Molecular modulators of polycystin signaling

多囊蛋白信号传导的分子调节剂

• 批准号: 10078607
• 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/10078607
• 关键词: ATAC-seq; Address; Adult; Affect; Affinity Chromatography; Antisense Oligonucleotides; Apoptosis; Attenuated; Autosomal Dominant Polycystic Kidney; Biological Assay; Biology; Cell Culture System; Cell Culture Techniques; Cell Cycle; Cell Line; Cell Proliferation; Cells; ChIP-seq; Chromatin; Chromatin Structure; Cilia; Complex; Consensus; Cyst; Data; Dialysis procedure; Disease; 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; Property; Proteins; Proteolytic Processing; Renal function; Research; Resolution; Ribosomes; Role; Signal Pathway; Signal Transduction; Structure; Time; Tissues; Transgenic Organisms; Translating; Transplantation; Transposase; Up-Regulation; Vitelliform macular dystrophy; base; 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.

常染色体显性多囊性肾脏疾病（ADPKD）是一种常见的遗传疾病，由 两种蛋白质中的突变，polcysytin-1（PC1）或polycystin-2（PC2）。超过二十年过去了 由于发现了编码这些蛋白质的基因，并且在 了解多囊素及其相关疾病的功能。尽管如此，仍然存在实质性 知识差距和缺乏关于PC1或PC2的精确功能的共识以及 ADPKD。鉴于我们期望对ADPKD的最佳疗法，解决这些差距具有很大的意义 最好基于对哺乳动物肾脏多囊这功能的基本理解而开发的。 当前对多囊菌功能的机械理解的许多理解是基于对候选途径的研究 从与分化，增殖等功能相关的已知细胞机制中得出 运输和信号传导。缺乏朝着polycystin中相互关联的统一功能途径的合并 尽管进行了广泛的研究 表明尚未识别出最近端多囊蛋白信号传导级联的关键成分。 实际上，多囊这被发现是复杂的，完全是新颖的蛋白质，这是有理由认为它们 可能在当前不理理解或研究的信号通路中起作用。我们 通过使用翻译核糖体应用无偏的体内转录组研究来利用此概念 亲和力纯化（TRAP）RNASEQ。由此，我们确定了细胞周期的上调和调节 氧化磷酸化作为体内的关键途径改变。其中，我们发现基因型依赖 纤毛相关的转录因子Glis2的上调，以前不考虑在多囊这 信号传导或ADPKD发病机理。我们在早期和成人模型中用GLIS2制成了PKD1的双突变体 并发现GLIS2剂量依赖于两者的囊肿形成。根据这些发现，我们建议 GLIS2是PC1功能下游效应子的候选者，这对于ADPKD中的囊肿进展至关重要。在 这项研究，我们将确定GLIS2的体内作用机理，并确定其对囊肿细胞的影响 由于PKD2引起的增殖，凋亡和ADPKD。我们将确定GLIS2是否是通过 遗传和药物治疗研究。我们将评估体内基因型是否依赖 我们确定的转录变化类似地扩展到具有PKD突变体的细胞培养系统 基因型。我们还将评估PKD突变细胞系中GLIS2蛋白的功能特性。在汇总中， 这些研究为多囊信号传导和ADPKD发病机理打开了新的研究方向。