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Knockout of circRNAs by base editing back-splice sites of circularized exons.

ABSTRACT: Many circular RNAs (circRNAs) are produced from back-splicing of exons of precursor mRNAs and are generally co-expressed with cognate linear RNAs. Methods for circRNA-specific knockout are lacking, largely due to sequence overlaps between forms. Here, we use base editors (BEs) for circRNA depletion. By targeting splice sites involved in both back-splicing and canonical splicing, BEs can repress circular and linear RNAs. Targeting sites predominantly for circRNA biogenesis, BEs could efficiently repress the production of circular but not linear RNAs. As hundreds of exons are predominantly back-spliced to produce circRNAs, this provides an efficient method to deplete circRNAs for functional study.

SUBMITTER: Gao X

PROVIDER: S-EPMC8744252 | biostudies-literature | 2022 Jan

REPOSITORIES: biostudies-literature

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Knockout of circRNAs by base editing back-splice sites of circularized exons.

Gao Xiang X Ma Xu-Kai XK Li Xiang X Li Guo-Wei GW Liu Chu-Xiao CX Zhang Jun J Wang Ying Y Wei Jia J Chen Jia J Chen Ling-Ling LL Yang Li L

Genome biology 20220110 1

Many circular RNAs (circRNAs) are produced from back-splicing of exons of precursor mRNAs and are generally co-expressed with cognate linear RNAs. Methods for circRNA-specific knockout are lacking, largely due to sequence overlaps between forms. Here, we use base editors (BEs) for circRNA depletion. By targeting splice sites involved in both back-splicing and canonical splicing, BEs can repress circular and linear RNAs. Targeting sites predominantly for circRNA biogenesis, BEs could efficiently ...[more]

PMID: 35012611

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Knockout of circRNAs by base editing back-splice sites of circularized exons

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Knockout of circRNAs by base editing back-splice sites of circularized exons

Project description:Knockout of circRNAs by base editing back-splice sites of circularized exons

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Cardiac circRNAs arise mainly from constitutive exons rather than alternatively spliced exons.

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Project description:Accurate pre-mRNA splicing is crucial for gene expression. The 5' splice site (5' ss)--the highly diverse element at the 5' end of introns--is initially recognized via base-pairing to the 5' end of the U1 small nuclear RNA (snRNA). However, many natural 5' ss have a poor match to the consensus sequence, and are predicted to be weak. Using genetic suppression experiments in human cells, we demonstrate that some atypical 5' ss are actually efficiently recognized by U1, in an alternative base-pairing register that is shifted by one nucleotide. These atypical 5' ss are phylogenetically widespread, and many of them are conserved. Moreover, shifted base-pairing provides an explanation for the effect of a 5' ss mutation associated with pontocerebellar hypoplasia. The unexpected flexibility in 5' ss-U1 base-pairing challenges an established paradigm and has broad implications for splice-site prediction algorithms and gene-annotation efforts in genome projects.

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