Molecular modulators of polycystin signaling

多囊蛋白信号传导的分子调节剂

• 批准号: 10561693
• 负责人: STEFAN SOMLO
• 金额: $ 42.4万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-03-01 至 2024-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10561693
• 关键词: ATAC-seq; Ablation; Address; Adult; Affect; Affinity Chromatography; Antisense Oligonucleotides; Apoptosis; Attenuated; Autosomal Dominant Polycystic Kidney; Biological Assay; Biology; Cell Culture System; Cell Culture Techniques; Cell Cycle Regulation; Cell Line; Cell Proliferation; Cells; ChIP-seq; Chromatin; Chromatin Structure; Cilia; Complex; Consensus; Cyst; Data; Dialysis procedure; Disease; Dissociation; Down-Regulation; GLIS2 gene; Gene Dosage; Genes; Genetic; Genetic Diseases; Genetic Transcription; Genotype; Growth; In Vitro; Inherited; Investigation; Kidney; Kidney Diseases; Knowledge; Maps; Modeling; Molecular; Mutation; Oxidative Phosphorylation; PKD2 protein; Pathogenesis; Pathway interactions; Patients; Polycystic Kidney Diseases; Post-Translational Protein Processing; Pre-Clinical Model; Proliferating; Property; Proteins; Proteolytic Processing; Renal function; Research; Resolution; Ribosomes; Role; Signal Pathway; Signal Transduction; Time; Tissues; Transgenic Organisms; Translating; Transplantation; Transposase; Up-Regulation; differential expression; dosage; early onset; expectation; field study; human disease; improved; in vitro Model; in vivo; in vivo Model; innovation; mouse model; mutant; novel; novel therapeutics; optimal treatments; overexpression; polycystic liver disease; programs; targeted treatment; therapeutic target; trafficking; transcription factor; transcriptome; transcriptome sequencing; transcriptomics

Autosomal dominant polycystic kidney disease (ADPKD) is a common genetic disease that results from mutations in either of two proteins, polcysytin-1 (PC1) or polycystin-2 (PC2). More than two decades have passed since the genes encoding these proteins were discovered and there has been significant progress in understanding the functions of polycystins and their associated disease. Nonetheless, there remain substantial gaps in knowledge and lack of consensus about the precise functions of PC1 or PC2 and the mechanisms of ADPKD. Resolution of these gaps is of great significance given our expectation that optimal therapies for ADPKD are best developed based on the fundamental understanding of polycystin function in the mammalian kidney. Much of the current mechanistic understanding of polycystin function is based on studies of candidate pathways drawn from amongst known cellular mechanisms associated with functions such as differentiation, proliferation, transport and signaling. The lack of coalescence toward an interrelated unifying functional pathway in polycystin biology and the persistence of gaps in understanding of in vivo polycystin function despite extensive investigation suggests that the critical components of the most proximal polycystin signaling cascade have yet to be identified. Indeed, the polycystins were discovered as complex, entirely novel proteins and it stands to reason that they may function in a signaling pathway that is not among those that are currently well understood or studied. We made use of this concept in by applying an unbiased in vivo transcriptomic study using Translating Ribosome Affinity Purification (TRAP) RNASeq. From this, we identified upregulation of cell cycle and down regulation of oxidative phosphorylation as key pathways alterations in vivo. Among these, we found genotype dependent upregulation of a cilia associated transcription factor, Glis2, not previously considered to function in polycystin signaling or ADPKD pathogenesis. We made double mutants of Pkd1 with Glis2 in early onset and adult models and found Glis2 dosage-dependent rescue of cyst formation in both. Based on these findings we propose that Glis2 is a candidate for a downstream effector of PC1 function that is critical for cyst progression in ADPKD. In this study, we will determine the in vivo mechanism of action of Glis2 and establish its effects on cyst cell proliferation, apoptosis and ADPKD due to Pkd2. We will determine whether Glis2 is a target for therapy through both genetic and pharmacotherapeutic studies. We will assess whether in vivo genotype dependent transcriptional changes we have identified are similarly extended to cell culture systems with Pkd mutant genotypes. We will also evaluate the functional properties of Glis2 protein in Pkd mutant cell lines. In aggregate, these studies open a new direction of investigation for polycystin signaling and ADPKD pathogenesis.

常染色体显性遗传性多囊肾病(ADPKD)是一种常见的遗传性疾病，由 两种蛋白质中的一种突变，多囊蛋白-1(PC1)或多囊蛋白-2(PC2)。二十多年过去了 自从编码这些蛋白质的基因被发现以来，在 了解多囊藻毒素的功能及其相关疾病。尽管如此，仍然有大量的 对PC1或PC2的确切功能以及PC2的机制认识上的差距和缺乏共识 ADPKD.鉴于我们期望ADPKD的最佳治疗方法 是基于对哺乳动物肾脏中多囊蛋白功能的基本了解而开发的。 目前对多囊蛋白功能的许多机制理解都是基于对候选通路的研究 从已知的与分化、增殖、 运输和信号发送。多囊蛋白缺乏向相互关联的统一功能通路的结合 生物学和体内多囊蛋白功能认识上的差距，尽管进行了广泛的研究 这表明，最近端多囊蛋白信号级联的关键成分尚未确定。 事实上，多囊藻毒素被发现是一种复杂的、全新的蛋白质，理所当然地它们 可能在目前尚未被充分了解或研究的信号通路中发挥作用。我们 利用这一概念，通过使用翻译核糖体进行无偏见的体内转录研究 亲和纯化(TRAP)RNAseq.由此，我们确定了细胞周期的上调和细胞周期的下调。 氧化磷酸化是体内改变的关键途径。其中，我们发现基因依赖于基因。 纤毛相关转录因子Glis2的上调，此前未被认为在多囊蛋白中起作用 信号转导或ADPKD发病机制。我们在早期发病和成年模型中用Glis2对PKD1进行了双突变 并发现Glis2对两者的囊性形成均有剂量依赖性的挽救作用。基于这些发现，我们建议 Glis2是PC1功能下游效应的候选者，PC1功能对ADPKD的囊性进展至关重要。在……里面 在本研究中，我们将确定Glis2的体内作用机制，并建立其对囊性细胞的作用 PKD2引起的细胞增殖、细胞凋亡和ADPKD我们将通过以下方式确定Glis2是否为治疗目标 包括遗传和药物治疗研究。我们将评估体内的基因型是否依赖于 我们已经确定的转录变化类似地延伸到具有PKD突变的细胞培养系统 基因分型。我们还将评估Glis2蛋白在PKD突变细胞系中的功能特性。总而言之， 这些研究为多囊蛋白信号转导和ADPKD发病机制的研究开辟了新的方向。