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Transcriptome-wide effects of cytosine and adenine deaminases on gene expression induced by base editors

ABSTRACT: Transcriptome-wide effects of cytosine and adenine deaminases on gene expression induced by base editors

PROVIDER: PRJNA666201 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR12734330_1.fastq.gz	Fastqsanger.gz
	SRR12734330_2.fastq.gz	Fastqsanger.gz
	SRR12734331_1.fastq.gz	Fastqsanger.gz
	SRR12734331_2.fastq.gz	Fastqsanger.gz
	SRR12734332_1.fastq.gz	Fastqsanger.gz

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Transcriptome-wide effects of cytosine and adenine deaminases on gene expression induced by base editors

Project description:We used RNA-sequencing data to explore the transcriptome-wide effects of cytosine and adenine deaminases on gene expression induced by base editors.

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Project description:CRISPR-guided DNA base editors enable the efficient installation of targeted single-nucleotide changes. Cytosine or adenine base editors (CBEs or ABEs), which are fusions of cytidine or adenosine deaminases to CRISPR-Cas nickases, can efficiently induce DNA C-to-T or A-to-G alterations in DNA, respectively. We recently demonstrated that both the widely used CBE BE3 (harboring a rat APOBEC1 cytidine deaminase) and the optimized ABEmax editor can induce tens of thousands of guide RNA-independent, transcriptome-wide RNA base edits in human cells with high efficiencies. In addition, we showed the feasibility of creating SElective Curbing of Unwanted RNA Editing (SECURE)-BE3 variants that exhibit substantially reduced unwanted RNA editing activities while retaining robust and more precise on-target DNA editing. Here we describe structure-guided engineering of SECURE-ABE variants that not only possess reduced off-target RNA editing with comparable on-target DNA activities but are also the smallest Streptococcus pyogenes Cas9 (SpCas9) base editors described to date. In addition, we tested CBEs composed of cytidine deaminases other than APOBEC1 and found that human APOBEC3A (hA3A) cytidine deaminase CBE induces substantial transcriptome-wide RNA base edits with high efficiencies. By contrast, a previously described “enhanced” A3A (eA3A) cytidine deaminase CBE or a human activation-induced cytidine deaminase (hAID) CBE induce substantially reduced or near background levels of RNA edits. In sum, our work describes broadly useful SECURE-ABE and -CBE base editors and reinforces the importance of minimizing RNA editing activities of DNA base editors for research and therapeutic applications.

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Project description:Adenine and cytosine base editors (ABEs and CBEs) represent a new genome editing technology that allows the programmable installation of A-to-G or C-to-T alterations on DNA. We engineered Streptococcus pyogenes Cas9-based adenine and cytosine base editor (SpACE) that enables efficient simultaneous introduction of A-to-G and C-to-T substitutions in the same base editing window on DNA.

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Replacing the SpCas9 HNH domain by deaminases generates compact base editors with an alternative targeting scope

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Project description:CRISPR adenine and cytosine base editors with reduced RNA off-target activities [ABE]

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