biostudies-literatureUnknownNuccio SPBiotechnology and Biological Sciences Research CouncilThe development of selective ligands to target DNA G-quadruplexes (G4s) and i-motifs (iMs) has revealed their relevance in transcriptional regulation. However, most of these ligands are unable to target individual G4s or iMs in the genome, severely limiting their scope. Herein, we describe a new Approach to Target Exact Nucleic Acid alternative structures (ATENA) that relies on the chemical conjugation of established G4 and iM ligands to a catalytically inactive Cas9 protein (dCas9), enabling their individual targeting in living cells. ATENA demonstrated that the selective targeting of the G4 present in the oncogene <i>c-MYC</i> leads to the suppression of transcripts regulated exclusively by one of its promoters (P1). Conversely, targeting the <i>c-MYC</i> iMs on the opposite strand leadsbioRxiv : the preprint server for biology2024.10.14.618195https://www.ebi.ac.uk/biostudies/studies/S-EPMC12330486biostudies-literatureChemically modified CRISPR-Cas9 enables targeting of individual G-quadruplex and i-motif structures, revealing ligand-dependent transcriptional perturbation.PMC12330486BB/R011605/1BB/W001616/1Sanchez-Cabanillas CMaher TELiu LSLer AJMagnani LSchneekloth JSNuccio SPGuneri DGalli SFlint GFullenkamp CRFan EZhu MAntonio MDCadoni EWaller ZNikoloudaki RfalseChemically modified CRISPR-Cas9 enables targeting of individual G-quadruplex and i-motif structures, revealing ligand-dependent transcriptional perturbation.The development of selective ligands to target DNA G-quadruplexes (G4s) and i-motifs (iMs) has revealed their relevance in transcriptional regulation. However, most of these ligands are unable to target individual G4s or iMs in the genome, severely limiting their scope. Herein, we describe a new Approach to Target Exact Nucleic Acid alternative structures (ATENA) that relies on the chemical conjugation of established G4 and iM ligands to a catalytically inactive Cas9 protein (dCas9), enabling their individual targeting in living cells. ATENA demonstrated that the selective targeting of the G4 present in the oncogene <i>c-MYC</i> leads to the suppression of transcripts regulated exclusively by one of its promoters (P1). Conversely, targeting the <i>c-MYC</i> iMs on the opposite strand leads2025-01-01T00:00:00Z2025 Jul2026-04-08T15:49:55.226Z2026-04-08T05:36:15.499ZS-EPMC123304864077750310.1101/2024.10.14.618195