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GFP报告系统揭示了Aquaporin-2表达的细胞间变异性

Lihe Chen¹, Adrian Murillo-de-Ozores¹, Euijung Park¹

¹Epithelial Systems Biology Laboratory, Systems Biology Center, National Heart, Lung, and Blood Institute, NIH, Bethesda, Maryland.

bioRxiv : the preprint server for biology

|December 25, 2025

概括

血管压素 (AVP) 调节水素-2 (Aqp2) 基因转录. 一个新的记者细胞系显示了与细胞周期状态相关的Aqp2表达异质性,为研究AVP调节提供了一个工具.

科学领域:

分子生物学分子生物学
细胞生物学细胞生物学
脏生理学脏生理学

背景情况:

血管压素 (AVP) 是脏中水的再吸收的关键调节剂.
水素-2 (Aqp2) 水道基因转录是收集管道主要细胞中的AVP作用的主要目标.
了解Aqp2基因调节对于了解脏水平衡至关重要.

研究的目的:

为研究Aqp2基因转录调节开发一个记者细胞系.
调查Aqp2表达的异质性背后的机制.
探索细胞周期和Aqp2表达之间的关系.

主要方法:

在mpkCCD细胞中使用CRISPR/Cas9设计了一个Aqp2记者细胞系,在内源Aqp2位点插入GFP磁带.
用dDAVP (血压素类似物) 和福斯科林刺激细胞,通过流细胞计评估GFP和Aqp2的表达.
进行了GFP阳性和GFP阴性群体的细胞分类,随后进行了再刺激,细胞循环分析 (流细胞计) 和RNA测序.

主要成果:

在dDAVP刺激和撤销时,Aqp2-GFP记者细胞系忠实地跟踪了内源的Aqp2表达.
在Aqp2表达中观察到显著的异质性,即使在刺激后,也只有不到50%的细胞表达GFP/Aqp2.
当GFP阴性和GFP阳性细胞重新生长时,它们产生了两种种群,这表明了动态调节.
细胞周期分析显示,GFP阳性细胞主要存在于G0/G1中,而GFP阴性细胞在G2/S中显示出更多的细胞.
RNA-seq在GFP阴性细胞中发现了细胞循环相关的转录的增加.

结论:

Aqp2基因表达的异质性与收集管道主要细胞的细胞周期状态密切相关.
开发的Aqp2-GFP记者细胞系是剖析Aqp2.2的转录调节的一个有价值的工具.
这一发现为脏水道表达的动态调节提供了新的见解.

关键词:

在Aqp2中,Aqp2是Aqp2.全国粮食计划署的记者在RNA-seqqq.细胞循环中的细胞循环.

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相关概念视频

Reporter Genes

Reporter Genes

Reporter genes are a type of protein-coding gene that are often tagged to a gene of interest. Once inside a target cell, reporter genes usually produce visually identifiable characteristics like fluorescence and luminescence when expressed along with the gene of interest. Thus, reporter genes “report” the presence or absence of genes of interest in an organism, determine the gene expression pattern, or track the physical location of a DNA segment or protein in the cell.

Aquaporins

Aquaporins

Aquaporins or AQPs are a family of integral membrane proteins whose primary function is to transport water, while some called aquaglyceroporins also transport glycerol. In addition, aquaporins have also been suspected to be involved in transporting volatile substances, such as carbon dioxide and ammonia, across membranes. Such AQPs that act as gas channels are often highly expressed in cells involved in the gaseous exchange, such as red blood cells, epithelial cells, and pulmonary capillaries.