miRNAs regulate lung epithelial progenitor cells in development and injury repair

描述（由申请人提供）：肺上皮祖细胞对于发育和成人气道再生和修复至关重要。控制其行为的机制包括发育过程中的增殖和分化以及气道损伤修复的进展在很大程度上是未知的。microRNAs（miRNAs）是基因表达的重要调节因子，但它们作为肺上皮祖细胞的调节因子的作用仍然很差。 明白 最近，我们已经证明miR-302/367是Gata6的直接靶点，Gata6是正常肺发育和气道再生所必需的转录因子，并调节肺上皮祖细胞增殖和分化以及细胞顶部-基底极性的平衡。我们已经开始探索miR-302/367介导的肺上皮祖细胞发育的重要性。miR-302/367在发育中的气道上皮中的过表达导致Sox 2+近端和Sox 9+远端祖细胞的扩增以及气道上皮细胞分化的降低。值得注意的是，我们最近的研究表明，与对照未损伤的肺相比，在萘损伤的肺中miR-302/367的表达显著增加，这表明miR-302/367在介导肺上皮基因转录和气道修复和再生期间的损伤反应中具有潜在作用。因此，我在这个提议中的工作假设是，miR-302/367作为Gata6依赖性调控网络的组成部分，涉及多个正反馈和负反馈回路，以调节肺上皮祖细胞的增殖和分化。 更好地了解miR-302/367如何调节肺上皮祖细胞在气道上皮发育和再生过程中对损伤的反应，将为多种肺部疾病提供重要的见解。 这将通过追求两个具体目标来实现： 具体目标1。K99阶段：确定miR-302/367在损伤后气道上皮发育和再生过程中调节肺上皮祖细胞行为的作用。a）使用ROSA-miR-302/367和floxed miR-302/367小鼠系确定miR-302/367的条件性获得和丧失对肺上皮祖细胞发育的影响。B）确定Wnt 5a的表达降低是否是miR-302/367功能获得突变体中肺上皮祖细胞扩增的原因。c）确定miR-302/367对于气道上皮中的稳态和再生的需求。 具体目标2。R00阶段：确定气道上皮发育和再生中miR-302/367调节上皮祖细胞行为的潜在机制。a）表征在发育和损伤后气道再生期间肺上皮中表达miR-302/367的细胞。B）确定miR-302/367在发育中的肺中调节的直接靶点和下游信号通路。 c）确定特异性miR-302/367调节的靶标或途径的调节是否会增强气道上皮修复和再生。

DESCRIPTION (provided by applicant): Lung epithelial progenitor cells are essential for development and adult airway regeneration and repair. The mechanisms controlling their behaviors including proliferation and differentiation during development and the progression of airway injury repair are largely unknown. MicroRNAs (miRNAs) are crucial modulators of gene expression, yet their involvement as regulators of lung epithelial progenitor cells is still poorly understood. Recently, We have demonstrated that miR-302/367 is a direct target of Gata6 -a transcription factor essential for proper lung development and airway regeneration- and regulates the balance of lung epithelial progenitor cell proliferation and differentiation as well s cell apical-basal polarity. We have begun to explore the importance of miR-302/367-mediated lung epithelial progenitor development. Overexpression of miR-302/367 in developing airway epithelium causes expansion of Sox2+ proximal and Sox9+ distal progenitor cells and decreased airway epithelial cell differentiation. Notably, our recent studies reveal that expression of miR- 302/367 is markedly increased in naphthalene-injured lungs compared to control uninjured lungs, suggesting a potential role for miR-302/367 in mediating lung epithelial gene transcription and the injury response during airway repair and regeneration. Therefore, my working hypothesis in this proposal is that miR-302/367 functions as components of a Gata6-dependent regulatory network involving multiple positive and negative feedback loops to modulate proliferation and differentiation of lung epithelial progenitor cells. A better understanding of how miR-302/367 regulates the behaviors of lung epithelial progenitor cells during airway epithelium development and regeneration in response to injury will provide important insights into multiple lung diseases. This will be accomplished by pursuing two specific aims: Specific Aim 1. K99 Phase: Determine the role of miR-302/367 in modulating the behaviors of lung epithelial progenitor cells during airway epithelium development and regeneration after injury. a) Determine the effects of conditional gain- and loss-of-miR-302/367 on lung epithelial progenitor cell development using ROSA-miR- 302/367 and floxed miR-302/367 mouse lines. b) Determine whether decreased expression of Wnt5a is responsible for the expansion of lung epithelial progenitors in miR-302/367 gain of function mutants. c) Determine the requirement of miR-302/367 for homeostasis and regeneration in airway epithelium. Specific Aim 2. R00 Phase: Determine the mechanisms underlying the regulation of epithelial progenitor cell behaviors by miR-302/367 in airway epithelium development and regeneration. a) Characterize miR-302/367 expressing cells in lung epithelium during development and airway regeneration after injury. b) Determine the direct targets and downstream signaling pathways regulated by miR-302/367 in developing lung. c) Determine whether modulation of specific miR-302/367-regulated targets or pathways would enhance airway epithelium repair and regeneration.