Back-splice junctions

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Tags: research, RNA-seq

To Be or Not to Be: Circular RNAs or mRNAs From Circular DNAs?

https://www.frontiersin.org/journals/genetics/articles/10.3389/fgene.2019.00940/full

回剪接结 (Back-splice junctions) 是环状RNA (circRNAs) 的一个标志性特征,环状RNA是存在于真核细胞中的一种非编码RNA分子类型。与线性RNA不同,环状RNA形成一个共价闭合的连续环,意味着它们没有5'或3'端。这种环形配置主要是通过一种称为回剪接的过程创建的。

在典型的(规范的)剪接中,前体mRNA的外显子以线性方式连接,形成成熟的信使RNA(mRNA),然后可以翻译成蛋白质。然而,在回剪接中,一个位于外显子末尾的下游剪接供体(splice donor)被连接到一个位于外显子起始处的上游剪接受体(splice acceptor)。这个过程创建了一个“回剪接结”,这是区分环状RNA和线性RNA的独特特征。

回剪接结不在典型的线性RNA分子中发现;它们是环状RNA独有的,并被用作环状RNA形成的确定性证据。这些环状RNA在生物学研究中非常感兴趣,因为它们在基因调控中的潜在作用,它们在细胞中相比线性RNAs的稳定性,以及它们在各种疾病中的含义,包括癌症。回剪接结和环状RNAs的研究是RNA生物学中的一个增长领域。

Characterization of ALTO-encoding circular RNAs expressed by Merkel cell polyomavirus and trichodysplasia spinulosa (纺锤体毛发异常增生症) polyomavirus

https://journals.plos.org/plospathogens/article?id=10.1371/journal.ppat.1009582

Fig 1. Identification of Merkel cell polyomavirus circRNAs.

https://doi.org/10.1371/journal.ppat.1009582.g001

NC_010277.2: Merkel cell polyomavirus isolate R17b, complete genome
(A) 
(Top panel) Schematic representation of potential circRNAs identified by the vircircRNA for MCPyV. 
V-shaped lines above the map indicate forward splicing events. 
Elliptical arcs below the map indicate predicted backsplicing. 
(Bottom panel) Red rectangles indicate ALTO open reading frames. 
Bold numbers above the map indicate the first base of ALTO start and stop codons, italic numbers below the map indicate the positions of exon boundaries. 
(B) (Left panel) RT-PCR analysis of four VP-MCC cell lines (MKL1, MKL-2, MS-1 and WaGa) with and without RNase R treatment. 
(Middle panel) Sanger sequencing of inverse PCR products from WaGa cells confirmed the backsplice junction of the predicted circRNAs. 
(Right panel) Schematic of the two different circALTO forms, we termed circALTO1 and circALTO2 based on the product length.

* (C) qRT-PCR analysis for circALTOs from VP-MCC (MKL-1, MKL-2, MS1, and WaGa) and VN-MCC (MCC13, MCC26 and UISO) after RNase R treatment. ACTB served as the normalization control. Values are the mean of three technical replicates; bars represent standard deviation. 
(D) Northern blot of total RNA after with and without RNase R treatment from VP-MCC cell line WaGa and VN-MCC cell line UISO using an ALTO-specific probe. Arrow indicates RNase R resistant band consistent with circALTO2. 
(E) Endpoint PCR confirmed the presence of circALTO2 in patient MCC tumors (upper panel), but not a non-malignant skin control or VN-MCC cell lines. circHIPK3 served as a control for RNA integrity (bottom panel).

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