Role of Par1 Polarity Proteins in Podocyte Development and Glomerular Disease

描述（由申请人提供）：本提案描述了一个在足细胞生物学和肾小球疾病方面的学术生涯发展的五年计划。PI已经完成了美国国立卫生研究院支持的儿科和发育肾病学结构化奖学金培训项目，现在将通过跨部门资源的独特整合来扩展她的科学技能。该项目将促进利用分子遗传学和细胞生物学的专业知识来研究肾小球疾病中维持足细胞极性的机制。Drs。Katalin Susztak和Anne Muesch将指导PI的科学发展。她的主要导师Susztak博士是内科/肾脏病学副教授，也是她所在领域确定肾小球疾病新机制的领导者。内部共同导师Muesch博士是发育和分子生物学副教授，是建立细胞极性机制的专家，将在肾小球疾病的背景下对其进行研究。此外，为了扩大PI的研究范围并促进她对足细胞极性发育的研究，PI正在接受来自哥伦比亚大学附近的外部顾问Jonathan Barasch博士的胚胎肾脏培养实践培训。Drs。Susztak、Muesch、Barasch和其他几位备受尊敬的发育肾病学家将组成一个咨询委员会，提供科学和职业建议。研究将集中于顶基极性蛋白Par1a/b在肾脏发育过程中建立足细胞极性和在肾小球疾病中维持足细胞极性的作用。最近在她导师的指导下进行的工作表明，Par1a/b在发育中的肾单位以及啮齿动物和人类足细胞中表达。在培养的足细胞中，Par1a/b功能的显性负抑制诱导细胞形状的改变和狭口隔膜蛋白的表达改变，狭口隔膜蛋白是支持肾小球滤过结构和功能的关键足细胞成分。在糖尿病肾病、肾病综合征和肾小球硬化模型中，肾小球Par1a/b表达发生改变。拟议的实验将需要强力霉素诱导的，足细胞特异性抑制Par1a/b功能，使用体内小鼠模型，这将允许研究Par1a/b功能在肾细胞发育期间（胚胎发育期间的诱导）和成年小鼠。通过腺病毒感染抑制Par1a/b在胚胎肾培养中的功能，将进一步研究Par1a/b在体外的功能。此外，Par1a/b在肾病综合征和肾小球硬化实验模型中的表达，使用大鼠诱导的嘌呤霉素氨基核苷肾病（PAN），以及在人肾病综合征中，使用收集的对照和患病人肾脏标本。具体目的包括：研究Par1a/b在肾细胞发育过程中对足细胞分化的影响；2)研究Par1a/b在维持足细胞极化结构和肾小球滤过功能中的作用；3)研究Par1a/b在肾病综合征和肾小球硬化的人类和实验模型中的表达。

DESCRIPTION (provided by applicant): This proposal describes a five year program for development of an academic career in podocyte biology and glomerular disease. The PI has completed an NIH-supported, structured fellowship training program in Pediatric and Developmental Nephrology and will now expand upon her scientific skills through a unique integration of interdepartmental resources. This program will promote expertise in use of molecular genetics and cell biology to investigate mechanisms that maintain podocyte polarity in the setting of glomerular disease. Drs. Katalin Susztak and Anne Muesch will mentor the PI's scientific development. Her primary mentor, Dr. Susztak, is an Associate Professor of Internal Medicine/ Nephrology and a leader in her field in identifying novel mechanisms of glomerular disease. The internal co-mentor, Dr. Muesch, an Associate Professor in Development and Molecular Biology, is an expert in mechanisms that establish cell polarity, which will be examined in the context of glomerular disease. In addition, to expand the PI's repertoire and facilitate her study of the development of podocyte polarity, the PI is receiving hands-on training in embryonic kidney culture from an external advisor, Dr. Jonathan Barasch, who is located nearby at Columbia University. Drs. Susztak, Muesch, Barasch and several other highly regarded developmental nephrologists will form an advisory committee to provide scientific and career advice. Research will focus on the role of apical-basal polarity proteins Par1a/b in establishing podocyte polarity during kidney development and in maintaining podocyte polarity in the setting of glomerular disease. Recent work performed under the guidance of her mentors demonstrated that Par1a/b is expressed in developing nephrons and in rodent and human podocytes. Dominant negative suppression of Par1a/b function in cultured podocytes induced changes of cell shape and altered expression of slit diaphragm proteins, which are key podocyte components that support glomerular filter structure and function. Glomerular Par1a/b expression was altered in rodent models of diabetic nephropathy, nephrotic syndrome and glomerulosclerosis. The proposed experiments will entail doxycycline-inducible, podocyte-specific suppression of Par1a/b function using an in vivo mouse model, which will allow study of Par1a/b function during nephron development (induction during embyrogenesis) and in adult mice. Adenoviral infection of constructs suppressing Par1a/b function in embryonic kidney culture will be used to further study the function of Par1a/b in vitro. In addition, Par1a/b expression in an experimental model of nephrotic syndrome and glomerulosclerosis, using puromycin aminonucleoside nephrosis (PAN) induced in rats, and in human nephrotic syndrome, using a collection of control and diseased human kidney specimens. Specific aims include: Examine Par1a/b function during nephron development on podocyte differentiation, 2) Examine Par1a/b function in maintaining a polarized podocyte structure and glomerular filter function, and 3) Examine expression of Par1a/b in human and experimental models of nephrotic syndrome and glomerulosclerosis. PUBLIC HEALTH RELEVANCE: Chronic and end-stage kidney disease (CKD and ESKD) results in significant morbidity and mortality. We propose to examine novel pathways that may provide insight into mechanisms of proteinuric kidney disease and focal glomerulosclerosis (FSGS), a leading cause of childhood CKD and ESKD. We will examine the role of Par1a/b in establishing and maintaining podocyte structure and define polarity protein expression in human nephrotic syndrome and glomerulosclerosis, potentialy identifying new targets for therapeutic intervention or biomarkers for progressive kidney disease. !