Identification of RNAs Engaged in Direct RNA-RNA Interaction with a Long Non-Coding RNA

Audrey Jacq (1) , Denis Becquet (1) , Séverine Guillen (1) , Bénédicte Boyer (1) , Maria-Montserrat Bello-Goutierrez (1) , Jean-Louis Franc (1) , Anne-Marie François-Bellan (1)

1 INP - Institut de neurophysiopathologie

The growing role attributed nowadays to long non-coding RNAs (lncRNA) in physiology and pathophysiology makes it crucial to characterize their interactome by identifying their molecular partners, DNA, proteins and/or RNAs. The latter can interact with lncRNA through networks involving proteins, but they can also be engaged in direct RNA/RNA interactions. We, therefore, developed an easy-to-use RNA pull-down procedure that allowed identification of RNAs engaged in direct RNA/RNA interaction with a lncRNA using psoralen, a molecule that cross-links only RNA/RNA interactions. Bioinformatics modeling of the lncRNA secondary structure allowed the selection of several specific antisense DNA oligonucleotide probes with a strong affinity for regions displaying a low probability of internal base pairing. Since the specific probes that were designed targeted accessible regions throughout the length of the lncRNA, the RNA-interaction zones could be delineated in the sequence of the lncRNA. When coupled with a high throughput RNA sequencing, this protocol can be used for the whole direct RNA interactome studies of a lncRNA of interest.

Domaines: Neurobiologie

Audrey Jacq, Denis Becquet, Séverine Guillen, Bénédicte Boyer, Maria-Montserrat Bello-Goutierrez, et al.. Identification of RNAs Engaged in Direct RNA-RNA Interaction with a Long Non-Coding RNA. Journal of visualized experiments : JoVE, 2021, 173, ⟨10.3791/62475⟩. ⟨hal-03375987⟩