Quantitative evaluation of all hexamers as exonic splicing elements

RNA剪接 小基因 生物 外显子 选择性拼接 遗传学 剪接 外显子剪接增强剂 背景(考古学) 计算生物学 核糖核酸 外显子跳跃 增强子 基因 基因表达 古生物学
作者
Shengdong Ke,Shulian Shang,Sergey Kalachikov,Irina Morozova,Lin Yu,James J. Russo,Jingyue Ju,Lawrence A. Chasin
出处
期刊:Genome Research [Cold Spring Harbor Laboratory Press]
卷期号:21 (8): 1360-1374 被引量:200
标识
DOI:10.1101/gr.119628.110
摘要

We describe a comprehensive quantitative measure of the splicing impact of a complete set of RNA 6-mer sequences by deep sequencing successfully spliced transcripts. All 4096 6-mers were substituted at five positions within two different internal exons in a 3-exon minigene, and millions of successfully spliced transcripts were sequenced after transfection of human cells. The results allowed the assignment of a relative splicing strength score to each mutant molecule. The effect of 6-mers on splicing often depended on their location; much of this context effect could be ascribed to the creation of different overlapping sequences at each site. Taking these overlaps into account, the splicing effect of each 6-mer could be quantified, and 6-mers could be designated as enhancers (ESEseqs) and silencers (ESSseqs), with an ESRseq score indicating their strength. Some 6-mers exhibited positional bias relative to the two splice sites. The distribution and conservation of these ESRseqs in and around human exons supported their classification. Predicted RNA secondary structure effects were also seen: Effective enhancers, silencers and 3' splice sites tend to be single stranded, and effective 5' splice sites tend to be double stranded. 6-mers that may form positive or negative synergy with another were also identified. Chromatin structure may also influence the splicing enhancement observed, as a good correspondence was found between splicing performance and the predicted nucleosome occupancy scores of 6-mers. This approach may prove of general use in defining nucleic acid regulatory motifs, substitute for functional SELEX in most cases, and provide insights about splicing mechanisms.
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