Function of FNDC3B in cardiovascular and pulmonary development

描述(申请人提供)：人类心血管和肺功能障碍通常危及生命，并产生巨大的经济负担。这种情况可能是由于发育异常或出生后感染、急性损伤或暴露于环境过敏原而引起的。这些疾病的高发病率为长期基础研究提供了明确的理论基础，以了解心血管和肺系统正常发育和动态平衡的分子基础。本研究的长期目标是了解包含3B的新型蛋白纤维连接蛋白III型结构域(FNDC3B)在哺乳动物心血管和肺发育和动态平衡中的功能。Fndc3b在心血管和肺脏系统中表达，Fndc3b突变的小鼠表现出呼吸窘迫并在出生后不久死亡。Fndc3b小鼠的心血管系统畸形，心脏表现为室间隔缺陷和致密性不全。肺发育也受到影响，传导气道上皮形态异常。FNDC3蛋白含有一个高度保守的WW结构域蛋白的共同结合位点。基于这些和其他发现，我们假设FNDC3B在肺上皮细胞和血管内皮细胞中发挥功能，调节正常肺和心血管功能所需的细胞-细胞黏附。我们还假设FNDC3B是分泌表面活性物质所必需的，并且FNDC3B通过与含有特定WW结构域的蛋白质结合来完成这些功能。为了验证这些假设，我们提出了三组实验。首先，我们将确定哪些细胞在小鼠的肺和心脏中表达FNDC3B。其次，我们将分析细支气管和肺泡上皮细胞的解剖和功能，并研究Fndc3b突变动物和对照动物正常肺功能所需蛋白质的表达是否受到影响。我们还将研究Fndc3b突变动物的细胞-细胞连接是否完整，以及Fndc3b突变动物的心房肌细胞和血管内皮细胞如板层小体、分泌颗粒、小凹和魏贝尔-帕拉德小体是否异常。第三，我们将使用(A)基于阵列的方法和(B)串联亲和纯化在小鼠呼吸道上皮细胞MEL15和HUVEC内皮细胞中表达的表位标记的FNDC3B，并随后进行质谱分析和数据库分析，来研究哪些WW蛋白与FNDC3B相互作用。这些研究将为FNDC3B在小鼠心血管和肺发育中的作用提供新的信息。这项基础研究的长期好处将是提高我们对心血管和肺发育以及体内平衡的分子和细胞基础的了解，这反过来将为诊断和最终治疗与这些基本系统功能障碍相关的人类疾病提供新的机会。 与公共卫生的相关性：新生儿心血管和肺功能异常是发病率和死亡率的常见原因，给人类带来巨大痛苦，并给卫生保健系统带来经济负担。了解这些疾病的原因是朝着改善此类疾病的诊断和治疗的长期目标迈出的重要一步。我们正在研究一种新的蛋白质FNDC3B的功能，它是小鼠正常的心肺发育和出生后存活所必需的。这项基础研究的长期好处将是增加我们对哺乳动物心肺发育的分子和细胞基础的了解，这反过来将改进与心血管和肺功能异常相关的人类疾病的诊断和最终治疗。

DESCRIPTION (provided by applicant): Human cardiovascular and pulmonary dysfunction is often life threatening and generates a huge economic burden. Such conditions can arise from abnormal development, or post-natal, from infection, acute injury or exposure to environmental allergen. The high incidence of these conditions provides a clear rationale for long- term basic research to understand the molecular basis for normal development and homeostasis of cardiovascular and pulmonary systems. The long-term goal of this research is to understand the function of the novel protein fibronectin type-III domain containing 3B (FNDC3B) in mammalian cardiovascular and pulmonary development and homeostasis. Fndc3b is expressed in cardiovascular and pulmonary systems and mice mutant for Fndc3b display respiratory distress and die shortly after birth. The cardiovascular system of Fndc3b mice is malformed and hearts display ventricular septal defects and non-compaction. Lung development is also affected and the conducting airway epithelium has abnormal morphology. FNDC3 proteins contain a highly conserved consensus binding site for WW-domain containing proteins. Based on these, and additional, findings we hypothesize that FNDC3B functions in pulmonary epithelial cells, and in vascular endothelial cells to modulate cell-cell adhesion required for normal pulmonary and cardiovascular function. We also hypothesize that FNDC3B is required for secretion of surfactant and that FNDC3B accomplishes each of these functions by binding to specific WW-domain containing proteins. To test these hypotheses, three sets of experiments are proposed. First, we will identify which cells express FNDC3B in the lung and heart of mice. Second, we will analyze the anatomy and function of epithelial cells lining the bronchiolar airway and alveoli and investigate whether the expression of proteins required for normal pulmonary function is affected in Fndc3b mutant and control animals. We shall also investigate whether cell- cell junctions are intact, and if unique secretory organelles required for normal function of airway epithelial cells, atrial myocytes and vascular endothelial cells such as lamellar bodies, secretory granules, caveolae and Weibel-Palade bodies are abnormal in Fndc3b mutant animals. Third, we will investigate which WW-proteins interact with FNDC3B using (a) an array based method and (b) by tandem-affinity purification of epitope- tagged FNDC3B expressed in mouse airway epithelial MEL15 cells and HUVEC endothelial cells followed by mass spectrometry and database analysis. These studies will provide new information about the function of FNDC3B in cardiovascular and pulmonary development in mice. The long-term benefit of this basic research will be to improve our knowledge of the molecular and cellular basis for cardiovascular and pulmonary development and homeostasis, which in turn will offer new opportunities for diagnosis, and ultimately treatment, of human conditions associated with dysfunction of these essential systems. PUBLIC HEALTH RELEVANCE: Abnormal cardiovascular and lung function in newborn infants is a common cause of morbidity and mortality that generates significant human suffering, and financial burden on the health-care system. Understanding the causes of these disorders is an important step towards the long- term goals of improved diagnosis and treatment for such conditions. We are investigating the function of a novel protein, FNDC3B, which is required for normal heart and lung development and post-natal survival of mice. The long-term benefit of this basic research will be to increase our knowledge of the molecular and cellular basis for cardio-pulmonary development in mammals, which in turn will offer improved diagnosis, and ultimately treatment, of human conditions associated with abnormal cardiovascular and pulmonary function.