3D imaging and deep sequencing of gene expression in the genital tubercle

生殖结节基因表达的 3D 成像和深度测序

基本信息

批准号：
8926970
负责人：
MARTIN J COHN
金额：
$ 29.45万
依托单位：
UNIVERSITY OF FLORIDA
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-09-30 至 2016-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8926970
关键词：
Affect Arts Atlases Cell Therapy Cells Child Cloaca Chamber Colon Colorectal Communities Congenital Abnormality Data Detection Development Developmental Anatomy Distal Endocrine Disruptors Endoderm Endoderm Cell Epithelial Cells Epithelium Exposure to Failure Foundations Future Gene Expression Gene Expression Profile Generations Genes Genetic Genetic Programming Genetic Screening Genetic study Genital system Genitalia Genitourinary system Goals Human Hypospadias Image Incidence Knowledge Limb Bud Limb Development Link Live Birth Maps Mesenchyme Modeling Molecular Molecular Profiling Morphogenesis Natural regeneration Noise Operative Surgical Procedures Optics Outcome Pattern Perinatal Exposure Play Population Protein Isoforms Public Health RNA Sequences RNA Splicing Reagent Relative (related person)Research Role Staging Stem cells Technology Three-Dimensional Imaging Tissues Transcript Tube Urethra Urothelium Variant ambiguous genitalia base cell type deep sequencing digital external genitalia gene discovery gene function innovation malformation molecular marker mouse model novel novel marker penis penis foreskin prepuce progenitor repaired research study tomography tool transcriptome sequencing urologic

项目摘要

DESCRIPTION (provided by applicant): The most common anomaly of the genitourinary system is hypospadias, a malformation of the external genitalia that is characterized by failure of urethral tube closure and incomplete formation of the prepuce (foreskin) and ventral penis. The incidence of hypospadias has more than doubled over the past 30 years, without explanation, and it is now the second most common birth defect in humans, affecting an estimated 1 of every 250 live births. Whether the causes of hypospadias are genetic, environmental, or a combination of both, development of preventative strategies for hypospadias and of cell-based therapies for repair of urologic tissues will require knowledge of the genetic programs that direct cell fate decisions and tissue morphogenesis. Identification of these mechanisms first requires the establishment of a foundation of descriptive data, in the form of gene expression patterns, for the different tissue compartments of the lower genitourinary tract. Moreover, transcriptional profiling will need to be linked to developmental anatomy in order to formulate hypotheses regarding gene function during morphogenesis, and to consider how multiple genes may interact within a tissue or cell population. In preliminary studies using microarrays, we identifie 86 novel markers of the urethral epithelium of the El 2.5 genital tubercle. Here we propose to use OPT to build a 3D reference model of the genital tubercle, and to map the expression domains of each of these genes onto the reference tubercle to create a 3D, multi-gene atlas of the gene expression in the developing external genitalia. We will build upon these results in two ways. Firstly, we will extend the OPT atlas to two additional stages of external genital development. Secondly, we will drill deeper into the expression profiling by using NexGen sequencing technology (RNA-seq) to characterize the quantitative patterns of gene expression and the distribution of splice variants in the endodermal cells fated to form the urethral epithelium. This combination of approaches will provide a wealth of new cell type-specific markers, uncover the quantitative molecular profiles of urethral progenitors, and place these data in a 3D developmental anatomical context that can be utilized by the research community for future hypothesis-based studies.

描述（由申请人提供）：泌尿生殖系统最常见的异常是尿道下裂，这是一种外生殖器畸形，其特征是尿道下裂。尿道管闭合以及包皮和阴茎腹侧形成不完整。过去 30 年来，尿道下裂的发病率在没有任何解释的情况下增加了一倍多，现在是人类第二常见的出生缺陷，估计每 250 名活产儿中就有 1 人受到影响。无论尿道下裂的原因是遗传、环境还是两者的结合，开发尿道下裂的预防策略和修复泌尿系统组织的细胞疗法都需要了解指导尿道下裂的遗传程序。细胞命运决定和组织形态发生。识别这些机制首先需要为下泌尿生殖道的不同组织区室以基因表达模式的形式建立描述性数据的基础。此外，转录谱需要与发育解剖学联系起来，以便提出有关形态发生过程中基因功能的假设，并考虑多个基因如何在组织或细胞群内相互作用。在使用微阵列的初步研究中，我们鉴定了 El 2.5 生殖结节尿道上皮的 86 个新标记。在这里，我们建议使用 OPT 构建生殖器结节的 3D 参考模型，并将每个基因的表达域映射到参考结节上，以创建发育中外生殖器中基因表达的 3D 多基因图谱。我们将以两种方式巩固这些成果。首先，我们将OPT图谱扩展到外生殖器发育的另外两个阶段。其次，我们将使用 NexGen 测序技术 (RNA-seq) 更深入地研究表达谱，以表征基因表达的定量模式以及注定形成尿道上皮的内胚层细胞中剪接变体的分布。这种方法的组合将提供大量新的细胞类型特异性标记，揭示尿道祖细胞的定量分子特征，并将这些数据置于 3D 发育解剖学背景中，研究界可以将其用于未来基于假设的研究。