Journal
NUCLEIC ACIDS RESEARCH
Volume 50, Issue 15, Pages 8834-8851Publisher
OXFORD UNIV PRESS
DOI: 10.1093/nar/gkac663
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Funding
- Deutsche Forschungsgemeinschaft (DFG) [SCHA 909/4-1]
- Jurgen Manchot Stiftung, Dusseldorf
- Stiftung fur AIDS-Forschung, Dusseldorf
- Forschungskommission of the Medical Faculty, Heinrich Heine Universitat Dusseldorf [2020-12]
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This study demonstrates the design and experimental validation of sequences with specific splicing regulatory properties, and develops a predictive model for 5' ss usage based on RNA-seq data, which contributes to splice site recognition.
Correct pre-mRNA processing in higher eukaryotes vastly depends on splice site recognition. Beyond conserved 5 ' ss and 3 ' ss motifs, splicing regulatory elements (SREs) play a pivotal role in this recognition process. Here, we present in silico designed sequences with arbitrary a priori prescribed splicing regulatory HEXplorer properties that can be concatenated to arbitrary length without changing their regulatory properties. We experimentally validated in silico predictions in a massively parallel splicing reporter assay on more than 3000 sequences and exemplarily identified some SRE binding proteins. Aiming at a unified 'functional splice site strength' encompassing both U1 snRNA complementarity and impact from neighboring SREs, we developed a novel RNA-seq based 5 ' ss usage landscape, mapping the competition of pairs of high confidence 5 ' ss and neighboring exonic GT sites along HBond and HEXplorer score coordinate axes on human fibroblast and endothelium transcriptome datasets. These RNA-seq data served as basis for a logistic 5 ' ss usage prediction model, which greatly improved discrimination between strong but unused exonic GT sites and annotated highly used 5 ' ss. Our 5 ' ss usage landscape offers a unified view on 5 ' ss and SRE neighborhood impact on splice site recognition, and may contribute to improved mutation assessment in human genetics.
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