Defining molecular signals that influence airway basal cell renewal and growth

项目总结 哺乳动物呼吸道的基底细胞是呼吸道假性复层上皮的驻留干细胞。 而了解它们是如何被监管和维护的，对于理解它们的异常将是重要的 疾病的增长，如慢性阻塞性肺病、囊性纤维化和癌症，以及开发新的治疗方法 为再生医学的目的利用BCS。河马信号通路已经成为一种 BC生长的重要调节器。河马信号控制转录调节因子YAP和YAP的活性 TAZ(YAP/TAZ)通过限制核YAP/TAZ在成熟分化的呼吸道细胞中的定位。核武大杂烩 活动已被证明可以促进呼吸道中BC的生长，并已被证明在 其他组织和器官中干细胞群体的多样性。YAP与维持呼吸道BC命运有关 通过与基底细胞特异性转录因子p63直接结合。气道基底部干细胞发挥作用 肺损伤修复的重要作用，但调节其生长和自我更新的信号很差 明白了。我们的研究以及其他研究表明，转录调控因子YAP的核活动 在呼吸道基底细胞生长中起重要作用，在这项建议中，我们的目标是了解 在这方面对YAP活动的监管。在这项提案中，我们的目标是使用遗传小鼠模型，该模型包含 有条件地敲除HIPPO通路，调节气道基底细胞中的LATS1和LATS2，以 了解河马不活动和随后的YAP核活动如何通过其 与p63的相互作用。我们的初步数据表明，Krt5阳性基底细胞中LATS1/2缺失导致 基底细胞增生，形成不同的基底细胞群。靠近地下室的那些 膜上有最高水平的核p63，因此这种表达在Krt5+细胞中减少或丢失 在增生性病变中更多的是管腔。我们的研究进一步表明，BCS中的YAP活性受到影响 通过信号，如FGF10，来自基底膜中的基质细胞，我们假设 这些信号影响YAP与p63的相互作用，以指导BC自我更新转录程序。我们的目标是 揭示描述基底膜信号如何转导的新的生化途径 通过BC受体酪氨酸激酶增加YAP-P63的功能活性，促进BC的自我更新和生长。 具体地说，我们建议：1)确定LATS1/2-耗竭和随后的YAP激活是否在基础气道中 细胞通过YAP-P63相互作用影响BC的维持；2)基底膜的信号如何 可能促进这些YAP/P63的相互作用；以及3)YAP-P63复合体是否差异调节基因表达 来促进公元前的命运。最终，这项研究将提供关于BC自我更新和扩张的分子洞察力 这可能为预防疾病中BC的扩展提供方向或指导新的方法来扩展 BCS体外再生治疗。

PROJECT SUMMARY Basal cells (BCs) of mammalian airways are resident stem cells of the pseudostratified epithelium of the airways and understanding how they are regulated and maintained will be important for understanding their aberrant growth in disease, such as COPD, cystic fibrosis and cancer, as well as for developing novel methods for harnessing BCs for the purposes of regenerative medicine. The Hippo signaling pathway has emerged as an important regulator of BC growth. Hippo signaling controls the activity of the transcriptional regulators YAP and TAZ (YAP/TAZ) by restricting nuclear YAP/TAZ localization in mature differentiated airway cells. Nuclear YAP activity has been shown to promote BC growth in the airways and has been shown to play important roles in a variety of stem cell populations in other tissues and organs. YAP is implicated in maintaining the airway BC fate through direct association with the basal cell specific transcription factor, p63. Airway basal stem cells play important roles lung injury repair, but the signals that regulate their growth and self-renewal are poorly understood. Our studies, as well as others, indicate that the nuclear activity of the transcriptional regulator YAP plays important roles in airway basal cell growth and in this proposal, we aim to understand the roles and regulation of YAP activity in this context. In this proposal we aim to use genetic mouse models which contain conditional knockouts of Hippo pathway regulating kinases, LATS1 and LATS2, in airway basal cells to understand how Hippo inactivity and subsequent YAP nuclear activity influences BC maintenance through its interaction with p63. Our preliminary data suggest that LATS1/2 deletion in Krt5 positive basal cells leads to basal cell hyperplasia and the formation of distinct populations of basal cells. Those adjacent to the basement membrane have the highest levels of nuclear p63, whereby this expression is reduced or lost in the Krt5+ cells that are more luminal in the hyperplastic lesion. Our studies further suggest that YAP activity in BCs is influenced by signals, such as FGF10, that come from stromal cells in the basement membrane, and we hypothesize that these signals influence YAP interactions with p63 to guide a BC self-renewal transcriptional program. We aim to unveil novel biochemical pathways that describe how signals from the basement membrane are transduced through BC receptor tyrosine kinases to increase YAP-p63 functional activity and BC self-renewal and growth. Specifically, we propose: 1) to determine if LATS1/2-depletion and subsequent YAP activation in airway basal cells influences BC maintenance through YAP-p63 interactions; 2) how signals from the basement membrane may promote these YAP/p63 interactions; and 3) if YAP-p63 complexes differentially regulate gene expression to promote BC fate. Ultimately this study will offer molecular insight into how BC self-renewal and expansion is regulated, which may offer directions for preventing BC expansion in disease or guide new methods to expand BCs ex vivo for regenerative therapy.