Diversity-generating mechanisms for mammalian intercalated cell lineages

项目总结/摘要 75万美国人患有终末期肾病（ESRD）。他们唯一的治疗选择是 透析，长期生存前景不佳，或治愈性肾移植，肾脏太少，无法满足 病人的需要。解决可移植肾脏短缺的一种方法是深入了解 基本的细胞类型以及它们是如何产生的，然后利用这种洞察力来产生功能性的，基于细胞的 换肾手术肾脏的生理相关上皮网络来自两种不同的细胞， 祖先群体肾单位祖细胞产生肾单位，其中大约有一百万个 在人肾脏中，邻近的输尿管祖细胞产生分支网络到收集系统， 与肾单位相连体内平衡由肾单位和肾细胞中的特化细胞类型维持。 集合管值得注意的是，水和盐以及pH平衡由主细胞（PC）调节，并插入 电池（IC）。有三种类型的闰细胞：A型闰细胞（A-ICs）、B型闰细胞 插入细胞（B-IC）和非A-非B插入细胞，在调节 pH稳态。有趣的是，McMahon小组最近证明了PC和IC都有双重起源。 来自肾单位和输尿管祖细胞，这是一个不寻常的发育过程。在这里，我的目的是确定 从不同的肾祖细胞群体产生相似但不相同的IC亚型的机制。在 具体目标1，我将集中于确定转录因子的目标，这些转录因子是已知的关键， 芯片Tfcp 2l 1和Foxi 1的开发。我还将分析转录因子的靶点， Dmrt 2将肾单位和输尿管上皮来源的IC-A细胞类型与 表达Hmx 2/Hmx 3的IC-B细胞类型。差异Hmx基因表达细分了IC-B群体： Hmx 2 + B-IC和非A-非B IC是肾单位来源的，Hmx 2 +/Hmx 3 + B-IC是输尿管谱系来源的。 我们假设Tfcp 2l 1和Foxi 1启动了一般IC发育程序，而Dmrt 2，Hmx 2， Hmx 3在IC亚型的产生中具有相互抑制作用。我将使用生物化学方法， 产生标记的等位基因，以鉴定这些转录因子在两种细胞的IC内的调节相互作用。 血统在具体目标2中，我将使用体内遗传方法和输尿管类器官模型来确定 这些因子在产生哺乳动物IC的功能性转录相互作用中的需求。采取 总之，这项拟议的工作将确定一个调控框架内的嵌入细胞的发展， 肾脏中的输尿管和肾单位谱系，并提供了新的见解， 参与哺乳动物肾脏功能的细胞类型。

Project Summary/Abstract Three-quarters of a million Americans have End Stage Renal Disease (ESRD). Their only treatment options are dialysis, with poor long term survival prospects, or a curative kidney transplant, with too few kidneys to meet patient need. One way to address the shortage of transplantable kidneys is to develop a thorough insight into essential cell types and how they are generated, and then to use this insight to generate functional, cell-based kidney replacements. The physiologically relevant epithelial networks of the kidney arise from two distinct progenitor populations. Nephron progenitors generate the nephrons, of which there are approximately one million in the human kidney, and adjacent ureteric progenitors generate the branched network to the collecting system, to which nephrons connect. Homeostasis is maintained by specialized cell types in both the nephron and the collecting duct. Notably, water and salt, and pH balance, are regulated by principal cells (PCs) and intercalated cells (ICs), respectively. There are three subtypes of intercalated cells: type A intercalated cells (A-ICs), type B intercalated cells (B-ICs) and non-A-non-B intercalated cells that perform distinct functions in the regulation of pH homeostasis. Interestingly, the McMahon group recently demonstrated both PCs and ICs have dual origins from both nephron and ureteric progenitor cells, an unusual developmental process. Here, I aim to identify the mechanisms generating similar, but not identical, IC subtypes from distinct kidney progenitor populations. In Specific Aim 1, I will focus on identifying the targets of transcription factors that are known to be essential for the development of ICs: Tfcp2l1 and Foxi1. I will also profile the targets of transcription factors that have not yet been described in ICs: Dmrt2 distinguishes both nephron and ureteric epithelial derived IC-A cell types from Hmx2/Hmx3 expressing IC-B cell types. Differential Hmx gene expression sub-divides the IC-B populations: Hmx2+ B-ICs and non-A-nonB ICs are nephron-derived and Hmx2+/Hmx3+ B-ICs are ureteric lineage derived. We hypothesize that Tfcp2l1 and Foxi1 initiate a general IC developmental program, while Dmrt2, Hmx2, and Hmx3 have mutually repressive actions in the generation of IC subtypes. I will use biochemical approaches and generate tagged alleles to identify regulatory interactions for these transcription factors within ICs of both lineages. In Specific Aim 2, I will use genetic approaches in vivo and a ureteric organoid model to determine the requirement of these factors in the functional transcriptional interplay generating mammalian ICs. Taken together, this proposed work will define a regulatory framework for the development of intercalated cells within the ureteric and nephron lineages in the kidney and provide new insights into the regulatory programs governing cell types involved in mammalian kidney function.