Small-molecule affinity capture of DNA/RNA quadruplexes and their identification in vitro and in vivo through the G4RP protocol

Isaline Renard (1) , Michael Grandmougin (1) , Apolline Roux (1) , Sunny Y. Yang , Pauline Lejault (1) , Marc Pirrotta (1) , Judy M. Y. Wong , David Monchaud (1)

1 ICMUB - Institut de Chimie Moléculaire de l'Université de Bourgogne [Dijon]

Guanine-rich DNA and RNA sequences can fold into higher-order structures known as G-quadruplexes (or G4-DNA and G4-RNA, respectively). The prevalence of the G4 landscapes in the human genome, transcriptome and ncRNAome (non-coding RNA), collectively known as G4ome, is strongly suggestive of biological relevance at multiple levels (gene expression , replication). Small-molecules can be used to track G4s in living cells for the functional characterization of G4s in both normal and disease-associated changes in cell biology. Here, we describe biotinylated biomimetic ligands referred to as Bio-TASQ and their use as molecular tools that allow for isolating G4s through affinity pull-down protocols. We demonstrate the general applicability of the method by purifying biologically relevant G4s from nucleic acid mixtures in vitro and from human cells through the G4RP-RT-qPCR protocol. Overall, the results presented here represent a step towards the optimization of G4-RNAs identification, a key step in studying G4s in cell biology and human diseases.

Domaines: Biochimie, Biologie Moléculaire; Biologie moléculaire

Isaline Renard, Michael Grandmougin, Apolline Roux, Sunny Y. Yang, Pauline Lejault, et al.. Small-molecule affinity capture of DNA/RNA quadruplexes and their identification in vitro and in vivo through the G4RP protocol. Nucleic Acids Research, 2019, 47 (11), pp.5502-5510. ⟨10.1093/nar/gkz215⟩. ⟨hal-02121536v2⟩