HL-FAM13A Regulates the beta-catenin/Wnt Pathway in Chronic Obstructive Pulmonary Disease

 描述(申请人提供)：HL-131。慢性阻塞性肺疾病(COPD)是美国第三大死亡原因。吸烟(CS)和遗传易感性也对COPD有很大影响。在病例对照研究中，一个新的基因FAM13A(具有序列相似性的家族，成员A，功能未知)一直与COPD的易感性有关，在全基因组关联研究中，FAM13A与普通人群样本中的肺功能一致。最近，FAM13A基因座上的COPD风险等位基因被证明与人类肺样本中FAM13A的表达增加有关。然而，这个新基因在CS诱导的损伤下如何在体内发挥作用仍不清楚。我们知识中的这种巨大差距极大地阻碍了将这些基因发现转化为对COPD病理生物学的更好理解和个性化医学的巨大潜力。通过一系列的体外和体内实验，我们证明了Fam13a基因缺陷小鼠(Fam13a-/-)表现出由CS或弹性蛋白酶引起的肺气肿减轻。我们还首次将新的Gwas基因FAM13A与典型的Wnt途径联系起来，该途径对多种肺部疾病的肺发育和肺修复/再生是必不可少的。在体外，Fam13a促进了蛋白磷酸酶2A(PP2A)与？连环蛋白的组装，促进了？连环蛋白的磷酸化和降解。在此，我们建议将我们的研究扩展到深入和全面地剖析Fam13a对Wnt通路的调节以及它们的串扰如何在CS诱导的损伤中介导肺修复。在目标1中，我们将利用一个敏感和特异的GFP报告鼠系来激活Wnt通路，以表征在CS暴露过程中Fam13a对Wnt通路活性的时空调节。在目标2中，我们将全面研究Fam13a如何通过抑制CS诱导的损伤模型中的Wnt途径来调节肺再生。在Aim 2.1中，我们将在SP-C(表面活性蛋白C)阳性细胞和兼性祖细胞中建立一个双缺陷(Fam13a和ç-catenin)小鼠系，用于损伤后肺复气，然后将这些小鼠暴露于CS，以确定激活Wnt通路是否导致对CS诱导的Fam13a-/-小鼠空域扩大的抵抗力。在Aim 2.2中将进一步探讨SP-C阳性细胞在CS损伤后肺再生中的作用。我们将在体内存在或不存在Fam13a的情况下，用他莫昔芬诱导SPC阳性细胞后用GFP示踪标记SPC阳性细胞，然后将小鼠暴露于急、慢性CS中，以确定Fam13-Wnt轴在SPC阳性细胞介导的肺再生中的作用。在目标3中，我们将使用来自已知FAM13A基因的COPD患者的原代肺泡和呼吸道上皮细胞，以确定不同的细胞增殖和分化是否与与FAM13A基因相关的不同β-连环蛋白水平相关。本项目完成后，我们将更加深入地了解Fam13a-ç-catenin/Wnt通路在CS诱导的损伤中的作用以及人类COPD的发病机制。

 DESCRIPTION (provided by applicant): HL-131. Chronic obstructive pulmonary disease (COPD) is the third leading cause of death in the U.S. It is also strongly influenced by cigarette smoking (CS) and genetic predisposition. A novel gene, FAM13A (family with sequence similarity 13, member A, with unknown function) has been consistently associated with susceptibility to COPD in case-control studies and with lung function in general population samples in genome-wide association studies (GWAS). Very recently, the COPD risk allele at the FAM13A locus was shown to be associated with increased expression of FAM13A in human lung samples. However, how this novel gene functions in vivo under CS-induced injury remains unknown. This substantial gap in our knowledge has greatly impeded translation of these genetic discoveries into a better understanding of COPD pathobiology and the huge potential of personalized medicine. Using a series of in vitro and in vivo approaches, we demonstrate that Fam13a-deficient mice (Fam13a-/-) showed attenuated emphysema induced by either CS or elastase. We also make the first connection between the novel GWAS gene FAM13A and the canonical Wnt pathway, essential for lung development as well as lung repair/regeneration in a variety of pulmonary diseases. In vitro, Fam13a facilitates assembly of protein phosphatase 2A(PP2A) with ß-catenin and promotes the phosphorylation and degradation of ß-catenin. Herein, we propose to extend our studies to a deep and comprehensive dissection of the regulation of the Wnt pathway by Fam13a and how their cross-talk mediates lung repair during CS-induced injury. In Aim 1, we will utilize a sensitive and specific GFP reporter mouse line for Wnt pathway activation to characterize the temporal and spatial regulation of Fam13a on the activity of the Wnt pathway during CS- exposure. In Aim 2, we will fully characterize how Fam13a modulates lung regeneration through inhibiting the Wnt pathway in the CS-induced injury model. In Aim 2.1,we will generate a double-deficient (Fam13a and ß- catenin) mouse line specifically in SP-C(surfactant protein C)-positive cells, facultative progenitor cells for lung reair after injury and then expose these mice to CS to determine whether activation of the Wnt pathway led to resistance to CS-induced airspace enlargement in Fam13a-/- mice. Roles of SP-C-positive cells in lung regeneration after CS-induced injury will be further explored in Aim 2.2. We will trace-label SPC-positive cells with GFP after tamoxifen induction in the presence or absence of Fam13a in vivo, then expose mice to acute and chronic CS to determine the role of the Fam13-Wnt axis in lung regeneration mediated by SPC-positive cells. In Aim 3, we will apply human primary alveolar and airway epithelial cells from COPD subjects with known genotypes at the FAM13A locus to determine whether differential cell proliferation and differentiation correlate with varying beta-catenin levels associated with FAM13A genotypes. Upon completion of this project, we will have a much deeper understanding of the role of Fam13a-ß-catenin/Wnt pathway during CS-induced injury and the mechanisms underlying human COPD.