Biological mechanisms of developmental stem cell plasticity

干细胞是组织发育、维持和再生的基本驱动力。为了确保这些过程的完整性，干细胞在分子身份和命运决定方面表现出可塑性，必须仔细控制。干细胞身份最好在基因和蛋白质表达水平上表征;尽管对整合多种生物过程的细胞器（如溶酶体）的影响知之甚少。溶酶体是细胞自噬（细胞的“废物处理系统”）的公认效应器。我们对这种细胞器功能的更广泛的理解仍在展开，尽管最近出现了已知影响细胞命运决定的过程的联系，包括代谢，表观遗传和细胞信号调节。因此，阐明溶酶体影响的干细胞群体和相应机制将填补关键的知识空白，并开辟新的发现途径。 我最近发现，神经干细胞的初始特化以溶酶体的突然激活为标志，这在亲本多能干细胞或发育相关的神经嵴细胞（NCC）中没有观察到。随后，NSC群体在溶酶体水平上表现出异质性，因为它们扩增、成熟并开始做出自我更新或分化的命运决定。提示维持“中等”溶酶体含量的关键要求，强制溶酶体活化至“高”水平导致功能失调的NSC表型和加速分化。因此，微调的溶酶体活化似乎对NSC身份规范和正常谱系发育都至关重要。我和我的团队将检验溶酶体表达动态调节NSC身份的采纳、维持和可塑性的假设。我的研究计划的首要目标是阐明干细胞身份的关键生物调节因子及其对细胞命运可塑性的影响。在接下来的五年里，我的团队将开始研究溶酶体在干细胞身份规范和谱系发育中的作用，使用我建立的二维（2D）多能和3D类器官干细胞模型系统。小鼠胚胎外植体将提供二次验证。我们将重点关注NSC和NCC，但也将研究其他干细胞群体。目的：1.定义干细胞鉴别质量标准中溶酶体的要求。2.阐明溶酶体活化对谱系发育的影响。3.研究溶酶体影响干细胞命运的生物学机制。这项工作将推进溶酶体生物学和控制干细胞身份和命运可塑性的过程的基础知识。它还将揭示易处理的细胞因子，为翻译研究人员开发组织再生策略和识别干细胞完整性与功能障碍的生物标志物。

Stem cells are essential drivers of tissue development, maintenance, and regeneration. To ensure integrity in these processes, stem cells exhibit plasticity in the molecular identities and fate decisions they can adopt, which must be carefully controlled. Stem cell identity is best characterized at the level of gene and protein expression; though the impact of cellular organelles like lysosomes, which integrate multiple biological processes, is poorly understood. Lysosomes are well-established effectors of autophagy, the cell's "waste disposal system". Our understanding of the wider breadth of function of this organelle is still unfolding, though links to processes that are known to impact cell fate decisions have recently emerged, including metabolic, epigenetic, and cell signaling regulation. Thus, elucidating the stem cell populations and corresponding mechanisms that lysosomes impact would fill a critical knowledge gap and open up new avenues for discovery. I recently discovered that the initial specification of NSCs is marked by a sudden activation of lysosomes, which is not observed in parental pluripotent stem cells or developmentally related neural crest cells (NCCs). Subsequently, NSC populations exhibit heterogeneity in lysosome levels as they expand, mature, and begin making fate decisions to self-renew or differentiate. Suggesting a critical requirement for maintenance of `moderate' lysosome content, forced lysosome activation to `high' levels leads to dysfunctional NSC phenotypes and accelerated differentiation. Finely tuned lysosome activation thus appears to be critical for both NSC identity specification and normal lineage development. My team and I will test the hypothesis that lysosome expression dynamics regulate the adoption, maintenance, and plasticity of NSC identity. The over-arching goal of my research program is to elucidate the critical biological regulators of stem cell identity and their impacts on cell fate plasticity. Over the next five years my team will begin to investigate the role of lysosomes in stem cell identity specification and lineage development, using my established two-dimensional (2D) pluripotent and 3D organoid stem cell model systems. Mouse embryo explants will provide secondary validation. We will focus on NSCs and NCCs, but additional stem cell populations will also be investigated. OBJECTIVES: 1. Define the requirement for lysosomes in stem cell identity specification. 2. Elucidate the impacts of lysosome activation on lineage development. 3. Investigate the biological mechanisms by which lysosomes impact stem cell fate. This work will advance basic knowledge on lysosome biology and the processes that control stem cell identity and fate plasticity. It will also reveal tractable cellular factors for translational researchers developing tissue regeneration strategies and identifying biomarkers of stem cell integrity versus dysfunction.