Global identification of mRNA-interacting circular RNAs by CLiPPR-Seq.
Nucleic Acids Res
; 52(6): e29, 2024 Apr 12.
Article
em En
| MEDLINE
| ID: mdl-38324478
ABSTRACT
Although the functional role of circular RNA (circRNA) interaction with microRNAs and proteins has been studied extensively, circRNA interactions with the protein-coding mRNAs in intact cells remain largely unknown. Here, by employing AMT-mediated proximity ligation of RNA-RNA duplexes followed by circRNA enrichment and deep sequencing, we report a novel Cross-Linking Poly(A) Pulldown RNase R Sequencing (CLiPPR-seq) technology which identified hundreds of mRNA-interacting circRNAs in three different cell types, including ßTC6, C2C12 and HeLa cells. Furthermore, CLiPP-seq without RNase R treatment was also performed to identify the mRNA expression in these cells. BLAST analysis of circRNAs in CLiPPR-seq sample with the mRNAs in CLiPP-seq samples determined their potential complementary sequences for circRNA-mRNA interaction. Pulldown of circRNAs and poly(A) RNAs confirmed the direct interaction of circRNAs with target mRNAs. Silencing of mRNA-interacting circRNAs led to the altered expression of target mRNAs in ßTC6 cells, suggesting the role of direct interaction of circRNAs with mRNAs in gene expression regulation. CLiPPR-seq thus represents a novel method for illuminating the myriad of uncharacterized circRNA-mRNA hybrids that may regulate gene expression.
Texto completo:
1
Coleções:
01-internacional
Base de dados:
MEDLINE
Assunto principal:
Técnicas Genéticas
/
Análise de Sequência de RNA
/
MicroRNAs
/
RNA Circular
Tipo de estudo:
Diagnostic_studies
Limite:
Humans
Idioma:
En
Ano de publicação:
2024
Tipo de documento:
Article