Changes in nuclear and cytoplasmic microRNA distribution following hypoxic stress

Purpose: MicroRNAs have well characterized roles in cytoplasmic gene regulation by binding mRNA and inhibiting translation. However, besides this post-transcriptional gene silencing, miRNAs have also been implicated in transcriptional gene regulation and alternative splicing, events that are restricted to the cell nucleus. The nuclear functions of miRNAs are currently not understood, and there is a paucity of systematic studies of miRNA nuclear-cytoplasmic distribution and fewer still which have investigated this in the context of physiological conditions. Methods: Here we performed nuclear-cytoplasmic fractionation in a mouse endothelial cell line in hypoxia and characterized the localization of miRNAs using small RNA sequencing. Results: We show here that there is a broad population of mature miRNAs in the cell nucleus and that it is also altered upon exposure to hypoxia. Conclusions: Our results strongly imply that miRNAs have extensive regulatory functions in the nucleus and expand potential therapeutic use of small RNA molecules. C166 endothelial cells were treated with hypoxia (NT, 2h and 24h timepoints) and fractionated into nuclear and cytoplasmic samples that were used for sequencing. Samples were done in duplicates.

Purpose: 微小RNA（MicroRNAs，miRNAs）通过结合信使RNA（mRNA）并抑制翻译过程，在细胞质基因调控中发挥着已被充分阐明的功能。然而，除了这种转录后基因沉默机制外，miRNAs还被证实参与转录水平的基因调控与可变剪接——这类过程此前被认为仅局限于细胞核内。当前学界对miRNAs的核功能尚缺乏清晰认知，且针对miRNA核质分布的系统性研究较为匮乏，而在生理条件下开展此类研究的报道更是寥寥无几。
Methods: 本研究针对缺氧处理下的小鼠内皮细胞系开展核质分离实验，并通过小RNA测序（small RNA sequencing）技术表征miRNAs的定位情况。
Results: 本研究证实，细胞核内存在广泛分布的成熟miRNAs群体，且其丰度会在缺氧处理后发生改变。
Conclusions: 本研究结果有力表明，miRNAs在细胞核内具备广泛的调控功能，同时拓展了小分子RNA分子的潜在治疗应用场景。
本研究对C166内皮细胞进行缺氧处理（设置正常对照NT、2小时及24小时三个时间点），随后将其分离为细胞核与细胞质样本用于测序，所有样本均设置生物学重复。