Role of the cdx homeobox genes in the regulation of wt1 and nephron development

cdx 同源盒基因在 wt1 和肾单位发育调节中的作用

• 批准号: 7579971
• 负责人: Alan J Davidson
• 金额: $ 35.25万
• 依托单位: MASSACHUSETTS GENERAL HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2008
• 资助国家: 美国
• 起止时间: 2008-03-03 至 2012-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7579971
• 关键词: Affect; Animals; Apoptosis; C.I. Solvent Yellow 56; Cell Proliferation; Cells; Characteristics; Chemicals; Child; Chronic; Chronic Kidney Failure; Code; Congenital Abnormality; Data; Defect; Development; Distal; Electron Microscopy; Embryo; Enzymes; Exhibits; Filtration; Fishes; Gene Expression; Genes; Homeobox; Homeobox Genes; Infant; Intermediate Mesoderm; Kidney; Kidney Diseases; Knowledge; Lead; Light; Link; Mammals; Metanephric structure; Modeling; Molecular; Nephrons; Paraxial Mesoderm; Pathway interactions; Pattern; Play; Positioning Attribute; Pronephric structure; Proteins; Regulation; Reporter; Role; Side; Signal Transduction; Staging; Stream; Testing; Transcription factor genes; Transgenic Organisms; Tretinoin; Vitamin A; Wilms' tumor suppressor 1a; Work; Zebrafish; base; design; functional genomics; in vivo; insight; knock-down; malformation; mutant; nephrogenesis; new therapeutic target; podocyte; prevent; promoter; research study; small molecule; solute; spatiotemporal; transcription factor

DESCRIPTION (provided by applicant): Congenital kidney malformations are the leading cause of chronic renal failure in children. At present, we have a poor understanding of the developmental pathways that control the formation of the nephron, the key functional unit of the kidney. We are using zebrafish to elucidate the embryonic pathways that govern the subdivision of the nephron into functionally distinct segments. The zebrafish embryo is ideal for these studies as the intermediate mesoderm on either side of the trunk forms two simple linear nephrons. From a functional genomics approach we have isolated a number of renal transcription factor and solute transporter genes. The expression patterns of these genes subdivide the zebrafish nephron into distinct proximal and distal segments much like the mammalian nephron. Combinations of transcription factors define each segment suggesting a `renal transcription factor code' determines segment identity. This code is disrupted in embryos deficient in the homeobox transcription factor genes cdx1a and cdx4, which act as master regulators of the hox genes. Doubly-deficient embryos display abnormal hox expression patterns, a posterior shift in the position of the kidney, and a loss of distal tubules. In addition, cdx mutants exhibit expanded expression of Wilms' tumor suppressor-1a (wt1a), a transcription factor implicated in metanephros development and podocyte differentiation in mammals. The cdx genes are expressed in the trunk (paraxial mesoderm) adjacent to the intermediate mesoderm. Expression of raldh2, encoding an enzyme involved in retinoic acid (RA) synthesis, is expanded in the paraxial mesoderm of cdx-deficient embryos. Knock-down of Raldh2 in cdx4 mutants is able to rescue the kidney positioning defect, whereas in wild-types it down-regulates wt1a expression. These results suggest that aberrant RA signaling is responsible for the renal defects in cdx mutants and led us to examine the role of RA in nephron segmentation. Using DEAB, a chemical antagonist of Raldh2, we found that RA is required at multiple stages of nephrogenesis to induce podocytes and the proximal tubules, and to suppress distal tubule fates. Based on these data we hypothesize that the cdx-hox pathway acts upstream of raldh2 to restrict RA synthesis and that RA induces podocytes and proximal tubule fates by up-regulating genes such as wt1a. The aims of this proposal are designed to test this hypothesis. We will characterize the cellular and molecular nephron defects in cdx- and RA-deficient animals and use rescue experiments and knock-down studies to determine the epistatic relationships between the cdx, hox, and raldh2 genes. Using chemical and molecular antagonists of RA, as well as a RA-responsive transgenic reporter line, we will determine where and when RA signaling is required to induce proximal nephron fates. These studies will establish a link between the cdx-hox and RA pathways in the control of nephron segmentation, shed new light on renal birth defects, and will be useful for developing new therapies to treat, or prevent, chronic renal failure.Birth defects that affect the kidneys are the leading cause of chronic renal failure in infants and children, however at present, we have a poor understanding of the genes that control kidney formation. We have found that retinoic acid, a derivative of Vitamin A, plays a central role in regulating normal renal development. This work advances our understanding of how kidneys arise in the embryo and may be lead to new therapeutic targets to treat or prevent chronic renal failure.

描述(由申请人提供)： 先天性肾畸形是儿童慢性肾功能衰竭的主要原因。目前，我们对控制肾脏的关键功能单位--肾单位的形成的发育途径知之甚少。我们正在使用斑马鱼来阐明控制肾单位细分为功能不同部分的胚胎途径。斑马鱼胚胎是这些研究的理想对象，因为躯干两侧的中间中胚层形成两个简单的线性肾单位。通过功能基因组学方法，我们分离了一些肾脏转录因子和溶质转运蛋白基因。这些基因的表达模式将斑马鱼的肾单位细分为不同的近端和远端片段，很像哺乳动物的肾单位。转录因子的组合定义了每个片段，表明‘肾脏转录因子编码’决定了片段的同一性。在同源盒转录因子基因cdx1a和cdx4缺失的胚胎中，这一密码被破坏，这两个基因是HOX基因的主要调节者。双缺陷胚胎表现出异常的HOX表达模式，肾脏位置后移，远端小管丢失。此外，CDX突变体显示Wilms肿瘤抑制因子-1a(Wt1a)的扩大表达，这是一种与哺乳动物后肾发育和足细胞分化有关的转录因子。CDX基因在与中间中胚层相邻的树干(旁中胚层)中表达。编码维甲酸(RA)合成的一种酶raldh2的表达在CDX缺陷胚胎的中胚层旁轴扩大。在cdx4突变体中敲除Raldh2能够挽救肾脏定位缺陷，而在野生型中Raldh2下调wt1a的表达。这些结果表明，RA信号的异常是CDX突变体肾脏缺陷的原因，并使我们研究了RA在肾单位分割中的作用。利用Raldh2的化学拮抗剂Deab，我们发现在肾脏发生的多个阶段都需要RA来诱导足细胞和近端小管，并抑制远端小管的命运。基于这些数据，我们假设CDX-HOX途径作用于raldh2上游，以限制RA的合成，并且RA通过上调wt1a等基因诱导足细胞和近端小管的命运。这项提议的目的是为了检验这一假设。我们将描述CDX和RA缺陷动物的细胞和分子肾单位缺陷，并使用挽救实验和击倒研究来确定CDX、Hox和raldh2基因之间的上位关系。利用RA的化学和分子拮抗剂，以及RA反应转基因报告系，我们将确定在哪里以及何时需要RA信号来诱导近端肾单位的命运。这些研究将在CDX-HOX和RA通路之间建立控制肾单位分段的联系，为肾脏出生缺陷提供新的线索，并将有助于开发治疗或预防慢性肾功能衰竭的新疗法。出生缺陷影响肾脏是婴儿和儿童慢性肾功能衰竭的主要原因，然而，目前我们对控制肾脏形成的基因知之甚少。我们发现维生素A的衍生物维甲酸在调节正常肾脏发育中起着核心作用。这项工作促进了我们对肾脏是如何在胚胎中形成的理解，并可能导致治疗或预防慢性肾功能衰竭的新的治疗靶点。