Defining the mechanism of Clara cell dedifferentiation into basal stem cells

总结 近年来，祖细胞和干细胞的定义越来越受到关注 群体的肺和解剖的分子机制，管理 这些细胞在气道再生和修复过程中的行为。最近的研究在多个 不同的器官系统已经证明， 由于祖细胞和干细胞群体比以前更具可塑性， 认可.然而，损伤后气道上皮细胞的可塑性一直不佳 记录在案。 白喉毒素诱导的细胞死亡与遗传谱系追踪相结合， 用于严格证明皮肤中不同类型的上皮干细胞， 肠可以相互转化。同样的策略组合也表明， 分化的肠腔上皮细胞可以“去分化”为肠干细胞， 细胞我们目前的初步数据表明，管腔克拉拉细胞可以转换， 转化为气道基底干细胞。 我们通过只表达白喉毒素（DTA）来消融气道基底干细胞， 基底细胞，导致这些基底细胞发生凋亡。针对这一 当基底干细胞耗尽时，管腔Clara细胞祖细胞开始复制， 这些Clara细胞开始表达基底细胞特异性标记p63和细胞角蛋白5。 此外，分选至纯度并离体培养的Clara细胞失去了它们的生物活性。 特征性Clara细胞标志物，开始表达Yap 1转录共激活因子， 然后表达对基底干细胞特异的标记。这些去分化的基底样 然后细胞开始复制并产生菌落。我们假设卢米纳尔·克拉拉 当基底干细胞库被激活时，细胞祖细胞可以转化为基底干细胞。 耗尽了为了验证这一假设，我们将谱系追踪Clara细胞的贡献， DTA介导的气道基底干细胞消融后的基底干细胞库。然后我们将 测试去分化的Clara细胞是否具有功能性干细胞能力。我们 并推测Yap 1转录辅激活因子是Clara细胞所必需的 去分化我们将使用小鼠遗传学来测试Yap 1的表达是否是 足够和/或必需的克拉拉细胞可塑性。

Summary Recently, increasing attention has been paid to defining the progenitor and stem cell populations of the lung and dissecting the molecular mechanisms that govern the behavior of these cells during airway regeneration and repair. Recent studies in multiple different organ systems have demonstrated that well-defined differentiated cells as well as progenitor and stem cell populations are much more plastic than previously recognized. However, the plasticity of airway epithelial cells after injury has not been well documented. Diphtheria toxin-induced cell death in combination with genetic lineage tracing has been used to stringently demonstrate that distinct types of epithelial stem cells in the skin and intestine can interconvert. The same pairing of strategies has also demonstrated that differentiated luminal intestinal epithelial cells can "dedifferentiate" into intestinal stem cells. We present preliminary data that suggests that a luminal Clara cell can convert into an airway basal stem cell following injury. We ablated airway basal stem cells by expressing diphtheria (DTA) toxin exclusively in basal cells, which causes those basal cells to undergo apoptosis. In response to this depletion of basal stem cells, luminal Clara cell progenitors begin replicating and then these Clara cells begin to express the basal cell-specific markers p63 and cytokeratin 5. Furthermore, Clara cells that are sorted to purity and cultured ex vivo lose their characteristic Clara cell markers, begin expressing the Yap1 transcriptional co-activator, and then express markers specific to basal stem cells. These dedifferentiated basal-like cells then start replicating and give rise to colonies. We hypothesize that luminal Clara cell progenitors can convert into basal stem cells when the basal stem cell pool is depleted. To test this hypothesis, we will lineage trace the contribution of Clara cells to the basal stem cell pool after DTA-mediated airway basal stem cell ablation. We will then test whether the dedifferentiated Clara cells possess functional stem cell capacity. We also hypothesize that the Yap1 transcriptional co-activator is necessary for Clara cell dedifferentiation. We will use mouse genetics to test whether Yap1 expression is sufficient and/or necessary for Clara cell plasticity.