ER stress and cytotoxicity. Furthermore, we have developed a novel transgenic knock-in (KI) mouse model expressing the most common clinical variant of ABCA3 (ABCA3E292V) that develops age dependent parenchymal lung remodeling. Our preliminary data demonstrate that this functional ABCA3 mutant disrupts cellular macroautophagy, producing cytotoxicity by lysosome-dependent and ER stress-independent pathways. Our working model proposes that ILD-associated ABCA3 mutations elicits a class-specific, distinct set of altered AT2 cell responses which then drives abnormal lung injury/remodeling. This project contains three thematically interrelated specific aims that address emerging themes critical to lung cell biology: protein quality control, organelle homeostasis, and cytoprotection. Specific Aim 1 will characterize the mechanisms underlying AT2 cell dysfunction from expression of the transport-deficient ABCA3E292V mutant in vivo. Specific Aim 2 will study the vulnerability of the ABCA3E292V lung to exogenous "2nd hits", and determine the role of concomitant ER stress in promoting and enhancing abnormal lung remodeling. Specific Aim 3 will utilize a second novel KI mouse model of ILD we recently developed bearing the most common ILD-associated mutation of the surfactant protein C (SP-C) gene, SP-CI73T, which we have recently shown to also profoundly disrupt cellular quality control including autophagy. Using this novel bi-genic model, we will then assess the consequences of SP-CI73T co-expression in modulating ABCA3E292V mutation associated pathology. Results from the proposed studies utilizing rare gene mutations in AT2 cells as models will not only enhance our knowledge of the molecular mechanisms underlying the pathophysiology of familial ILD, but also more broadly promote a better understanding of the significance of AT2 cell dysfunction-induced aberrant lung remodeling and provide strategies for future development of targeted therapies for sporadic lung fibrosis.