miRNAs regulate lung epithelial progenitor cells in development and injury repair

描述(申请人提供)：肺上皮祖细胞对于发育和成人呼吸道的再生和修复是必不可少的。控制其行为的机制，包括发育过程中的增殖和分化以及呼吸道损伤修复的进展，在很大程度上尚不清楚。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仍然很少 明白了。最近，我们已经证明miR-302/367是GATA6的直接靶点，GATA6是肺发育和呼吸道再生所必需的转录因子，它调节肺上皮祖细胞增殖和分化的平衡，以及S细胞的尖基两极。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。在发育中的呼吸道上皮中，miR-302/367的过度表达导致Sox2+近端和Sox9+远端祖细胞的扩张，并降低了呼吸道上皮细胞的分化。值得注意的是，我们最近的研究表明，miR-302/367在萘损伤的肺组织中的表达明显高于对照未损伤的肺组织，这表明miR-302/367在介导肺上皮细胞基因转录和呼吸道修复和再生过程中的损伤反应中具有潜在的作用。因此，我在这个建议中的工作假设是miR-302/367作为GATA6依赖的调控网络的组成部分，涉及多个正负反馈环来调节肺上皮祖细胞的增殖和分化。更好地了解miR-302/367在呼吸道上皮细胞发育和再生过程中如何调节肺上皮祖细胞的行为，将为多种肺部疾病提供重要的见解。这将通过追求两个特定的目标来实现：特定的目标1.K99阶段：确定miR-302/367在调节肺上皮祖细胞在呼吸道上皮细胞发育和损伤后再生过程中的作用。A)使用ROSA-miR-302/367和FLOXED的miR-302/367小鼠系，确定条件获得和丢失miR-302/367对肺上皮祖细胞发育的影响。B)确定Wnt5a表达降低是否与miR-302/367肺上皮祖细胞的扩增有关。C)确定miR-302/367对呼吸道上皮细胞动态平衡和再生的需求。特定目的2.R00期：确定miR-302/367在呼吸道上皮细胞发育和再生中调控上皮祖细胞行为的机制。A)鉴定miR-302/367在肺上皮细胞发育和损伤后呼吸道再生过程中的表达。B)确定miR-302/367在发育肺中调控的直接靶点和下游信号通路。C)确定调节特定miR-302/367调节的靶点或通路是否会促进呼吸道上皮的修复和再生。

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.