Transcriptional Regulatory Mechanisms of Salivary Gland Branching Morphogenesis

项目摘要 在胚胎发育过程中，增殖和分化之间的微妙平衡 唾液腺的干细胞/祖细胞(SP)必须受到严格的调控，以维持动态平衡。 这对正常的组织发育和再生至关重要。因此，更好地理解 驱动SP细胞功能的调节机制将提供对唾液腺的基本见解 形态发生，并帮助确定对损伤后组织再生至关重要的途径 在患病的状态下。转录因子p63，特别是Δnp63亚型，在 富含上皮性组织的SP细胞群，包括皮肤和腺体器官，如唾液 腺体(SG)。P63缺失小鼠的戏剧性表型突显了p63在SG中的关键重要性， 它们显示了组织形态发生的一个完整的障碍。尽管p63在SG中的某些功能 发展已有报道，我们目前对p63在早期发育中的作用的了解 SG的形态发生和分化阶段，包括分枝形态发生，是相当有限的。在……里面 特别是，p63决定细胞命运的分子机制以及它如何控制特定的 缺乏早期SG分枝过程中的基因表达程序。因此，确定ΔNP63驱动的 调控网络，特别是在分枝形态发生的背景下，将是必不可少的第一步 在我们对SG干细胞/祖细胞生物学的理解中，我们的长期目标是开发新的策略 用于再生和重新设计富含上皮性器官(如SG)的复杂分支结构。 为了解决这些知识差距，我们将利用小鼠模型和复杂的基因组方法来研究 三个独立的兴趣领域。首先，我们将使用条件性基因敲除小鼠模型来确定角色 ΔNp63在涎腺分支形态发生(AIM1)中的表达这样的研究是非常需要的，因为他们将发现 首次研究了p63在SG分枝形态发生中的功能作用。第二，我们将执行机械化 研究确定由ΔNp63控制的关键靶基因和通路，并确定如何丢失 Δ的Np63改变了全球基因表达程序，并影响了细胞在分枝过程中的命运轨迹 单细胞分辨率形态发生(AIM2)。最后，我们将利用体外组织外植体来检测其作用。 Wnt信号通路，特别是sFRP1，在介导ΔNp63的功能中的作用，并检测sFRP1是否 能够恢复在p63丢失时观察到的分枝缺陷。总体而言，这些研究具有 对进一步了解ΔNp63依赖的转录和细胞网络具有重要意义 对于SG的生物学功能，特别是当它与分枝有关时。我们对这一现象的理解取得了进展 推动SG分支和发展的潜在机制可以为两者的再生提供范例 老鼠和人类的SGS和更好的信息疗法来治疗发育障碍和 SG的疾病。

Project Summary During embryonic development, the delicate balance between proliferation and differentiation of stem/progenitor (SP) cells of the salivary gland must be tightly regulated to maintain a homeostatic balance, which is critical for normal tissue development and regeneration. Therefore, a better understanding of the regulatory mechanisms driving SP cell function will provide fundamental insights into salivary gland morphogenesis and help identify pathways that are critical for tissue regeneration following injury, damage, or during diseased states. The transcription factor p63, specifically the ΔNp63 isoform, is highly expressed in the SP cell populations of epithelial rich tissues including those of the skin and glandular organs such as the salivary gland (SG). The critical importance of p63 in the SG is highlighted by the dramatic phenotype of p63-null mice, which display a complete block in tissue morphogenesis. Although some aspects of p63 function in SG development have been reported, our current knowledge regarding the role of p63 in the early developmental stages of SG morphogenesis and differentiation, including branching morphogenesis, is rather limited. In particular, the molecular mechanisms through which p63 dictates cell fate decisions and how it controls specific gene expression programs during early SG branching, is lacking. Therefore, identifying the ΔNp63-driven regulatory networks, particularly in the context of branching morphogenesis, will serve as an essential first step in our understanding of SG stem/progenitor cell biology with the long-term goals of developing new strategies for regeneration and for re-engineering the complex branching architecture of epithelial rich organs like the SG. To address these knowledge gaps, we will utilize mouse models and sophisticated genomic approaches to study three independent areas of interest. First, we will use conditional knockout mouse models to determine the role of ΔNp63 in salivary gland branching morphogenesis (Aim1). Such studies are much needed as they will identify for the first time, the functional role of p63 in SG branching morphogenesis. Second, we will perform mechanistic studies to identify critical target genes and pathways that are governed by ΔNp63 and determine how the loss of ΔNp63 alters the global gene expression program and affects cell fate trajectories during branching morphogenesis at single cell resolution (Aim2). Finally, we will utilize ex vivo tissue explants to examine the role of the WNT signaling pathway, and specifically Sfrp1, in mediating the function of ΔNp63 and test whether Sfrp1 is able to restore the branching defects observed upon the loss of p63. Collectively, these studies have the potential to further our understanding of the ΔNp63-dependent transcriptional and cellular networks important for the biological function of the SG particularly as it pertains to branching. Advances in our understanding of the underlying mechanisms driving SG branching and development can provide a paradigm for regeneration of both mouse and human SGs and better inform therapies to treat patients with developmental dysfunctions and diseases of the SG.