Capture of RNA G-quadruplex structures using an l-RNA aptamer.

Autor:	Lam SY; Department of Chemistry and State Key Laboratory of Marine Pollution, City University of Hong Kong Kowloon Tong Hong Kong SAR 999077 China., Umar MI; Department of Chemistry and State Key Laboratory of Marine Pollution, City University of Hong Kong Kowloon Tong Hong Kong SAR 999077 China.; RNA Molecular Biology Group, National Institute of Arthritis and Musculoskeletal and Skin Diseases (NIAMS), National Institutes of Health Bethesda MD USA., Zhao H; Department of Chemistry and State Key Laboratory of Marine Pollution, City University of Hong Kong Kowloon Tong Hong Kong SAR 999077 China., Zhao J; Department of Chemistry and State Key Laboratory of Marine Pollution, City University of Hong Kong Kowloon Tong Hong Kong SAR 999077 China., Kwok CK; Department of Chemistry and State Key Laboratory of Marine Pollution, City University of Hong Kong Kowloon Tong Hong Kong SAR 999077 China.; Shenzhen Research Institute of City University of Hong Kong Shenzhen China ckkwok42@cityu.edu.hk.
Jazyk:	angličtina
Zdroj:	RSC chemical biology [RSC Chem Biol] 2024 Aug 29. Date of Electronic Publication: 2024 Aug 29.
DOI:	10.1039/d4cb00161c
Abstrakt:	G-quadruplexes (dG4 and rG4) are nucleic acid secondary structures formed by the self-assembly of certain G-rich sequences, and they have distinctive chemical properties and play crucial roles in fundamental biological processes. Small molecule G4 ligands were shown to be crucial in characterizing G4s and understanding their functions. Nevertheless, concerns regarding the specificity of these synthetic ligands for further investigation of G4s, especially for rG4 isolation purposes, have been raised. In comparison to G4 ligands, we propose a novel magnetic bead-based pulldown assay that enables the selective capture of general rG4s using functionalized l-Apt.4-1c from both simple buffer and complex media, including total RNA and the cell lysate. We found that our l-RNA aptamer can pulldown general rG4s with a higher efficiency and specificity than the G4 small molecule ligand BioTASQ v.1 in the presence of non-target competitors, including dG4 and non-G4 structures. Our findings reveal that biotinylated l-aptamers can serve as effective molecular tools for the affinity-based enrichment of rG4 of interest using this new assay, which was also verified by quantitative reverse transcription-polymerase chain reaction (RT-qPCR) on endogenous transcripts. This work provides new and important insights into rG4 isolation using a functionalized l-aptamer, which can potentially be applied in a transcript-specific or transcriptome-wide manner in the future. Competing Interests: The authors declare that they have no conflicts of interest. (This journal is © The Royal Society of Chemistry.)
Databáze:	MEDLINE
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