Defining signaling networks in epithelial homeostasis

摘要 本提案中工作大纲的目标是了解控制细胞命运的基本信号， 维持组织内环境稳定。上皮组织表现出其组成细胞的内在能力， 组织和维持一个稳定的形式和功能状态。许多疾病状态缺乏这些内在控制。 人体的上皮组织处于不断更新的状态。我们对信号的理解 控制这些复杂的上皮组织如何保持强健组织的系统是不完整的。基本 为了取得进展，我们需要在单细胞水平上对信号传导进行定量理解， 生理条件，以揭示系统水平的行为，可以有针对性的治疗。我们专注于 蛋白激酶作为细胞中信号传导的关键介质，众所周知，其在细胞内的细胞凋亡中发挥重要作用。 组织功能和疾病的驱动因素。我们的全激酶组研究已经确定GSK3和CLK3是主要的 控制增殖和分化之间平衡的组织内稳态调节剂。在项目1中， 我们假设GSK3需要多种抑制性输入，这些输入独特地产生不同的命运结果， 包括干细胞、过渡扩增和分化。我们将提供第一个系统级映射 GSK3动力学的多个输入以及这些动力学如何解码成不同的细胞结果 更新上皮的过程。在项目2中，我们假设CLK3是通过以下途径控制干细胞命运的看门人： 转录组调控我们将确定干细胞龛中的CLK3活性作为表达调节因子， 以及剪接Wnt靶基因以促进干性。我们的方法使用高通量定量 显微镜以测量生理类器官稳态培养模型中的单细胞行为。我们 研究将确定两种关键激酶GSK3和CLK3的调节机制，并发现新的 更新和再生上皮细胞的准确组织所需的信号电路，揭示新的 治疗再生组织疾病的策略。

ABSTRACT The goal of the work outline in this proposal is to understand the fundamental signaling that controls cell fate to maintain tissue homeostasis. Epithelial tissues demonstrate an intrinsic ability for their constituent cells to organize and maintain a steady-state of form and function. Many disease states lack these intrinsic controls. Epithelial tissues of the human body are in a constant state of renewal. Our understanding of the signaling systems that control how these complex epithelial tissues maintain robust organization is incomplete. Essential for progress, we need a quantitative understanding of signaling at the single-cell level in the context of physiological conditions to reveal systems-level behaviors that can be targeted therapeutically. We focus on protein kinases as critical mediators of signaling in the cell, which are well known to play prominent roles in tissue function and drivers of disease. Our kinome-wide studies have identified GSK3 and CLK3 as major tissue homeostasis regulators that govern the balance between proliferation and differentiation. In Project 1, we hypothesize GSK3 requires multiple suppressive inputs that uniquely produce different fate outcomes ranging from stem cells, transit-amplifying, and differentiated. We will provide the first systems-level mapping for multiple inputs onto GSK3 dynamics and how these dynamics are decoded into distinct cellular outcomes of renewing epithelium. In Project 2, we hypothesize CLK3 is a gatekeeper controlling stem cell fate through transcriptome regulation. We will define the activity of CLK3 in the stem cell niche as a regulator of expression and splicing of Wnt-target genes to promote stemness. Our approach uses high-throughput quantitative microscopy to measure single-cell behaviors in physiological organoid homeostatic culture models. Our research will define the regulatory mechanisms for two critical kinases, GSK3 and CLK3, and discover novel signaling circuitry needed for the accurate organization of renewing and regenerative epithelia, uncovering new strategies for treating diseases of regenerative tissues.