Functional Analysis of the Role of Transcription Factor 21 (Tcf21) in Nephron Progenitor Cells in Congenital Anomalies of the Kidney and Urinary Tract

项目摘要/摘要 这是Gal Finer，医学博士，博士，助理，K08职业发展奖的申请表 西北大学范伯格医学院教授。此申请者曾接受过临床培训 儿科肾病和免疫学、遗传学和分子生物学方面的研究培训。她的长期工作 职业目标是成为一名独立资助的研究人员，拥有肾脏开发方面的专业知识。这 提案将使申请者获得发育生物学和相关实验方面的高级技能 方法：研究方法。为了实现这些目标，芬纳博士组建了一支拥有专业知识的多学科指导团队 参加校外资助的研究，她将参加课程、研讨会和会议。 这个职业发展奖中包含的项目的广泛的、长期的目标是破译 导致先天性肾和尿路畸形(CAKUT)的分子机制 确定新的治疗靶点。CAKUT是前三位慢性肾脏疾病的主要原因 这与健康儿童的存活率低30倍有关。然而， CAKUT的发病机制知之甚少，在缺乏预防或治疗的情况下， 护理主要依靠透析和移植。肾单位祖细胞是一种多能细胞。 这对于确保出生时正常的肾单位数量和正常的肾脏大小至关重要。已经证明，在 正常发育，这些细胞在自我更新和谱系特异性分化之间取得平衡 (NPC动力学)。然而，人们对控制这些“干细胞样”细胞的基本机制知之甚少。 夸金实验室已经确定了一种新的bHLH转录因子TCF21，它对肾脏发育至关重要。 我们初步表明，TCF21控制Cited1和WNT4的表达，这两个β-连环蛋白靶标是 参与鼻咽癌的自我更新和分化。这一提议的中心假设是TCF21控制 调控Wnt/β-catenin信号通路的鼻咽癌动力学。我们将利用TCF21独特的小鼠遗传模型- 表现CAKUT谱型肾发育不良的耗竭的鼻咽癌细胞与TCF21在鼻咽癌中的作用无关 差异化。我们的具体目标是破译TCF21调节鼻咽癌的分子机制。 为了达到我们的目标，我们将进行β-连环蛋白途径的分子分析，遗传育种的分析， 肾源性生态位的形态分析、谱系追踪和单细胞RNA测序。 这项工作将由肾脏和高血压科主任苏珊·夸金博士指导 以及西北大学范伯格学院心血管和肾脏研究所的主任。Dr。 夸金是一位享誉国际的发展性肾病专家，具有丰富的指导经验。这个 这项提议的单细胞RNA分析将由黛博拉·温特博士指导，因为她在 基因组研究中的计算生物学和生物信息学。该项目还将受益于特殊的 夸金博士实验室和西北大学范伯格医学院的研究环境。

PROJECT SUMMARY/ABSTRACT This is an application for a K08 Career Development Award for Gal Finer, MD, PhD, an Assistant Professor at the Feinberg School of Medicine of Northwestern University. This applicant has clinical training in Pediatric Nephrology and research training in immunology, genetics, and molecular biology. Her long-term career goal is to become an independently funded researcher with expertise in renal development. This proposal will position the applicant to gain advanced skills in developmental biology and pertinent experimental methods. To achieve these goals, Dr. Finer has assembled a multidisciplinary mentoring team with expertise in extramurally funded research, and she will be participating in coursework, seminars, and conferences. The broad, long-term objectives of the project included in this career development award are to decipher the molecular mechanisms that cause congenital anomalies of the kidney and urinary tract (CAKUT) and to identify novel therapeutic targets. CAKUT is the leading cause of chronic kidney disease in the first three decades of life and is associated with survival rate that is 30 times lower than that of healthy children. However, very little is known about the pathogenesis of CAKUT, and in the absence of prevention or curative treatment, care relies mainly on dialysis and transplantation. The nephron progenitor cells (NPCs) are multipotent cells that are critical for ensuring normal nephron number and normal kidney size at birth. It has been shown that, in normal development, these cells strike a balance between self-renewal and lineage-specific differentiation (NPC dynamics). However, the basic mechanisms that control these “stem-like” cells are poorly understood. The Quaggin lab has identified a novel bHLH transcription factor, Tcf21, that is critical for kidney development. We preliminary show that Tcf21 controls the expression of Cited1 and Wnt4, two β-catenin targets that are involved in NPC self-renewal and differentiation. The central hypothesis of this proposal is that Tcf21 controls NPC dynamics by regulating Wnt/β-catenin signaling. We will utilize unique genetic mouse models of Tcf21- depleted NPCs that present CAKUT-spectrum renal dysplasia to dissociate the effects of Tcf21 on NPC differentiation. Our Specific Aims are to decipher the molecular mechanisms by which Tcf21 regulates NPCs. To address our aims, we will perform molecular analysis of the β-catenin pathway, analysis of genetic breeding, morphological analysis of the nephrogenic niche, lineage tracing, and single-cell RNA-sequencing. This work will be mentored by Dr. Susan Quaggin, Head of the Division of Nephrology and Hypertension and the Director of the Feinberg School Cardiovascular and Renal Institute at Northwestern University. Dr. Quaggin is an internationally acclaimed developmental nephrologist with extensive mentoring experience. The single-cell RNA analysis of this proposal will be mentored by Dr. Deborah Winter for her expertise in computational biology and bioinformatics in genomic studies. This project will also benefit from the exceptional research environment of Dr. Quaggin’s lab and the Northwestern University Feinberg School of Medicine.