Mirzadeh Azad Fatemeh, Taheri Bajgan Elham, Naeli Parisa, Rudov Alexander, Bagheri Moghadam Mahrokh, Sadat Akhtar Mozhgan, Gholipour Akram, Mowla Seyed Javad, Malakootian Mahshid
Molecular Genetics Department, Faculty of Biological Sciences, Tarbiat Modares University, Tehran, Iran.
Patrick G. Johnston Centre for Cancer Research, Queen's University Belfast, Belfast, UK.
Cell J. 2022 Oct 1;24(10):569-576. doi: 10.22074/cellj.2022.8205.
The human large intergenic non-coding RNA-regulator of reprogramming program () is known as a stem cell specific linc-RNA. linc-ROR counteracts differentiation via sequestering microRNA-145 (miR-145) that targets OCT4 transcript. Despite the research on the expression and function, the exact structure of transcripts is not clear. Considering the contribution of alternative splicing in transcripts structures and function, identifying different spliced variants of is necessary for further functional analyses. We aimed to find the alternatively spliced transcripts of and investigate their expression pattern in stem and cancer cell lines and during neural differentiation of NT2 cells as a model for understanding linc-ROR role in stem cell and differentiation.
In this experimental study, locus was scanned for identifying novel exons. Different primer sets were used to detect new spliced variants by reverse transcription polymerase chain reaction (RT-PCR) and direct sequencing. Quantitative PCR (qPCR) and RT-PCR were employed to profile expression of transcripts in different cell lines and during neural differentiation of stem cells.
We could discover 13 novel spliced variants of linc-ROR harboring unique array of exons. Our work uncovered six novel exons, some of which were the product of exonized transposable elements. Monitoring expression profile of the spliced variants in a panel of pluripotent and non-pluripotent cells exhibited that all transcripts were primarily expressed in pluripotent cells. Moreover, the examined spliced variants showed a similar downregulation during neural differentiation of NT2 cells.
Altogether, our data showed despite the difference in the structure and composition of exons, various spliced variants of showed similar expression pattern in stem cells and through differentiation.
人类重编程程序的大型基因间非编码RNA调节因子（linc-ROR）是一种干细胞特异性长链非编码RNA。linc-ROR通过隔离靶向OCT4转录本的微小RNA-145（miR-145）来对抗分化。尽管对其表达和功能进行了研究，但其转录本的确切结构尚不清楚。考虑到可变剪接在转录本结构和功能中的作用，鉴定linc-ROR的不同剪接变体对于进一步的功能分析是必要的。我们旨在寻找linc-ROR的可变剪接转录本，并研究它们在干细胞和癌细胞系以及作为理解linc-ROR在干细胞和分化中作用模型的NT2细胞神经分化过程中的表达模式。
在本实验研究中，扫描linc-ROR基因座以鉴定新外显子。使用不同的引物组通过逆转录聚合酶链反应（RT-PCR）和直接测序检测新的剪接变体。采用定量PCR（qPCR）和RT-PCR分析linc-ROR转录本在不同细胞系以及干细胞神经分化过程中的表达情况。
我们发现了13种具有独特外显子阵列的linc-ROR新剪接变体。我们的研究发现了6个新外显子，其中一些是外显子化转座元件的产物。监测一组多能和非多能细胞中linc-ROR剪接变体的表达谱显示，所有转录本主要在多能细胞中表达。此外，所检测的linc-ROR剪接变体在NT2细胞神经分化过程中表现出类似的下调。
总之，我们的数据表明，尽管外显子的结构和组成存在差异，但linc-ROR的各种剪接变体在干细胞中以及分化过程中表现出相似的表达模式。
