Mechanisms of beta-catenin signaling in alveolar epithelial cell differentiation

描述（由申请方提供）：肺泡上皮由两种不同的细胞群组成：II型（AT 2）和I型（AT 1）细胞。AT 2细胞作为兼性祖细胞用于维持和修复肺泡上皮，而AT 1细胞被认为是终末分化的。Wnt/β-连环蛋白信号在肺发育中起着双重作用，指导祖细胞的维持/扩增与分化。在任何细胞或组织环境（包括肺）中指定这些作用中的每一个的机制，特别是Wnt/β-连环蛋白依赖性信号传导在肺修复和成人肺泡上皮细胞（AEC）分化中的作用在很大程度上是未知的。我们最近开发的新型抑制剂差异调节b-连环蛋白和其转录辅激活因子p300或CREB结合蛋白（CBP）之间的相互作用，使我们提出了一个模型来解释Wnt/<$-连环蛋白信号传导的二分活性，其中CBP/<$-连环蛋白介导的转录对祖细胞的维持至关重要，而p300/<$-连环蛋白介导的转录对祖细胞的维持至关重要。β-连环蛋白介导的转录对于分化的起始和细胞效能的降低至关重要。本提案的总体目标是调查 Wnt/<$-catenin信号传导的作用，特别是<$-catenin与CBP的差异相互作用 和p300，在正常和损伤的成人肺中指导AEC分化和表型转变。β-连环蛋白/p300相互作用受p300磷酸化调节，而转录因子加塔-6（其促进AT 1细胞表型）通过与Wnt信号传导的相互作用控制发育/再生期间祖细胞维持与分化之间的平衡。我们假设，在正常和损伤的成人肺中，（1）β-连环蛋白/p300依赖性信号在AT 1细胞分化中起关键调节作用，（2）p300磷酸化调节β-连环蛋白/p300相互作用以促进AT 1细胞表型，（3）AT 1细胞表型的调节涉及Wnt 5a和加塔-6之间的相互作用。我们将利用我们良好表征的体外AEC分化模型和我们在表征AEC分化和遗传修饰小鼠方面的专业知识，结合我们的新型抑制剂，差异调节辅激活因子/β-连环蛋白相互作用，以解决以下具体目标：1）研究差异辅激活因子/β-连环蛋白相互作用在成体AEC体外分化过程中的作用; 2）表征成体AEC转分化过程中差异共激活因子/<$-连环蛋白相互作用的潜在机制;和3）探索体内成体AEC转分化中差异共激活因子/<$-连环蛋白相互作用。阐明这种发育相关的Wnt信号通路在促进正常上皮细胞分化中的作用的潜在机制对于理解肺损伤后的修复机制至关重要。促进正常AEC分化的独特小分子抑制剂（并且目前正在临床试验中评估）的可用性另外提供了这些发现的快速转化的可能性，以通过增强上皮修复来改善结果。

DESCRIPTION (provided by applicant): Alveolar epithelium is comprised of two distinct cell populations: type II (AT2) and type I (AT1) cells. AT2 cells serve as facultative progenitors for maintenance and repair of alveolar epithelium, while AT1 cells have been viewed as terminally differentiated. Wnt/¿-catenin signaling plays dichotomous roles in lung development, directing progenitor maintenance/expansion vs differentiation. Mechanisms that specify each of these roles in any cellular or tissue context (including the lung), and in particular the roles of Wnt/¿-catenin-dependent signaling in lung repair and adult alveolar epithelial cell (AEC) differentiation are largely unknown. Our recent development of novel inhibitors that differentially modulate interactions between b-catenin and either of its transcriptional coactivators, p300 or CREB binding protein (CBP), have led us to propose a model to explain dichotomous activities of Wnt/¿-catenin signaling, in which CBP/¿-catenin-mediated transcription is critical for progenitor cell maintenance while p300/¿-catenin-mediated transcription is critical for initiation of differentiation and decreased cellular potency. The overall goal of this proposal is to investigate the role of Wnt/¿-catenin signaling, and especially differential interactions of ¿-catenin with CBP and p300, in directing AEC differentiation and phenotypic transitions in normal and injured adult lung. ¿-catenin/p300 interactions are modulated by p300 phosphorylation, while the transcription factor GATA-6 (which promotes AT1 cell phenotype) controls the balance between progenitor maintenance vs differentiation during development/regeneration through interactions with Wnt signaling. We hypothesize that, in normal and injured adult lung, (1) ¿-catenin/p300-dependent signaling plays a critical regulatory role in AT1 cell differentiation, (2) p300 phosphorylation modulates b-catenin/p300 interactions to promote AT1 cell phenotype, and (3) regulation of AT1 cell phenotype involves interactions between Wnt5a and GATA-6. We will use our well-characterized models of AEC differentiation in vitro and our expertise in characterizing AEC differentiation and genetically modified mice, in conjunction with our novel inhibitors, to differentially regulate coactivator/¿-catenin interactions in order to address the following Specifc Aims: 1) Investigate the role of differential coactivator/b-catenin interactions during adult AEC differentiation in vitro; 2) Characterize mechanisms underlying differential coactivator/¿-catenin interactions during adult AEC transdifferentiation; and 3) Explore differential coactivator/¿-catenin interactions in adult AEC transdifferentiation in vivo. Elucidating mechanisms underlying the role of this developmentally relevant Wnt signaling pathway in promoting normal epithelial cell differentiation is critically important to understanding mechanisms of repair following lung injury. Availability of unique small molecule inhibitors that promote normal AEC differentiation (and that are currently being evaluated in clinical trials) additionally offers the possibility of apid translation of these findings to improve outcome by enhancing epithelial repair.