Creating high-resolution multi-omics molecular atlases for developing urogenital organs

创建用于发育泌尿生殖器官的高分辨率多组学分子图谱

• 批准号: 10491224
• 负责人: FENG CHEN
• 金额: $ 46.64万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-30 至 2026-07-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10491224
• 关键词: 3-Dimensional; Ablation; Age; Anatomy; Atlases; Cell Nucleus; Cells; Cellular Structures; Childhood; Communities; Computer Analysis; Data; Databases; Development; Dimensions; Disease; Disease model; Epigenetic Process; Fertilization; Gene Expression; Gene Silencing; Genes; Genitourinary system; Human; Imaging technology; Infrastructure; Institution; Kidney; Light; Lower urinary tract; Lymphatic; Maps; Mesenchyme; Microscopy; Modeling; Molecular; Morbidity - disease rate; Mus; Nerve; Organ; Organ Model; Penetration; Phenotype; Population; Procedures; Process; Prostate; Reference Values; Reproducibility; Research; Research Personnel; Resolution; Signal Transduction; Small Nuclear RNA; Structure; System; Technology; Time; Ureter; Ureteropelvic junction obstruction; Urinary tract; Work; analysis pipeline; biological systems; body system; congenital anomalies of the kidney; epigenetic profiling; experience; experimental study; male; microscopic imaging; mortality; mouse model; multiple omics; multiplexed imaging; new technology; prenatal; progenitor; reproductive organ; transcriptome; transcriptome sequencing; transcriptomics; tumor

The GenitoUrinary Development Molecular Anatomy Project (GUDMAP) has been providing valuable references for the research community studying urogenital development and diseases. It is a recurring theme that rapidly advancing new technologies are instrumental in enhancing and expanding reference databases. Cross fertilization of atlas building efforts spanning various organ systems and disease types will undoubtedly boost the technology penetration across these projects. To build multi-dimensional atlases of developing urogenital organs that incorporate the latest multi-omics and spatial molecular mapping technologies, we have assembled a team with expertise both in urogenital development and multi-dimensional, multi-platform, molecular atlas building. We propose to utilize the infrastructure we developed at our institution for the NCI Human Tumor Atlas Network (HTAN) and other large scale projects as a springboard to help effectively and efficiently propel GUDMAP to the next level with transcriptome-wide coverage, single cell level resolution, and spatial mapping with unprecedented clarity. We will take advantage of our experience in the incorporation of single nucleus (sn) RNA-seq and snATAC-seq to establish a comprehensive epigenetic and transcriptomic landscape in targeted urogenital organs and structures (lower urinary tract (LUT), selected male reproductive organs, kidney vasculature, lymphatics, and nerves) at single cell resolution (Aim 1). We will then add the spatial dimension to this molecular landscape to build 2D and 3D molecular atlases by incorporating spatial transcriptomics (ST), CODEX, and light sheet microscopy (LSM) (Aim 2). In Aim 3, we will extend our study to disease models, focusing on murine models of congenital anomalies of the kidney and the urinary tract (CAKUT). With the proposed experiments, we aim at building multidimensional molecular atlases for developing urogenital organs at unprecedented cellular resolution and gene coverage with the highest efficiency possible. Aim 1: Characterize the epigenetic and transcriptomic landscapes of developing urogenital organs with single cell omics We will perform integrated transcriptomic and epigenetic profiling of the developing/maturing LUT, male reproductive organs, and the kidney at E16.5, NB, and 3 weeks of age. Although the focus for the kidney will be on vasculature, lymphatics, and nerves, since there is a lack of single cell omics data on most of the selected stages, our data will also help to strengthen GUDMAP data for the broadest use by the research community. Aim 2: Construct multi-dimensional molecular atlases for developing urogenital organs using spatial transcriptomics and advanced imaging technologies A major challenge for atlas building in biological systems has been spatially assigning large number of molecular features to the anatomical and cellular structures. We have successfully established experimental procedures and computational analyses pipelines for spatial transcriptomics, CODEX, and light sheet microscopic imaging, to map gene expression data, including transcriptome-wide data to cells and structures. We will use these technologies to analyze the developing/maturing lower urinary tract, male reproductive organs, and the kidney at E16.5, NB, and 3 weeks of age for the construction of truly multi-dimensional, multiplatform molecular atlases. Aim 3: Building molecular atlases for key urogenital structures using murine CAKUT models with cell ablation or gene inactivation CAKUT occurs in many different forms representing a significant cause of morbidity and mortality in the pediatric population. We have generated and analyzed several CAKUT murine models in the past. Building atlases of the target organs for these models will provide high resolution, spatially registered molecular references for key stages of disease initiation and progression. Moreover, such atlases will help researchers better understand normal urogenital development by knowing the tolerance of the systems and processes in dealing with various disturbances. We will use a highly reproducible murine model of CAKUT with inactivation of canonical Smad signaling in ureteral mesenchyme, causing a uniform ureteropelvic junction (UPJ) obstruction phenotype prenatally. We will use the technologies outlined in Aims 1 and 2 to build molecular atlases of the relevant structures (ureter, kidney, UPJ) at key time points and compare the atlases of defective development with those of normal development.

