Cell Fate Decisions in Epithelial Stem Cell Lineages

R35赠款摘要 我的实验室对了解细胞分化是如何在一个连续的 更新上皮组织这些组织的保守特征尚未完全了解，并统一研究 跨实验系统包括一个灵活的利基结构，“过境放大”阶段， 具有显著的细胞可塑性，相邻干细胞谱系竞争小生境的能力 占用率为了理解组织的这些新特性，我们专注于允许 用于在细胞分辨率下研究天然体内环境中的细胞行为。我们的主要模型 系统是果蝇卵巢的滤泡上皮，我们最近将研究扩展到了 小鼠肠上皮我们在过去十年的贡献包括确定来源， 卵泡干细胞（FSC）小生境配体的身份，定义了FSC的自我更新网络，描述了新的 促进干细胞和子细胞命运分离的机制， 作为理解干细胞生态位竞争的模型。我们目前的研究是 调查三个相互关联的领域首先，我们制作了果蝇卵巢的细胞图谱， 身份，位置和基因表达谱的十几个已知的和新的细胞类型。这个项目 提供了有用的新工具，使我们能够研究组织中细胞的谱系可塑性， 例如，导致发现小生境细胞可以在生理应激期间转化为干细胞。在 此外，这些工具为我们提供了一个新的机会，研究动态种群的小生境细胞是如何能够 以在不断变化的组织需求中维持稳定的FSC库。第二，我们正在研究 控制干细胞生态位竞争的机制。我们已经确定了一大类等位基因， 我们正在利用遗传学、定量成像和数学建模， 理解选择一个血统而不是另一个的基础。我们还将这些研究扩展到 小鼠肠上皮细胞，并发现该过程至少部分保守。第三，我们是 研究细胞内pH（pHi）在调节细胞命运决定中的作用。我们证明了pHi 在FSC谱系和小鼠胚胎干细胞的分化过程中增加， pHi升高是分化所必需的。在未发表的研究中，我们发现了一个类似的要求， 增加小鼠肠干细胞谱系中的pHi。目前，我们专注于了解pHi 调节细胞命运，重点是候选的“pH传感器”蛋白，如β-连环蛋白，具有pKa 在生理范围内。对于这些蛋白质来说，质子的获得或丢失就像翻译后的蛋白质一样。 修饰，从而将pH动态与可能影响细胞命运的蛋白质活性变化联系起来。通过研究 这些涌现出的特性，在良好的特点果蝇和小鼠上皮细胞，我们正在获得详细的 对正常组织稳态过程的深入了解，将为更好地理解 在衰老和疾病期间组织内稳态是如何失效的。

R35 GRANT ABSTRACT My lab is broadly interested in understanding how cellular differentiation is controlled within a continuously renewing epithelial tissue. Conserved features of these tissues that are not fully understood and unify studies across experimental systems include a flexible niche structure, a “transit amplification” stage which typically has significant cellular plasticity, and the ability of neighboring stem cell lineages to compete for niche occupancy. To understand these emergent properties of tissues, we have focused on approaches that allow for the study of cell behaviors within the native, in vivo context in at cellular resolution. Our primary model system is the follicle epithelium of the Drosophila ovary, and we have recently extended our studies into the mouse intestinal epithelium. Our contributions over the past ten years include identifying the source and identity of the follicle stem cell (FSC) niche ligands, defining a self-renewal network for FSCs, describing new mechanisms that promote the segregation of stem cell and daughter cell fates, and the establishment and use of the FSC lineage as a model for understanding stem cell niche competition. Our current studies are investigating three interconnected areas. First, we created a cell atlas of the Drosophila ovary that describes the identity, position, and gene expression profile of over a dozen known and novel cell types. This project has provided useful new tools that are allowing us to investigate the lineage plasticity of cells in the tissue and has led, for example, to the discovery that niche cells can convert to stem cells during physiological stress. In addition, these tools provide us with a new opportunity to study how a dynamic population of niche cells is able to maintain a stable pool of FSCs amid changing tissue demands. Second, we are investigating the molecular mechanisms that govern stem cell niche competition. We have identified a broad class of alleles that cause hypercompetition for the niche, and we are using genetics, quantitative imaging, and mathematical modeling to understand the basis for selection of one lineage over another. We have also extended these studies into the mouse intestinal epithelium and found that the process is at least partially conserved. Third, we are investigating the role of intracellular pH (pHi) in regulating cell fate decisions. We demonstrated that pHi increases during differentiation in both the FSC lineage and mouse embryonic stem cells, and that this increase in pHi is necessary for differentiation. In unpublished studies, we discovered a similar requirement for increased pHi in the mouse intestinal stem cell lineage. Currently, we are focused on understanding how pHi regulates cell fate, with an emphasis on candidate “pH sensor” proteins, such as β-catenin, that have a pKa within the physiological range. For these proteins, the gain or loss of a proton functions like a post-translational modification, thus linking pH dynamics to changes in protein activities that may affect cell fate. By studying these emergent properties within well-characterized Drosophila and mouse epithelia, we are gaining detailed insights into the process of normal tissue homeostasis that will provide a foundation for a better understanding of how tissue homeostasis fails during aging and disease.