转录因子HNF1β控制调节肾细胞结构和紧密连接完整性的转录网络。

IF 3.7 2区 医学 Q1 PHYSIOLOGY American Journal of Physiology-renal Physiology Pub Date : 2023-02-01 DOI:10.1152/ajprenal.00199.2022
Lotte E Tholen, Femke Latta, Joost H A Martens, Joost G J Hoenderop, Jeroen H F de Baaij
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引用次数: 0

摘要

肝细胞核因子(HNF)1β基因(HNF1B)突变导致常染色体显性小管间质性肾病,这是一种罕见的异质性疾病,其特征为肾囊肿和/或畸形,年轻人的成熟型糖尿病,低镁血症和低钾血症。电解质紊乱可能发生在肾元远端,这对Mg2+和Ca2+重吸收的微调是重要的。因此,我们旨在研究HNF1β在肾元远端部分的转录网络。我们结合HNF1β染色质免疫沉淀测序和mRNA表达数据来确定肾远曲小管细胞系(mpkDCT)中HNF1β的直接靶点。基因本体术语通路分析证明了数据集中细胞极性、细胞-细胞连接和细胞骨架通路的富集。在这些通路中直接或间接受HNF1β调控的基因包括顶端和底侧极性复合物的成员,包括碎屑蛋白同源物3 (Crb3)、分配缺陷6同源物-β (Pard6b)和LLGL Scribble细胞极性复合物组分2 (Llgl2)。在表达显性阴性Hnf1b的小鼠内髓集管3细胞的单层中,与野生型细胞相比,通过上皮电阻值降低和荧光素通透性增加(0.4 kDa)可以观察到,紧密连接的完整性受到损害。显性阴性Hnf1b的表达也导致生长在膜上的细胞高度下降(30%),表面增加(58.5%)。此外,与野生型球体相比,表达显性阴性Hnf1b的细胞形成的三维球体的大小减少了30%。总之,这些发现表明,HNF1β在肾元远端指导一个调节紧密连接完整性和细胞结构的转录网络。转录因子肝细胞核因子(HNF)1β的遗传缺陷导致以电解质紊乱、肾囊肿和糖尿病为特征的异质性疾病。通过结合rna测序和HNF1β染色质免疫沉淀测序数据,我们发现了新的富集于细胞极性通路的HNF1β靶点。新发现的靶标包括极性配合物Crb3、Pard6b和Llgl2。肾上皮细胞的功能分析显示,Hnf1b突变细胞的紧密连接完整性降低,典型的立方体形态丧失。
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Transcription factor HNF1β controls a transcriptional network regulating kidney cell structure and tight junction integrity.

Mutations in the hepatocyte nuclear factor (HNF)1β gene (HNF1B) cause autosomal dominant tubulointerstitial kidney disease, a rare and heterogeneous disease characterized by renal cysts and/or malformation, maturity-onset diabetes of the young, hypomagnesemia, and hypokalemia. The electrolyte disturbances may develop in the distal part of the nephron, which is important for fine-tuning of Mg2+ and Ca2+ reabsorption. Therefore, we aimed to study the transcriptional network directed by HNF1β in the distal part of the nephron. We combined HNF1β chromatin immunoprecipitation-sequencing and mRNA expression data to identify direct targets of HNF1β in a renal distal convoluted tubule cell line (mpkDCT). Gene Ontology term pathway analysis demonstrated enrichment of cell polarity, cell-cell junction, and cytoskeleton pathways in the dataset. Genes directly and indirectly regulated by HNF1β within these pathways included members of the apical and basolateral polarity complexes including Crumbs protein homolog 3 (Crb3), partitioning defective 6 homolog-β (Pard6b), and LLGL Scribble cell polarity complex component 2 (Llgl2). In monolayers of mouse inner medullary collecting duct 3 cells expressing dominant negative Hnf1b, tight junction integrity was compromised, as observed by reduced transepithelial electrical resistance values and increased permeability for fluorescein (0.4 kDa) compared with wild-type cells. Expression of dominant negative Hnf1b also led to a decrease in height (30%) and an increase in surface (58.5%) of cells grown on membranes. Moreover, three-dimensional spheroids formed by cells expressing dominant negative Hnf1b were reduced in size compared with wild-type spheroids (30%). Together, these findings demonstrate that HNF1β directs a transcriptional network regulating tight junction integrity and cell structure in the distal part of the nephron.NEW & NOTEWORTHY Genetic defects in transcription factor hepatocyte nuclear factor (HNF)1β cause a heterogeneous disease characterized by electrolyte disturbances, kidney cysts, and diabetes. By combining RNA-sequencing and HNF1β chromatin immunoprecipitation-sequencing data, we identified new HNF1β targets that were enriched for cell polarity pathways. Newly discovered targets included members of polarity complexes Crb3, Pard6b, and Llgl2. Functional assays in kidney epithelial cells demonstrated decreased tight junction integrity and a loss of typical cuboidal morphology in mutant Hnf1b cells.

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来源期刊
CiteScore
8.40
自引率
7.10%
发文量
154
审稿时长
2-4 weeks
期刊介绍: The American Journal of Physiology - Renal Physiology publishes original manuscripts on timely topics in both basic science and clinical research. Published articles address a broad range of subjects relating to the kidney and urinary tract, and may involve human or animal models, individual cell types, and isolated membrane systems. Also covered are the pathophysiological basis of renal disease processes, regulation of body fluids, and clinical research that provides mechanistic insights. Studies of renal function may be conducted using a wide range of approaches, such as biochemistry, immunology, genetics, mathematical modeling, molecular biology, as well as physiological and clinical methodologies.
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