泌尿生殖发育分子解剖学项目（GUDMAP）一直为研究泌尿生殖发育和疾病的研究界提供有价值的参考。快速发展的新技术有助于增强和扩展参考数据库，这是一个反复出现的主题。跨越不同器官系统和疾病类型的图集构建工作的交叉施肥无疑将促进这些项目的技术渗透。为了构建融合最新多组学和空间分子图谱技术的泌尿生殖器官发育多维图谱，我们组建了一支在泌尿生殖发育和多维、多平台分子图谱构建方面具有专业知识的团队。我们建议利用我们在 NCI 人类肿瘤图谱网络 (HTAN) 和其他大型项目中开发的基础设施作为跳板，以帮助有效和高效地将 GUDMAP 推向新的水平，实现全转录组覆盖、单细胞水平分辨率，以及前所未有的清晰度空间映射。我们将利用我们在单核 (sn) RNA-seq 和 snATAC-seq 结合方面的经验，在目标泌尿生殖器官和结构（下尿路 (LUT)、选定的男性生殖器官、单细胞分辨率下的肾脏脉管系统、淋巴管和神经）（目标 1）。然后，我们将通过结合空间转录组学 (ST)、CODEX 和光片显微镜 (LSM)，将空间维度添加到该分子景观中，以构建 2D 和 3D 分子图谱（目标 2）。在目标 3 中，我们将把研究扩展到疾病模型，重点关注肾脏和泌尿道先天性异常 (CAKUT) 的小鼠模型。通过所提出的实验，我们的目标是建立多维分子图谱，以前所未有的细胞分辨率和基因覆盖率以尽可能高的效率发育泌尿生殖器官。 目标 1：利用单细胞组学表征发育中泌尿生殖器官的表观遗传和转录组学景观 我们将对 E16.5、NB 和 3 周龄时发育/成熟的 LUT、雄性生殖器官和肾脏进行综合转录组和表观遗传分析。尽管肾脏的重点将放在脉管系统、淋巴管和神经上，但由于大多数选定阶段缺乏单细胞组学数据，我们的数据也将有助于加强 GUDMAP 数据，以供研究界最广泛使用。 目标 2：利用空间转录组学和先进成像技术构建用于发育泌尿生殖器官的多维分子图谱 在生物系统中构建图谱的一个主要挑战是在空间上将大量分子特征分配给解剖和细胞结构。我们已经成功建立了空间转录组学、CODEX 和光片显微成像的实验程序和计算分析管道，以绘制基因表达数据，包括转录组范围的数据到细胞和结构。我们将使用这些技术来分析 E16.5、NB 和 3 周龄时发育/成熟的下尿路、男性生殖器官和肾脏，以构建真正的多维、多平台分子图谱。 目标 3：使用细胞消融或基因失活的小鼠 CAKUT 模型构建关键泌尿生殖结构的分子图谱 CAKUT 以多种不同形式发生，是儿科人群发病和死亡的重要原因。我们过去已经生成并分析了多个 CAKUT 小鼠模型。为这些模型构建靶器官图谱将为疾病发生和进展的关键阶段提供高分辨率、空间注册的分子参考。此外，这样的图谱将帮助研究人员通过了解系统和过程在处理各种干扰时的耐受性，更好地了解正常的泌尿生殖发育。我们将使用高度可重复的 CAKUT 小鼠模型，使输尿管间充质中的经典 Smad 信号失活，从而在产前引起均匀的肾盂输尿管连接部 (UPJ) 阻塞表型。我们将利用目标1和目标2中概述的技术来构建关键时间点相关结构（输尿管、肾脏、UPJ）的分子图谱，并将发育缺陷的图谱与正常发育的图谱进行比较。