Gene regulatory networks in the proximal tubules of the mammalian kidney

哺乳动物肾脏近端小管的基因调控网络

• 批准号: 10031038
• 负责人: Joo-Seop Park
• 金额: $ 48.19万
• 依托单位: CINCINNATI CHILDRENS HOSP MED CTR
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-06-09 至 2023-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10031038
• 关键词: ATAC-seq; Address; Adult; Binding; CCL21 gene; Carrier Proteins; Cell Differentiation process; Cells; ChIP-seq; Chromatin; DNA; Data; Development; Developmental Process; Distal; Down-Regulation; Engineering; Epithelial Cells; Exhibits; Filtration; Gene Expression; Genes; Genetic Models; Glycolysis; Goals; HNF4A gene; Hand; Henle' s loop; Human; Injury; Kidney; Knowledge; Maintenance; Mammalian Cell; Maps; Molecular; Mus; Mutant Strains Mice; Na(+)-K(+)-Exchanging ATPase; Natural regeneration; Nephrons; Pathogenicity; Phenotype; Physiological; Play; Polydipsia; Polyuria; Production; Public Health; Pump; Regulator Genes; Renal corpuscle structure; Replacement Therapy; Research; Role; Sodium; Specific qualifier value; Syndrome; Testing; Tissue-Specific Gene Expression; Tissues; Water; blood filtration; cell type; clinically relevant; fatty acid oxidation; genome-wide; improved; kidney cell; loss of function; loss of function mutation; mutant; nephrogenesis; nuclease; progenitor; single-cell RNA sequencing; small molecule; solute; sound; stem cells; tool; transcription factor; transcriptome sequencing

Project Summary/Abstract Nephron segments are composed of specific types of epithelial cells that perform distinct physiological functions and collectively act as a blood filtration unit. Along the proximo-distal axis, the renal corpuscle is followed by the proximal tubule (PT), loop of Henle, and distal tubule. To successfully engineer kidney cells/tissue for replacement or to stimulate nephron regeneration, it is essential to understand (1) how the different segments of the nephron are specified during development and (2) how their segmental identities are maintained in the adult kidney. We propose to use Hnf4a, a transcription factor specifically expressed in PT cells, as an entry point to study mammalian nephron segmentation. One of the major functions of the PT is reabsorption of water and vital small molecules from the filtrate. Defective PT function causes Fanconi renotubular syndrome (FRTS), characterized by polydipsia, polyuria, and glucosuria and loss-of-function of HNF4A gene causes FRTS in humans. We found that, in mice, the deletion of Hnf4a specifically in the nephron lineage caused massive downregulation of PT-specific genes, disruption in PT development, and an FRTS-like phenotype. These results strongly suggest that Hnf4a plays a key role in PT development. Hnf4a is the most highly expressed transcription factor in adult PT cells in the mouse kidney. Our analysis of publicly available PT-specific ATAC-seq data in the mouse adult kidney shows that the most highly enriched DNA motif in open chromatin domains in PT cells is the Hnf4a motif. Taken together, these results suggest that Hnf4a may serve as a master regulator in adult PT cells. In order to better understand the functions of Hnf4a in adult PT cells, we have mapped genome-wide binding of Hnf4a in mouse adult PT cells. Our preliminary data show that Hnf4a directly binds to genes involved in fatty acid oxidation (FAO) and also SLC genes encoding solute carrier proteins. These two sets of genes are critical for reabsorption, the major function of PT cells. To better understand how Hnf4a regulates the specification of PT cells during nephrogenesis, we will determine the role of Hnf4a in early stages of PT development and identify direct target genes of Hnf4a. To investigate if Hnf4a plays a role in the maintenance/function of adult PT cells, we will test if Hnf4a regulates the expression of FAO and SLC genes in adult PT cells. We will also test if Hnf4a is required for the maintenance of open chromatin regions in adult PT cells. Our proposed studies will fill a longstanding gap of knowledge in the molecular mechanisms underlying specification and maintenance of PT cells of the mammalian nephron and significantly improve our understanding of the pathogenic mechanisms of FRTS.

项目摘要/摘要 肾单位段由特定类型的上皮细胞组成，这些上皮细胞执行不同的生理功能 功能，并共同充当血液过滤单元。沿近远轴线，肾小体 其次是近端小管(PT)、Henle环和远端小管。成功改造肾脏 为了替代细胞/组织或刺激肾单位再生，有必要了解(1) 肾单位的不同节段是在发育过程中指定的，以及(2)它们的节段身份是如何确定的 维持在成人肾脏中。我们建议使用HNF4a，一种在PT中特异表达的转录因子 细胞，作为研究哺乳动物肾单位分割的切入点。工商局的主要职能之一是 从滤液中重新吸收水和重要的小分子。PT功能缺陷导致Fanconi 肾小管综合征(FRTS)，以多饮、多尿、糖尿和肾功能减退为特征 HNF4a基因会导致人类的FRTS。我们发现，在小鼠中，HNF4a基因的缺失特异性地存在于肾单位 血统导致PT特异基因的大量下调，PT发育中断，以及类似FRTS的 表型。这些结果有力地表明HNF4a在甲状旁腺功能亢进的发生中起关键作用。HNF4a是最多的 转录因子在小鼠肾脏成体PT细胞中的高表达。我们对公众可获得的数据的分析 小鼠成年肾脏中PT特异的ATAC-seq数据表明，在开放的 PT细胞中的染色质结构域是HNF4a基序。综上所述，这些结果表明HNF4a可能对 作为成年PT细胞的主调节器。为了更好地了解HNF4a在成年PT细胞中的功能， 我们已经定位了HNF4a在小鼠成年PT细胞中的全基因组结合。我们的初步数据显示， HNF4a直接与脂肪酸氧化相关基因(FAO)以及编码溶质的SLC基因结合 载体蛋白。这两组基因对PT细胞的主要功能--重吸收至关重要。为了更好地 了解HNF4a在肾脏形成过程中如何调节PT细胞的规格，我们将确定其作用 并鉴定HNF4a的直接靶基因。调查HNF4a是否 在成年甲状旁腺细胞的维持/功能中起作用，我们将测试HNF4a是否调节FAO的表达 和成体PT细胞中的SLC基因。我们还将测试是否需要HNF4a来维持开放染色质 成体甲状旁腺细胞的区域。我们提出的研究将填补分子方面长期存在的知识空白。 哺乳动物肾单位PT细胞的特性和维持机制 提高对FRTS发病机制的认识。